G2Cdb::Gene report

Gene id
G00002468
Gene symbol
RPSA (HGNC)
Species
Homo sapiens
Description
ribosomal protein SA
Orthologue
G00001219 (Mus musculus)

Databases (7)

Gene
ENSG00000168028 (Ensembl human gene)
3921 (Entrez Gene)
905 (G2Cdb plasticity & disease)
RPSA (GeneCards)
Literature
150370 (OMIM)
Marker Symbol
HGNC:6502 (HGNC)
Protein Sequence
P08865 (UniProt)

Synonyms (4)

  • 37LRP
  • LRP
  • SA
  • p40

Literature (58)

Pubmed - other

  • C-terminal fragment of human laminin-binding protein contains a receptor domain for venezuelan equine encephalitis and tick-borne encephalitis viruses.

    Malygin AA, Bondarenko EI, Ivanisenko VA, Protopopova EV, Karpova GG and Loktev VB

    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, 630090, Russia.

    Polyclonal and monoclonal antibodies (MABs) to human laminin-binding protein (LBP) can efficiently block the penetration of some alpha- and flaviviruses into the cell. A panel of 13 types of MABs to human recombinant LBP was used for more detailed study of the mechanism of this process. Competitive analysis has shown that MABs to LBP can be divided into six different competition groups. MABs 4F6 and 8E4 classified under competition groups 3 and 4 can inhibit the replication of Venezuelan equine encephalitis virus (VEEV), which is indicative of their interaction with the receptor domain of LBP providing for binding with virions. According to enzyme immunoassay and immunoblotting data, polyclonal anti-idiotypic antibodies to MABs 4F6 and 8E4 modeling paratopes of the LBP receptor domain can specifically interact with VEEV E2 protein and tick-borne encephalitis virus (TBEV) E protein. Mapping of binding sites of MABs 4F6 and 8E4 with LBP by constructing short deletion fragments of the human LBP molecule has shown that MAB 8E4 interacts with the fragment of amino acid residues 187-210, and MAB 4F6 interacts with the fragment of residues 263-278 of LBP protein, which is represented by two TEDWS peptides separated by four amino acid residues. This suggested that the LBP receptor domain interacting with VEEV E2 and TBEV E viral proteins is located at the C-terminal fragment of the LBP molecule. A model of the spatial structure of the LBP receptor domain distally limited by four linear loops (two of which are represented by experimentally mapped regions of amino acid residues 187-210 and 263-278) as well as the central beta-folded region turning into the alpha-helical site including residues 200-216 of the LBP molecule and providing for the interaction with the laminin-1 molecule has been proposed.

    Biochemistry. Biokhimiia 2009;74;12;1328-36

  • Interactions of the 67 kDa laminin receptor and its precursor with laminin.

    Fatehullah A, Doherty C, Pivato G, Allen G, Devine L, Nelson J and Timson DJ

    School of Biological Sciences, Queen's University Belfast, Medical Biology Centre, 97 Lisburn Road, Belfast BT9 7BL, UK.

    The 67LR (67 kDa laminin receptor) enables cells to interact with components of the extracellular matrix. The molecule is derived from the 37LRP (37 kDa laminin receptor precursor); however, the precise molecular mechanism of this conversion is unknown. Recombinant 37LRP, expressed in and purified from Escherichia coli, bound to human laminin in a SPR (surface plasmon resonance) experiment. 67LR isolated from human breast-cancer-derived cells in culture was also shown to bind to laminin by SPR. However, the kinetics of association are qualitatively different. 37LRP, but not 67LR, binds to heparan sulfate. The binding of 37LRP to heparan sulfate did not affect the interaction of 37LRP with laminin. In contrast, heparan sulfate reduces the extent of binding of laminin to 67LR. Taken together, these results show that 37LRP has some of the biological activities of 67LR, even prior to the conversion event. However, the conversion affects the sites of interaction with both laminin and heparan sulfate.

    Bioscience reports 2009;30;2;73-9

  • Down-regulation of 67LR reduces the migratory activity of human glioma cells in vitro.

    Chen FX, Qian YR, Duan YH, Ren WW, Yang Y, Zhang CC, Qiu YM and Ji YH

    School of Life Sciences, Shanghai University, Shanghai 200444, PR China. chenfuxue@staff.shu.edu.cn

    Objectives: Glioma is the most common brain tumor in central nervous system. Traditional therapies are not effective to cure this disease. Experimental evidence indicates that the 67 kDa elastin-laminin receptor (67LR) subunit is a high-affinity non-integrin laminin-binding protein that is over-expressed on the tumor cell surface in a variety of human carcinomas, and directly correlates with a higher proliferation rate of malignant cells and tendency to metastasize. However, little is known of the expression and function of 67LR in glioma cells.

    Methods: In this study, we estimated whether 67LR was constitutively over-expressed in high-grade astrocytomas by immunohistochemical staining and Western blotting, and investigated the role of a low level of 67LR expression in glioma cell line-U251 by constructing an interfering RNA expression plasmid.

    Results: The results showed that the 67LR had an enhanced over-expression in high-grade astrocytomas against normal brain tissues samples, and that the migratory activity of glioma cells was reduced after the down-regulation of the 67LR gene by RNAi.

    Discussion: It was hypothesized that a low level of 67LR expression could reduce migratory activity of glioma cells, which further proved that 67LR played an important role in glioma invasion by mediating tumor cell functions leading to sarcomata. This study provided a new alternative to gene therapy for glioma treatment.

    Brain research bulletin 2009;79;6;402-8

  • Laminin receptor involvement in the anti-angiogenic activity of pigment epithelium-derived factor.

    Bernard A, Gao-Li J, Franco CA, Bouceba T, Huet A and Li Z

    Université Pierre et Marie Curie, Univerisité Paris 06, UR4, Aging, Stress and Inflammation and Institut Fédératif de Recherche 83, 75252 Paris, France.

    Pigment epithelium-derived factor (PEDF) is a multifunctional protein with neurotrophic, anti-oxidative, and anti-inflammatory properties. It is also one of the most potent endogenous inhibitors of angiogenesis, playing an important role in restricting tumor growth, invasion, and metastasis. Studies show that PEDF binds to cell surface proteins, but little is known about how it exerts its effects. Recently, research identified phospholipase A(2)/nutrin/patatin-like phospholipase domain-containing 2 as one PEDF receptor. To identify other receptors, we performed yeast two-hybrid screening using PEDF as bait and discovered that the non-integrin 37/67-kDa laminin receptor (LR) is another PEDF receptor. Co-immunoprecipitation, His tag pulldown, and surface plasmon resonance assays confirmed the interaction between PEDF and LR. Using the yeast two-hybrid method, we further restricted the LR-interacting domain on PEDF to a 34-amino acid (aa) peptide (aa 44-77) and the PEDF-interacting domain on LR to a 91-aa fragment (aa 120-210). A 25-mer peptide named P46 (aa 46-70), derived from 34-mer, interacts with LR in surface plasmon resonance assays and binds to endothelial cell (EC) membranes. This peptide induces EC apoptosis and inhibits EC migration, tube-like network formation in vitro, and retinal angiogenesis ex vivo, like PEDF. Our results suggest that LR is a real PEDF receptor that mediates PEDF angiogenesis inhibition.

    The Journal of biological chemistry 2009;284;16;10480-90

  • Hypoxia-mediated up-regulation of MGr1-Ag/37LRP in gastric cancers occurs via hypoxia-inducible-factor 1-dependent mechanism and contributes to drug resistance.

    Liu L, Sun L, Zhang H, Li Z, Ning X, Shi Y, Guo C, Han S, Wu K and Fan D

    State Key Laboratory of Cancer Biology, Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi'an, China.

    Our previous study demonstrated hypoxia-inducible factor-1(HIF-1) could prompt multidrug resistance (MDR) phenotype and MGr1-Ag/37LRP, a novel drug-resistance protein was reported by our labortary, associated with multidrug resistance in gastric cancer. Given this association, we hypothesized that MGr1-Ag/37LRP contributed to HIF-1-dependent hypoxia-induced MDR phenotype. Initial experiments revealed that blocking MGr1-Ag/37LRP expression by siRNA in gastric cancer cells effectively reversed multidrug resistance phenotype induced by hypoxia. Subsequent analysis of MGr1-Ag/37LRP mRNA and protein in gastric cancer cells revealed a time-dependent manner increase with hypoxia. While the up-regulation of MGr1-Ag/37LRP was abolished by HIF-1 inhibition with siRNA. Studies using luciferase promoter constructs revealed a significant increase in activity in cells subject to hypoxia and such hypoxia inducibility was lost in cells co-transfected siRNA targeting HIF-1. Analysis of the MGr1-Ag/37LRP promoter revealed several potential binding sites for HIF-1. Electrophoretic mobility shift assay and chromatin immunoprecipitation demonstrated a functional HIF-1 binding site within MGr1-Ag/37LRP gene regulatory sequence located at -16 to -11 relative to the transcriptional initiation point. These observations demonstrate that MGr1-Ag/37LRP is actively engaged by hypoxia and represent a novel HIF-1 target. Such results suggest hypoxia-elicited MGr1-Ag/37LRP expression as a pathway for resistance of gastric cancer to chemotherapeutics.

    International journal of cancer 2009;124;7;1707-15

  • 67-kDa laminin receptor in human bile duct carcinoma.

    Li D, Chen J, Gao Z, Li X, Yan X, Xiong Y and Wang S

    Hepatobiliary Surgery Institute, Southwest Hospital, Third Military Medical University, Chongqing, PR China.

    Abnormal interaction of epithelial cells with laminin component of basement membrane may account for altered biological behavior of cells, influencing proliferation, adhesion, and motility. In the current study, we investigated the role of 67-kDa laminin receptor (67LR), a high affinity receptor for laminin, in aggressiveness of bile duct carcinoma.

    Methods: Fifty-two paraffin-embedded specimens and 22 fresh tissues of patients with bile duct carcinoma were analyzed using immunohistologic and real-time polymerase chain reaction techniques, respectively. Expression of 67LR on the bile duct carcinoma QBC939 cells was examined by flow cytometry. The effects of 67LR on the adhesive and invasive abilities of QBC939 cells were determined by adhesion and invasion assay in vitro.

    Results: Both at the mRNA and protein level, bile duct carcinoma cells expressed a higher level of 67LR than normal epithelial cells (p < 0.01). The expression of 67LR was correlated inversely with differentiation extent of tumor (p < 0.05). The 67LR level was significantly increased in patients with lymph node metastases than in patients without lymph node involvement (p < 0.01). Flow cytometry showed that 69.9 +/- 1.1% of QBC939 cells expressed 67LR. Expression of 67LR increased the adhesion and invasion of QBC939 cells. 60 and 120 min of incubation of 67LR antibodies, MLuC5, induced 46.3 +/- 2.2 and 61.3 +/- 2.1% inhibition of adhesion, respectively. The invasive ability of QBC939 cells to Matrigel was also reduced by 67LR antibodies, MLuC5.

    Conclusions: Overexpressing 67LR on bile duct carcinoma was associated with invasion and metastasis of bile duct carcinoma. Blockage of 67LR suppressed adhesion and invasion of bile duct carcinoma.

    European surgical research. Europaische chirurgische Forschung. Recherches chirurgicales europeennes 2009;42;3;168-73

  • Occurrence of autoantibodies to annexin I, 14-3-3 theta and LAMR1 in prediagnostic lung cancer sera.

    Qiu J, Choi G, Li L, Wang H, Pitteri SJ, Pereira-Faca SR, Krasnoselsky AL, Randolph TW, Omenn GS, Edelstein C, Barnett MJ, Thornquist MD, Goodman GE, Brenner DE, Feng Z and Hanash SM

    Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA. jiqiu@fhcrc.org

    Purpose: We have implemented a high throughput platform for quantitative analysis of serum autoantibodies, which we have applied to lung cancer for discovery of novel antigens and for validation in prediagnostic sera of autoantibodies to antigens previously defined based on analysis of sera collected at the time of diagnosis.

    Proteins from human lung adenocarcinoma cell line A549 lysates were subjected to extensive fractionation. The resulting 1,824 fractions were spotted in duplicate on nitrocellulose-coated slides. The microarrays produced were used in a blinded validation study to determine whether annexin I, PGP9.5, and 14-3-3 theta antigens previously found to be targets of autoantibodies in newly diagnosed patients with lung cancer are associated with autoantibodies in sera collected at the presymptomatic stage and to determine whether additional antigens may be identified in prediagnostic sera. Individual sera collected from 85 patients within 1 year before a diagnosis of lung cancer and 85 matched controls from the Carotene and Retinol Efficacy Trial (CARET) cohort were hybridized to individual microarrays.

    Results: We present evidence for the occurrence in lung cancer sera of autoantibodies to annexin I, 14-3-3 theta, and a novel lung cancer antigen, LAMR1, which precede onset of symptoms and diagnosis.

    Conclusion: Our findings suggest potential utility of an approach to diagnosis of lung cancer before onset of symptoms that includes screening for autoantibodies to defined antigens.

    Journal of clinical oncology : official journal of the American Society of Clinical Oncology 2008;26;31;5060-6

  • Green tea polyphenol EGCG signaling pathway through the 67-kDa laminin receptor.

    Tachibana H

    tatibana@agr.kyushu-u.ac.jp

    Nihon yakurigaku zasshi. Folia pharmacologica Japonica 2008;132;3;145-9

  • E3 ubiquitin ligase SIAH1 mediates ubiquitination and degradation of TRB3.

    Zhou Y, Li L, Liu Q, Xing G, Kuai X, Sun J, Yin X, Wang J, Zhang L and He F

    State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, 27 Taiping Road, Beijing 100850, China.

    Tribbles 3 homolog (TRB3) is recently identified as a scaffold-like regulator of various signal transducers and has been implicated in several processes including insulin signaling, NF-kappaB signaling, lipid metabolism and BMP signaling. To further understand cellular mechanisms of TRB3 regulation, we performed a yeast two-hybrid screen to identify novel TRB3 interacting proteins and totally obtained ten in-frame fused preys. Candidate interactions were validated by co-immunoprecipitation assays in mammalian cells. We further characterized the identified proteins sorted by Gene Ontology Annotation. Its interaction with the E3 ubiquitin ligase SIAH1 was further investigated. SIAH1 could interact with TRB3 both in vitro and in vivo. Importantly, SIAH1 targeted TRB3 for proteasome-dependent degradation. Cotransfection of SIAH1 could withdraw up-regulation of TGF-beta signaling by TRB3, suggesting SIAH1-induced degradation of TRB3 represents a potential regulatory mechanism for TGF-beta signaling.

    Cellular signalling 2008;20;5;942-8

  • Green tea polyphenol epigallocatechin-3-gallate signaling pathway through 67-kDa laminin receptor.

    Umeda D, Yano S, Yamada K and Tachibana H

    Laboratory of Food Chemistry, Division of Applied Biological Chemistry, Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, Fukuoka 812-8581, Japan.

    (-)-Epigallocatechin-3-gallate (EGCG), the principal polyphenol in green tea, has been shown to be a potent chemopreventive agent. Recently, 67-kDa laminin receptor (67LR) has been identified as a cell surface receptor for EGCG that mediates the anticancer activity of EGCG. Indeed, expression of 67LR confers EGCG responsiveness to tumor cells; however, the molecular basis for the anticancer activity of EGCG in vivo is not entirely understood. Here we show that (i) using a direct genetic screen, eukaryotic translation elongation factor 1A (eEF1A) is identified as a component responsible for the anticancer activity of EGCG; (ii) through both eEF1A and 67LR, EGCG induces the dephosphorylation of myosin phosphatase targeting subunit 1 (MYPT1) at Thr-696 and activates myosin phosphatase; and (iii) silencing of 67LR, eEF1A, or MYPT1 in tumor cells results in abrogation of EGCG-induced tumor growth inhibition in vivo. Additionally, we found that eEF1A is up-regulated by EGCG through 67LR. Overall, these findings implicate both eEF1A and MYPT1 in EGCG signaling for cancer prevention through 67LR.

    The Journal of biological chemistry 2008;283;6;3050-8

  • The 67 kDa laminin receptor: structure, function and role in disease.

    Nelson J, McFerran NV, Pivato G, Chambers E, Doherty C, Steele D and Timson DJ

    School of Biological Sciences, Queen's University Belfast, Medical Biology Centre, 97 Lisburn Road, Belfast BT9 7BL, UK.

    The 67LR (67 kDa laminin receptor) is a cell-surface receptor with high affinity for its primary ligand. Its role as a laminin receptor makes it an important molecule both in cell adhesion to the basement membrane and in signalling transduction following this binding event. The protein also plays critical roles in the metastasis of tumour cells. Isolation of the protein from either normal or cancerous cells results in a product with an approx. molecular mass of 67 kDa. This protein is believed to be derived from a smaller precursor, the 37LRP (37 kDa laminin receptor precursor). However, the precise mechanism by which cytoplasmic 37LRP becomes cell-membrane-embedded 67LR is unclear. The process may involve post-translational fatty acylation of the protein combined with either homo- or hetero-dimerization, possibly with a galectin-3-epitope-containing partner. Furthermore, it has become clear that acting as a receptor for laminin is not the only function of this protein. 67LR also acts as a receptor for viruses, such as Sindbis virus and dengue virus, and is involved with internalization of the prion protein. Interestingly, unmodified 37LRP is a ribosomal component and homologues of this protein are found in all five kingdoms. In addition, it appears to be strongly associated with histones in the eukaryotic cell nucleus, although the precise role of these interactions is not clear. Here we review the current understanding of the structure and function of this molecule, as well as highlighting areas requiring further research.

    Bioscience reports 2008;28;1;33-48

  • Immunochemical and single molecule force spectroscopy studies of specific interaction between the laminin binding protein and the West Nile virus surface glycoprotein E domain II.

    Bogachek MV, Protopopova EV, Loktev VB, Zaitsev BN, Favre M, Sekatskii SK and Dietler G

    State Research Center of Virology and Biotechnology , Koltsovo, Novosibirsk Region 630559, Russia.

    ELISA and Western blot immunochemical data attest an effective and highly specific interaction of the surface glycoprotein E domain II (DII) of the tick born encephalitis and Dengue viruses with the laminin binding protein (LBP). Based on a highly conservative structure of the DII in different flaviviruses we propose a similarly effective interaction between the LBP and the DII of the surface glycoprotein E of the West Nile virus. We report the results of studies of this interaction by immunochemical and single molecule force spectroscopy methods. The specific binding between these species is confirmed by both methods.

    Journal of molecular recognition : JMR 2008;21;1;55-62

  • siRNA-mediated silencing of the 37/67-kDa high affinity laminin receptor in Hep3B cells induces apoptosis.

    Susantad T and Smith DR

    Molecular Pathology Laboratory, Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus, Salaya, Nakorn Pathom, Thailand.

    The laminin-binding protein, variously called the 37/67-kDa high affinity laminin receptor or p40, mediates the attachment of normal cells to the laminin network, and also has a role as a ribosomal protein. Over-expression of this protein has been strongly correlated with the metastatic phenotype. However, few studies have investigated the cellular consequence of the ablation of this gene's expression. To address this issue, the expression of the 37/67-kDa high affinity laminin receptor was knocked out with several siRNA constructs via RNA interference in transformed liver (Hep3B) cells. In each case where the message was specifically ablated, apoptosis was induced, as determined by annexin V/propidium iodide staining, and by double staining with annexin V and an antibody directed against the 37/67-kDa high affinity laminin receptor. These results suggest that this protein plays a critical role in maintaining cell viability.

    Cellular & molecular biology letters 2008;13;3;452-64

  • Laminin receptor 1: a novel protein interacting with human circadian clock protein, hPer1.

    Wang Y, Liu Y, Hu L, Lu F, Jiang Z, Wan C and Wang Z

    Health Ministry Key Laboratory of Chronobiology, West China Medical Center, Sichuan University, Chengdu, Sichuan 610041, China.

    The circadian clock is the central timing system that controls numerous physiologic processes. The current model of these oscillators is based on autoregulatory transcription and translation feedback loops of these circadian genes in which Period1 (Per1) gene occupies a central position. The laminin receptor 1 (Lamr1) and its precursor are expressed in most tissues and play important roles in several physiologic and pathologic processes, including cell differentiation, growth, migration and cancer invasion. The present study showed that Lamr1 was a novel protein that interacted with human circadian clock protein hPer1 by the yeast two-hybrid system and co-immunoprecipitation, which was expressed in many tissues and did not display circadian rhythm. The expression of hPer1 was knocked down to 84.9% by the hPer1 RNA interfering test, but the expression levels of Lamr1 was not depressed by the hPer1 RNA interfering test. The results suggest that Lamr1 is a novel protein that interacts with human circadian clock protein hPer1 and Lamr1 is not a direct efferent element of circadian clock.

    Neurological research 2007;29;5;429-34

  • The 37/67-kilodalton laminin receptor is a receptor for adeno-associated virus serotypes 8, 2, 3, and 9.

    Akache B, Grimm D, Pandey K, Yant SR, Xu H and Kay MA

    Stanford University, Department of Pediatrics, 300 Pasteur Drive, Room G305, Stanford, CA 94305-5208, USA.

    Adeno-associated virus serotype 8 (AAV8) is currently emerging as a powerful gene transfer vector, owing to its capability to efficiently transduce many different tissues in vivo. While this is believed to be in part due to its ability to uncoat more readily than other AAV serotypes such as AAV2, understanding all the processes behind AAV8 transduction is important for its application and optimal use in human gene therapy. Here, we provide the first report of a cellular receptor for AAV8, the 37/67-kDa laminin receptor (LamR). We document binding of LamR to AAV8 capsid proteins and intact virions in vitro and demonstrate its contribution to AAV8 transduction of cultured cells and mouse liver in vivo. We also show that LamR plays a role in transduction by three other closely related serotypes (AAV2, -3, and -9). Sequence and deletion analysis allowed us to map LamR binding to two protein subdomains predicted to be exposed on the AAV capsid exterior. Use of LamR, which is constitutively expressed in many clinically relevant tissues and is overexpressed in numerous cancers, provides a molecular explanation for AAV8's broad tissue tropism. Along with its robust transduction efficiency, our findings support the continued development of AAV8-based vectors for clinical applications in humans, especially for tumor gene therapy.

    Funded by: NHLBI NIH HHS: HL66948, U01 HL066948

    Journal of virology 2006;80;19;9831-6

  • The involvement of the 67 kDa laminin receptor-mediated modulation of cytoskeleton in the degranulation inhibition induced by epigallocatechin-3-O-gallate.

    Fujimura Y, Umeda D, Kiyohara Y, Sunada Y, Yamada K and Tachibana H

    Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.

    Recently, we have reported that (-)-epigallocatechin-3-O-gallate (EGCG) acts as an inhibitor of degranulation. However, the inhibitory mechanism for degranulation is still poorly understood. Here we show that suppression of exocytosis-related myosin II regulatory light chain phosphorylation and alteration of actin remodeling are involved in the inhibitory effect of EGCG on the calcium ionophore-induced degranulation from human basophilic KU812 cells. Surface plasmon resonance assay also revealed that EGCG binds to the cell surface, and the disruption of lipid rafts resulted in reduction of EGCG's ability. We have previously identified the raft-associated 67kDa laminin receptor (67LR) as an EGCG receptor on the cell surface. Treatment of the cells with anti-67LR antibody or RNA interference-mediated downregulation of 67LR expression abolished the effects of EGCG. These findings suggest that EGCG-induced inhibition of the degranulation includes the primary binding of EGCG to the cell surface 67LR and subsequent modulation of cytoskeleton.

    Biochemical and biophysical research communications 2006;348;2;524-31

  • Contribution of the 37-kDa laminin receptor precursor in the anti-metastatic PSP94-derived peptide PCK3145 cell surface binding.

    Annabi B, Currie JC, Bouzeghrane M, Dulude H, Daigneault L, Garde S, Rabbani SA, Panchal C, Wu JJ and Béliveau R

    Laboratoire d'Oncologie Moléculaire, Département de Chimie, Université du Québec à Montréal, Que., Canada.

    Purpose: PCK3145 is an anti-metastatic synthetic peptide with promising therapeutic efficacy against hormone-refractory prostate cancer. The characterization of the PCK3145 peptide cell surface binding/internalization mechanisms and of the receptors involved remained to be explored.

    Results: [(14)C]PCK3145 cell surface binding assays showed rapid and transient kinetic profile, that was inhibited by RGD peptides, laminin, hyaluronan, and type-I collagen. RGD peptides were however unable to inhibit PCK3145 intracellular uptake. Far-Western ligand binding studies enabled the identification of the 37-kDa laminin receptor precursor (37LRP) as a potential ligand for PCK3145. Overexpression of the recombinant 37LRP indeed led to an increase in PCK3145 binding but unexpectedly not to its uptake.

    Conclusions: Our data support the implication of laminin receptors in cell surface binding and in transducing PCK3145 anti-metastatic effects, and provide a rational for targeting cancers that express high levels of such laminin receptors.

    Biochemical and biophysical research communications 2006;346;1;358-66

  • Characterization of 67 kD laminin receptor, a protein whose gene is overexpressed on treatment of cells with anti-benzo[a]pyrene-7,8-diol-9,10-epoxide.

    An SJ, Chen JK, Chen HJ, Chang W, Jiang YG, Wei QY and Chen XM

    Department of Occupational and Environmental Health, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan Hubei 430030, China.

    The molecular mechanisms potentially related to tumorigenesis induced by anti-benzo[a]pyrene-7,8-diol-9,10-epoxide (anti-BPDE) were investigated by suppression subtractive hybridization of the human bronchial epithelial cells (16HBE) carcinoma induced by BPDE-transformed 16HBE cells (16HBE-C). The 67 kD laminin receptor gene (67LR1) is one of the screened overexpressed genes in 16HBE-C cells when compared with 16HBE. In order to understand the main functions of 67LR1 gene, we amplified the full length of 67LR1 gene using reverse transcription-polymerase chain reaction (RT-PCR) method. The amplified gene products were inserted into pcDNA 3.1 Directional TOPO expression vector. We then transfected 16HBE cells with this vector and derived stable transfected 16HBE cell lines containing the 67LR1 gene by using lipofectin and G418 selection protocols. The expression products of transfected genes were analyzed by semiquantitative RT-PCR. Soft agar growth assay was carried out to identify the malignant features of 67LR1 gene. The stable transfected cell lines can form colonies in soft agar. Further, the transfected cells showed morphological changes compared to the control cells, such as the obvious pseudopods. These data suggest that the 67LR1 gene may be related to malignant transformation induced by the anti-BPDE. The 67LR1 protein may be related to the directionality of cell movement.

    Toxicological sciences : an official journal of the Society of Toxicology 2006;90;2;326-30

  • Proteomic analysis of SUMO4 substrates in HEK293 cells under serum starvation-induced stress.

    Guo D, Han J, Adam BL, Colburn NH, Wang MH, Dong Z, Eizirik DL, She JX and Wang CY

    Center for Biotechnology and Genomic Medicine, Medical College of Georgia, 1120 15th Street, CA4098, Augusta, GA 30912, USA.

    The substrates of SUMO4, a novel member for the SUMO gene family, were characterized in HEK293 cells cultured under serum starvation by proteomic analysis. We identified 90 SUMO4 substrates including anti-stress proteins such as antioxidant enzymes and molecular chaperones or co-chaperones. The substrates also include proteins involved in the regulation of DNA repair and synthesis, RNA processing, protein degradation, and glucose metabolism. Several SUMO4-associated transcription factors were characterized by Western blot analyses. AP-1 was selected for in vitro conjugation assays to confirm SUMO4 sumoylation of these transcription factors. Further functional analyses of the transcription factors suggested that SUMO4 sumoylation represses AP-1 and AP-2alpha transcriptional activity, but enhances GR DNA binding capacity. These results demonstrate that SUMO4 sumoylation may play an important role in the regulation of intracellular stress.

    Biochemical and biophysical research communications 2005;337;4;1308-18

  • Towards a proteome-scale map of the human protein-protein interaction network.

    Rual JF, Venkatesan K, Hao T, Hirozane-Kishikawa T, Dricot A, Li N, Berriz GF, Gibbons FD, Dreze M, Ayivi-Guedehoussou N, Klitgord N, Simon C, Boxem M, Milstein S, Rosenberg J, Goldberg DS, Zhang LV, Wong SL, Franklin G, Li S, Albala JS, Lim J, Fraughton C, Llamosas E, Cevik S, Bex C, Lamesch P, Sikorski RS, Vandenhaute J, Zoghbi HY, Smolyar A, Bosak S, Sequerra R, Doucette-Stamm L, Cusick ME, Hill DE, Roth FP and Vidal M

    Center for Cancer Systems Biology and Department of Cancer Biology, Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Boston, Massachusetts 02115, USA.

    Systematic mapping of protein-protein interactions, or 'interactome' mapping, was initiated in model organisms, starting with defined biological processes and then expanding to the scale of the proteome. Although far from complete, such maps have revealed global topological and dynamic features of interactome networks that relate to known biological properties, suggesting that a human interactome map will provide insight into development and disease mechanisms at a systems level. Here we describe an initial version of a proteome-scale map of human binary protein-protein interactions. Using a stringent, high-throughput yeast two-hybrid system, we tested pairwise interactions among the products of approximately 8,100 currently available Gateway-cloned open reading frames and detected approximately 2,800 interactions. This data set, called CCSB-HI1, has a verification rate of approximately 78% as revealed by an independent co-affinity purification assay, and correlates significantly with other biological attributes. The CCSB-HI1 data set increases by approximately 70% the set of available binary interactions within the tested space and reveals more than 300 new connections to over 100 disease-associated proteins. This work represents an important step towards a systematic and comprehensive human interactome project.

    Funded by: NCI NIH HHS: R33 CA132073; NHGRI NIH HHS: P50 HG004233, R01 HG001715, RC4 HG006066, U01 HG001715; NHLBI NIH HHS: U01 HL098166

    Nature 2005;437;7062;1173-8

  • Proteomics of human umbilical vein endothelial cells applied to etoposide-induced apoptosis.

    Bruneel A, Labas V, Mailloux A, Sharma S, Royer N, Vinh J, Pernet P, Vaubourdolle M and Baudin B

    Service de Biochimie A, Hôpital Saint-Antoine, AP-HP, Paris, France. arnaud.bruneel@sat.ap-hop-paris.fr

    We have undertaken to continue the proteomic study of human umbilical vein endothelial cells (HUVECs) using the combination of 2-DE, automated trypsin digestion, and PMF analysis after MALDI-TOF MS and peptide sequencing using nano LC-ESI-MS/MS. The overall functional characterization of the 162 identified proteins from primary cultures of HUVECs confirms the metabolic capabilities of endothelium and illustrates various cellular functions more related to cell motility and angiogenesis, protein folding, anti-oxidant defenses, signal transduction, proteasome pathway and resistance to apoptosis. In comparison with controls cells, the differential proteomic analysis of HUVECs treated by the pro-apoptotic topoisomerase inhibitor etoposide further revealed the variation of eight proteins, namely, GRP78, GRP94, valosin-containing protein, proteinase inhibitor 9, cofilin, 37-kDa laminin receptor protein, bovine apolipoprotein, and tropomyosin. These data suggest that etoposide-induced apoptosis of human vascular endothelial cells results from the intricate involvement of multiple apoptosis processes including at least the mitochondrial and the ER stress pathways. The presented 2-D pattern and protein database, as well as the data related to apoptosis of HUVECs, are available at http://www.huvec.com.

    Proteomics 2005;5;15;3876-84

  • A human protein-protein interaction network: a resource for annotating the proteome.

    Stelzl U, Worm U, Lalowski M, Haenig C, Brembeck FH, Goehler H, Stroedicke M, Zenkner M, Schoenherr A, Koeppen S, Timm J, Mintzlaff S, Abraham C, Bock N, Kietzmann S, Goedde A, Toksöz E, Droege A, Krobitsch S, Korn B, Birchmeier W, Lehrach H and Wanker EE

    Max Delbrueck Center for Molecular Medicine, 13092 Berlin-Buch, Germany.

    Protein-protein interaction maps provide a valuable framework for a better understanding of the functional organization of the proteome. To detect interacting pairs of human proteins systematically, a protein matrix of 4456 baits and 5632 preys was screened by automated yeast two-hybrid (Y2H) interaction mating. We identified 3186 mostly novel interactions among 1705 proteins, resulting in a large, highly connected network. Independent pull-down and co-immunoprecipitation assays validated the overall quality of the Y2H interactions. Using topological and GO criteria, a scoring system was developed to define 911 high-confidence interactions among 401 proteins. Furthermore, the network was searched for interactions linking uncharacterized gene products and human disease proteins to regulatory cellular pathways. Two novel Axin-1 interactions were validated experimentally, characterizing ANP32A and CRMP1 as modulators of Wnt signaling. Systematic human protein interaction screens can lead to a more comprehensive understanding of protein function and cellular processes.

    Cell 2005;122;6;957-68

  • Laminin binding protein, 34/67 laminin receptor, carries stage-specific embryonic antigen-4 epitope defined by monoclonal antibody Raft.2.

    Katagiri YU, Kiyokawa N, Nakamura K, Takenouchi H, Taguchi T, Okita H, Umezawa A and Fujimoto J

    Department of Developmental Biology, National Research Institute for Child Health and Development, Setagaya-ku, Tokyo 157-8535, Japan. kata@nch.go.jp

    We previously produced monoclonal antibodies against the detergent-insoluble microdomain, i.e., the raft microdomain, of the human renal cancer cell line ACHN. Raft.2, one of these monoclonal antibodies, recognizes sialosyl globopentaosylceramide, which has the stage-specific embryonic antigen (SSEA)-4 epitope. Although the mouse embryonal carcinoma (EC) cell line F9 does not express SSEA-4, some F9 cells stained with Raft.2. Western analysis and matrix-assisted laser desorption ionization-time of flight mass spectrometry identified the Raft.2 binding molecule as laminin binding protein (LBP), i.e., 34/67 laminin receptor. Weak acid treatment or digestion with Clostridium perfringens sialidase reduced Raft.2 binding to LBP on nitrocellulose sheets and [(14)C]galactose was incorporated into LBP, indicating LBP to have a sialylated carbohydrate moiety. Subcellular localization analysis by sucrose density-gradient centrifugation and examination by confocal microscopy revealed LBP to be localized on the outer surface of the plasma membrane. An SSEA-4-positive human EC cell line, NCR-G3 cells, also expressed Raft.2-binding LBP.

    Biochemical and biophysical research communications 2005;332;4;1004-11

  • 67-kDa laminin receptor promotes internalization of cytotoxic necrotizing factor 1-expressing Escherichia coli K1 into human brain microvascular endothelial cells.

    Kim KJ, Chung JW and Kim KS

    Division of Pediatrics Infectious Diseases, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287, USA.

    Escherichia coli K1 is the most common Gram-negative organism causing meningitis, and its invasion of human brain microvascular endothelial cells (HBMEC) is a prerequisite for penetration into the central nervous system. We have reported previously that cytotoxic necrotizing factor 1 (CNF1) contributes to E. coli K1 invasion of HBMEC and interacts with 37-kDa laminin receptor precursor (37LRP) of HBMEC, which is a precursor of 67-kDa laminin receptor (67LR). In the present study, we examined the role of 67LR in the CNF1-expressing E. coli K1 invasion of HBMEC. Immunofluorescence microscopy and ligand overlay assays showed that 67LR is present on the HBMEC membrane and interacts with CNF1 protein as well as the CDPGYIGSR laminin peptide. 67LR was up-regulated and clustered at the sites of E. coli K1 on HBMEC in a CNF1-dependent manner. Pretreatment of CNF1+ E. coli K1 with recombinant 37-kDa laminin receptor precursor reduced the invasion rate to the level of Deltacnf1 mutant, and the invasion rate of CNF1+ E. coli K1 was enhanced in 67LR-overexpressing HBMEC, indicating 67LR is involved in the CNF1+ E. coli K1 invasion of HBMEC. Coimmunoprecipitation analysis showed that, upon incubation with CNF1+ E. coli K1 but not with Deltacnf1 mutant, focal adhesion kinase and paxillin were recruited and associated with 67LR. When immobilized onto polystyrene beads, CNF1 was sufficient to induce internalization of coupled beads into HBMEC through interaction with 67LR. Taken together, this is the first demonstration that E. coli K1 invasion of HBMEC occurs through the ligand-receptor (CNF1-67LR) interaction, and 67LR promotes CNF1-expressing E. coli K1 internalization of HBMEC.

    Funded by: NIAID NIH HHS: AI 47225; NINDS NIH HHS: NS 26310

    The Journal of biological chemistry 2005;280;2;1360-8

  • Nucleolar proteome dynamics.

    Andersen JS, Lam YW, Leung AK, Ong SE, Lyon CE, Lamond AI and Mann M

    Department of Biochemistry and Molecular Biology, Campusvej 55, DK-5230 Odense M, Denmark.

    The nucleolus is a key organelle that coordinates the synthesis and assembly of ribosomal subunits and forms in the nucleus around the repeated ribosomal gene clusters. Because the production of ribosomes is a major metabolic activity, the function of the nucleolus is tightly linked to cell growth and proliferation, and recent data suggest that the nucleolus also plays an important role in cell-cycle regulation, senescence and stress responses. Here, using mass-spectrometry-based organellar proteomics and stable isotope labelling, we perform a quantitative analysis of the proteome of human nucleoli. In vivo fluorescent imaging techniques are directly compared to endogenous protein changes measured by proteomics. We characterize the flux of 489 endogenous nucleolar proteins in response to three different metabolic inhibitors that each affect nucleolar morphology. Proteins that are stably associated, such as RNA polymerase I subunits and small nuclear ribonucleoprotein particle complexes, exit from or accumulate in the nucleolus with similar kinetics, whereas protein components of the large and small ribosomal subunits leave the nucleolus with markedly different kinetics. The data establish a quantitative proteomic approach for the temporal characterization of protein flux through cellular organelles and demonstrate that the nucleolar proteome changes significantly over time in response to changes in cellular growth conditions.

    Funded by: Wellcome Trust: 073980

    Nature 2005;433;7021;77-83

  • Immunoaffinity profiling of tyrosine phosphorylation in cancer cells.

    Rush J, Moritz A, Lee KA, Guo A, Goss VL, Spek EJ, Zhang H, Zha XM, Polakiewicz RD and Comb MJ

    Cell Signaling Technology Inc., 166B Cummings Center, Beverly, Massachusetts 01915, USA.

    Tyrosine kinases play a prominent role in human cancer, yet the oncogenic signaling pathways driving cell proliferation and survival have been difficult to identify, in part because of the complexity of the pathways and in part because of low cellular levels of tyrosine phosphorylation. In general, global phosphoproteomic approaches reveal small numbers of peptides containing phosphotyrosine. We have developed a strategy that emphasizes the phosphotyrosine component of the phosphoproteome and identifies large numbers of tyrosine phosphorylation sites. Peptides containing phosphotyrosine are isolated directly from protease-digested cellular protein extracts with a phosphotyrosine-specific antibody and are identified by tandem mass spectrometry. Applying this approach to several cell systems, including cancer cell lines, shows it can be used to identify activated protein kinases and their phosphorylated substrates without prior knowledge of the signaling networks that are activated, a first step in profiling normal and oncogenic signaling networks.

    Funded by: NCI NIH HHS: 1R43CA101106

    Nature biotechnology 2005;23;1;94-101

  • The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC).

    Gerhard DS, Wagner L, Feingold EA, Shenmen CM, Grouse LH, Schuler G, Klein SL, Old S, Rasooly R, Good P, Guyer M, Peck AM, Derge JG, Lipman D, Collins FS, Jang W, Sherry S, Feolo M, Misquitta L, Lee E, Rotmistrovsky K, Greenhut SF, Schaefer CF, Buetow K, Bonner TI, Haussler D, Kent J, Kiekhaus M, Furey T, Brent M, Prange C, Schreiber K, Shapiro N, Bhat NK, Hopkins RF, Hsie F, Driscoll T, Soares MB, Casavant TL, Scheetz TE, Brown-stein MJ, Usdin TB, Toshiyuki S, Carninci P, Piao Y, Dudekula DB, Ko MS, Kawakami K, Suzuki Y, Sugano S, Gruber CE, Smith MR, Simmons B, Moore T, Waterman R, Johnson SL, Ruan Y, Wei CL, Mathavan S, Gunaratne PH, Wu J, Garcia AM, Hulyk SW, Fuh E, Yuan Y, Sneed A, Kowis C, Hodgson A, Muzny DM, McPherson J, Gibbs RA, Fahey J, Helton E, Ketteman M, Madan A, Rodrigues S, Sanchez A, Whiting M, Madari A, Young AC, Wetherby KD, Granite SJ, Kwong PN, Brinkley CP, Pearson RL, Bouffard GG, Blakesly RW, Green ED, Dickson MC, Rodriguez AC, Grimwood J, Schmutz J, Myers RM, Butterfield YS, Griffith M, Griffith OL, Krzywinski MI, Liao N, Morin R, Morrin R, Palmquist D, Petrescu AS, Skalska U, Smailus DE, Stott JM, Schnerch A, Schein JE, Jones SJ, Holt RA, Baross A, Marra MA, Clifton S, Makowski KA, Bosak S, Malek J and MGC Project Team

    The National Institutes of Health's Mammalian Gene Collection (MGC) project was designed to generate and sequence a publicly accessible cDNA resource containing a complete open reading frame (ORF) for every human and mouse gene. The project initially used a random strategy to select clones from a large number of cDNA libraries from diverse tissues. Candidate clones were chosen based on 5'-EST sequences, and then fully sequenced to high accuracy and analyzed by algorithms developed for this project. Currently, more than 11,000 human and 10,000 mouse genes are represented in MGC by at least one clone with a full ORF. The random selection approach is now reaching a saturation point, and a transition to protocols targeted at the missing transcripts is now required to complete the mouse and human collections. Comparison of the sequence of the MGC clones to reference genome sequences reveals that most cDNA clones are of very high sequence quality, although it is likely that some cDNAs may carry missense variants as a consequence of experimental artifact, such as PCR, cloning, or reverse transcriptase errors. Recently, a rat cDNA component was added to the project, and ongoing frog (Xenopus) and zebrafish (Danio) cDNA projects were expanded to take advantage of the high-throughput MGC pipeline.

    Funded by: PHS HHS: N01-C0-12400

    Genome research 2004;14;10B;2121-7

  • Interaction of Doppel with the full-length laminin receptor precursor protein.

    Yin SM, Sy MS, Yang HY and Tien P

    Institute of Microbiology, Chinese Academy of Science, Beijing 100080, People's Republic of China.

    Doppel (Dpl) is a homolog of normal cellular prion protein (PrPc) with unknown functions. Ectopic expression of Dpl in the central nervous system (CNS) causes neurotoxicity and this effect is rescued by the expression of PrPc. However, the molecular basis for the protective effect of PrPc remains unclear. Using a yeast two-hybrid system, we showed that Dpl binds the full-length 37-kDa laminin receptor precursor protein (LRP), one of the receptors of PrPc. The interaction was also validated by immunoprecipitation and immunoblotting using transfected cell lines and in vivo derived tissues. Further mapping experiments showed that although the middle fragment containing residues 100-220 of LRP was able to interact with Dpl, deletion of the N-terminal domain of the full-length LRP abolished its interaction with Dpl. These results suggest that while both PrPc and Dpl interact with LRP, the domains that are involved in the binding are not the same. Our results may have implications for the molecular mechanisms of Dpl-PrPc antagonism and physiological roles of Dpl.

    Archives of biochemistry and biophysics 2004;428;2;165-9

  • The prion-like protein Doppel fails to interact with itself, the prion protein and the 37 kDa/67 kDa laminin receptor in the yeast two-hybrid system.

    Hundt C and Weiss S

    Laboratorium für Molekulare Biologie-Genzentrum-Institut für Biochemie der Ludwig-Maximilians-Universität München, Feodor-Lynen-Str. 25, D-81377 Munich, Germany.

    The prion-like protein termed Doppel (Dpl) shows approx. 25% sequence identity with all known prion proteins (PrP). We recently showed that the cellular PrP is dimeric under native conditions, a finding which was confirmed by the investigation of its crystal structure. Human PrP further interacts with its cellular receptor, the 37 kDa/67 kDa laminin receptor (LRP/LR). Here we report that human Doppel fails to interact with (i). itself, (ii). the human 37 kDa/67 kDa LRP/LR, and (iii). the human cellular prion protein (huPrP) in the yeast two-hybrid system. Our findings suggest that Dpl and PrP are not related or are only marginally related with respect to their ligand binding behaviour.

    Biochimica et biophysica acta 2004;1689;1;1-5

  • Lamr1 functional retroposon causes right ventricular dysplasia in mice.

    Asano Y, Takashima S, Asakura M, Shintani Y, Liao Y, Minamino T, Asanuma H, Sanada S, Kim J, Ogai A, Fukushima T, Oikawa Y, Okazaki Y, Kaneda Y, Sato M, Miyazaki J, Kitamura S, Tomoike H, Kitakaze M and Hori M

    Department of Internal Medicine and Therapeutics, Osaka University Graduate School of Medicine, 2-2 A8 Yamadaoka, Suita, Osaka 565-0871, Japan.

    Arrhythmogenic right ventricular dysplasia (ARVD) is a hereditary cardiomyopathy that causes sudden death in the young. We found a line of mice with inherited right ventricular dysplasia (RVD) caused by a mutation of the gene laminin receptor 1 (Lamr1). This locus contained an intron-processed retroposon that was transcribed in the mice with RVD. Introduction of a mutated Lamr1 gene into normal mice by breeding or by direct injection caused susceptibility to RVD, which was similar to that seen in the RVD mice. An in vitro study of cardiomyocytes expressing the product of mutated Lamr1 showed early cell death accompanied by alteration of the chromatin architecture. We found that heterochromatin protein 1 (HP1) bound specifically to mutant LAMR1. HP1 is a dynamic regulator of heterochromatin sites, suggesting that mutant LAMR1 impairs a crucial process of transcriptional regulation. Indeed, mutant LAMR1 caused specific changes to gene expression in cardiomyocytes, as detected by gene chip analysis. Thus, we concluded that products of the Lamr1 retroposon interact with HP1 to cause degeneration of cardiomyocytes. This mechanism may also contribute to the etiology of human ARVD.

    Nature genetics 2004;36;2;123-30

  • Complete sequencing and characterization of 21,243 full-length human cDNAs.

    Ota T, Suzuki Y, Nishikawa T, Otsuki T, Sugiyama T, Irie R, Wakamatsu A, Hayashi K, Sato H, Nagai K, Kimura K, Makita H, Sekine M, Obayashi M, Nishi T, Shibahara T, Tanaka T, Ishii S, Yamamoto J, Saito K, Kawai Y, Isono Y, Nakamura Y, Nagahari K, Murakami K, Yasuda T, Iwayanagi T, Wagatsuma M, Shiratori A, Sudo H, Hosoiri T, Kaku Y, Kodaira H, Kondo H, Sugawara M, Takahashi M, Kanda K, Yokoi T, Furuya T, Kikkawa E, Omura Y, Abe K, Kamihara K, Katsuta N, Sato K, Tanikawa M, Yamazaki M, Ninomiya K, Ishibashi T, Yamashita H, Murakawa K, Fujimori K, Tanai H, Kimata M, Watanabe M, Hiraoka S, Chiba Y, Ishida S, Ono Y, Takiguchi S, Watanabe S, Yosida M, Hotuta T, Kusano J, Kanehori K, Takahashi-Fujii A, Hara H, Tanase TO, Nomura Y, Togiya S, Komai F, Hara R, Takeuchi K, Arita M, Imose N, Musashino K, Yuuki H, Oshima A, Sasaki N, Aotsuka S, Yoshikawa Y, Matsunawa H, Ichihara T, Shiohata N, Sano S, Moriya S, Momiyama H, Satoh N, Takami S, Terashima Y, Suzuki O, Nakagawa S, Senoh A, Mizoguchi H, Goto Y, Shimizu F, Wakebe H, Hishigaki H, Watanabe T, Sugiyama A, Takemoto M, Kawakami B, Yamazaki M, Watanabe K, Kumagai A, Itakura S, Fukuzumi Y, Fujimori Y, Komiyama M, Tashiro H, Tanigami A, Fujiwara T, Ono T, Yamada K, Fujii Y, Ozaki K, Hirao M, Ohmori Y, Kawabata A, Hikiji T, Kobatake N, Inagaki H, Ikema Y, Okamoto S, Okitani R, Kawakami T, Noguchi S, Itoh T, Shigeta K, Senba T, Matsumura K, Nakajima Y, Mizuno T, Morinaga M, Sasaki M, Togashi T, Oyama M, Hata H, Watanabe M, Komatsu T, Mizushima-Sugano J, Satoh T, Shirai Y, Takahashi Y, Nakagawa K, Okumura K, Nagase T, Nomura N, Kikuchi H, Masuho Y, Yamashita R, Nakai K, Yada T, Nakamura Y, Ohara O, Isogai T and Sugano S

    Helix Research Institute, 1532-3 Yana, Kisarazu, Chiba 292-0812, Japan.

    As a base for human transcriptome and functional genomics, we created the "full-length long Japan" (FLJ) collection of sequenced human cDNAs. We determined the entire sequence of 21,243 selected clones and found that 14,490 cDNAs (10,897 clusters) were unique to the FLJ collection. About half of them (5,416) seemed to be protein-coding. Of those, 1,999 clusters had not been predicted by computational methods. The distribution of GC content of nonpredicted cDNAs had a peak at approximately 58% compared with a peak at approximately 42%for predicted cDNAs. Thus, there seems to be a slight bias against GC-rich transcripts in current gene prediction procedures. The rest of the cDNAs unique to the FLJ collection (5,481) contained no obvious open reading frames (ORFs) and thus are candidate noncoding RNAs. About one-fourth of them (1,378) showed a clear pattern of splicing. The distribution of GC content of noncoding cDNAs was narrow and had a peak at approximately 42%, relatively low compared with that of protein-coding cDNAs.

    Nature genetics 2004;36;1;40-5

  • The molecular mechanics of eukaryotic translation.

    Kapp LD and Lorsch JR

    Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, Maryland 21205-2185, USA. lkapp@jhmi.edu

    Great advances have been made in the past three decades in understanding the molecular mechanics underlying protein synthesis in bacteria, but our understanding of the corresponding events in eukaryotic organisms is only beginning to catch up. In this review we describe the current state of our knowledge and ignorance of the molecular mechanics underlying eukaryotic translation. We discuss the mechanisms conserved across the three kingdoms of life as well as the important divergences that have taken place in the pathway.

    Annual review of biochemistry 2004;73;657-704

  • The laminin receptor modulates granulocyte-macrophage colony-stimulating factor receptor complex formation and modulates its signaling.

    Chen J, Cárcamo JM, Bórquez-Ojeda O, Erdjument-Bromage H, Tempst P and Golde DW

    Department of Pharmacology, Weill Graduate School of Medical Sciences, Cornell University, New York, NY 10021, USA.

    Basement membrane matrix proteins are known to up-regulate granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling in neutrophils and mononuclear phagocytes, but the mechanisms involved are poorly understood. We used the intracellular portion of the alpha subunit of the GM-CSF receptor (alphaGMR) to search for interacting proteins and identified the 67-kDa laminin receptor (LR), a nonintegrin matrix protein receptor expressed in several types of host defense cells and certain tumors, as a binding partner. LR was found to interact with the beta subunit of the GMR (betaGMR) as well. Whereas GM-CSF functions by engaging the alphaGMR and betaGMR into receptor complexes, LR inhibited GM-CSF-induced receptor complex formation. Laminin and fibronectin binding to LR was found to prevent the binding of betaGMR to LR and relieved the LR inhibition of GMR. These findings provide a mechanistic basis for enhancing host defense cell responsiveness to GM-CSF at transendothelial migration sites while suppressing it in circulation.

    Funded by: NCI NIH HHS: CA 08748, CA30388, P30 CA008748, R01 CA030388

    Proceedings of the National Academy of Sciences of the United States of America 2003;100;24;14000-5

  • Identification of genes up-regulated in urothelial tumors: the 67-kd laminin receptor and tumor-associated trypsin inhibitor.

    Diggle CP, Cruickshank S, Olsburgh JD, Pellegrin S, Smith B, Banks RE, Selby PJ, Knowles MA, Southgate J and Harnden P

    Cancer Research United Kingdom Clinical Centre, St. James's University Hospital, Leeds, UK.

    Studies investigating changes in gene expression in urothelial carcinoma have generally compared tumors of different stages and grades but comparisons between low-grade, noninvasive tumors and normal urothelium are needed to identify genes involved in early tumor development. We isolated the urothelium from a low-grade tumor and corresponding normal mucosa by laser capture microdissection on frozen sections. The RNA extracted was amplified to generate suppressive subtractive cDNA libraries. Random sequencing of cDNA clones identified approximately 100 unique species. Of these 83% were known genes, 15% had homology to genes with an unknown function in humans, and 2% did not show homology to any published gene sequence. Two of the known genes, the 67-kd laminin receptor (67LR) and tumor-associated trypsin inhibitor (TATI), had previously been associated with metastatic progression in many tumor types, although 67LR has not been investigated in urothelial tumors. Immunolabeling of the original tissue with antibodies against these two genes confirmed overexpression, validating our strategy: 67LR was not expressed in the normal urothelium but was present in the tumor, whereas TATI expression was confined to umbrella cells in the normal urothelium, but extended to all cell layers in the tumor. We investigated both markers further in a separate series of tumors of different stages and grades. TATI was more consistently overexpressed than 67LR in all tumor grades and stages. Levels of secreted TATI were significantly higher in urine samples from patients with tumors compared to controls. Our strategy, combining laser capture microdissection and cDNA library construction, has identified genes that may be involved in the early phases of urothelial tumor development rather than with disease progression, highlighting the importance of comparing tumor with normal rather than just tumors of different stages and grades.

    The American journal of pathology 2003;163;2;493-504

  • Transcript-selective translational silencing by gamma interferon is directed by a novel structural element in the ceruloplasmin mRNA 3' untranslated region.

    Sampath P, Mazumder B, Seshadri V and Fox PL

    Department of Cell Biology, The Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.

    Transcript-selective translational control of eukaryotic gene expression is often directed by a structural element in the 3' untranslated region (3'-UTR) of the mRNA. In the case of ceruloplasmin (Cp), induced synthesis of the protein by gamma interferon (IFN-gamma) in U937 monocytic cells is halted by a delayed translational silencing mechanism requiring the binding of a cytosolic inhibitor to the Cp 3'-UTR. Silencing requires the essential elements of mRNA circularization, i.e., eukaryotic initiation factor 4G, poly(A)-binding protein, and poly(A) tail. We here determined the minimal silencing element in the Cp 3'-UTR by progressive deletions from both termini. A minimal, 29-nucleotide (nt) element was determined by gel shift assay to be sufficient for maximal binding of the IFN-gamma-activated inhibitor of translation (GAIT), an as-yet-unidentified protein or complex. The interaction was shown to be functional by an in vitro translation assay in which the GAIT element was used as a decoy to overcome translational silencing. Mutation analysis showed that the GAIT element contained a 5-nt terminal loop, a weak 3-bp helix, an asymmetric internal bulge, and a proximal 6-bp helical stem. Two invariant loop residues essential for binding activity were identified. Ligation of the GAIT element immediately downstream of a luciferase reporter conferred the translational silencing response to the heterologous transcript in vitro and in vivo; a construct containing a nonbinding, mutated GAIT element was ineffective. Translational silencing of Cp, and possibly other transcripts, mediated by the GAIT element may contribute to the resolution of the local inflammatory response following cytokine activation of macrophages.

    Funded by: NHLBI NIH HHS: HL29582, HL67725, P01 HL029582, R01 HL067725

    Molecular and cellular biology 2003;23;5;1509-19

  • p53-dependent downregulation of metastasis-associated laminin receptor.

    Modugno M, Tagliabue E, Ardini E, Berno V, Galmozzi E, De Bortoli M, Castronovo V and Ménard S

    Molecular Targeting Unit, Department of Experimental Oncology, Istituto Nazionale Tumori, 20133 Milan, Italy.

    Based on observations suggesting a role for the tumor suppressor protein p53 in regulating expression of the 67-kDa laminin receptor precursor, 37LRP, we analysed the 37LRP promoter activity in a wild-type p53 (wt p53) ovarian carcinoma cell line and in a cisplatin-resistant subline with mutated p53. We observed an increased promoter activity in wt p53 cells as compared to the mutated-p53 line when the first intron of the 37LRP gene was present in the reporter construct. Cotransfection experiments showed that the promoter is downregulated by both wt and mutated p53. Deletion analysis of the first intron localized an enhancer activity in the first 5' 214 bp that upregulates both 37LRP and SV40 promoter activity and is repressed by both wt and mutant p53. Cotransfection, mutagenesis and gel-shift experiments identified a functional AP-2 cis-acting element in this intron region that is repressed by increased levels of both wt and mutated p53. Coimmunoprecipitation studies revealed AP-2 in physical association in vivo with both wt and mutated p53, indicating for the first time that interaction of p53 with AP-2 is involved in the repression mechanism and in the regulation of genes involved in cancer growth and progression.

    Oncogene 2002;21;49;7478-87

  • Multidrug-resistance-associated protein MGr1-Ag is identical to the human 37-kDa laminin receptor precursor.

    Shi Y, Zhai H, Wang X, Wu H, Ning X, Han Y, Zhang D, Xiao B, Wu K and Fan D

    Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Shaanxi Province, PR China.

    We report the isolation and functional characterization of the gene encoding MGr1-Ag, a multidrug-resistance-associated protein. A lambdagt11 cDNA library derived from colorectal carcinoma SW480 cells was screened with monoclonal antibody MGr1. DNA homology analysis of 22 positive clones (designated R1-R22) suggested human 37-kDa laminin receptor precursor (37LRP, R7/R9/R15/R16/R19/R20) and a novel gene (R22) as candidate genes encoding MGr1-Ag. Western blot analysis showed that anti-R20 serum reacted with a unique protein band that was consistent with MGr1-Ag, while anti-R22 serum could not react with MGr1-Ag. The coding gene for MGr1-Ag was amplified using reverse transcription-PCR. Sequence analysis revealed that the MGr1-Ag and 37LRP genes shared the same coding sequence. An in vitro drug sensitivity assay indicated that down-regulation of 37LRP by an antisense technique could significantly enhance the cytotoxicity of anticancer drugs to gastric cancer cells. Thus we draw the conclusion that MGr1-Ag is identical to 37LRP.

    Cellular and molecular life sciences : CMLS 2002;59;9;1577-83

  • Directed proteomic analysis of the human nucleolus.

    Andersen JS, Lyon CE, Fox AH, Leung AK, Lam YW, Steen H, Mann M and Lamond AI

    Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230, Odense M, Denmark.

    Background: The nucleolus is a subnuclear organelle containing the ribosomal RNA gene clusters and ribosome biogenesis factors. Recent studies suggest it may also have roles in RNA transport, RNA modification, and cell cycle regulation. Despite over 150 years of research into nucleoli, many aspects of their structure and function remain uncharacterized.

    Results: We report a proteomic analysis of human nucleoli. Using a combination of mass spectrometry (MS) and sequence database searches, including online analysis of the draft human genome sequence, 271 proteins were identified. Over 30% of the nucleolar proteins were encoded by novel or uncharacterized genes, while the known proteins included several unexpected factors with no previously known nucleolar functions. MS analysis of nucleoli isolated from HeLa cells in which transcription had been inhibited showed that a subset of proteins was enriched. These data highlight the dynamic nature of the nucleolar proteome and show that proteins can either associate with nucleoli transiently or accumulate only under specific metabolic conditions.

    Conclusions: This extensive proteomic analysis shows that nucleoli have a surprisingly large protein complexity. The many novel factors and separate classes of proteins identified support the view that the nucleolus may perform additional functions beyond its known role in ribosome subunit biogenesis. The data also show that the protein composition of nucleoli is not static and can alter significantly in response to the metabolic state of the cell.

    Current biology : CB 2002;12;1;1-11

  • Laminin-1-induced migration of multiple myeloma cells involves the high-affinity 67 kD laminin receptor.

    Vande Broek I, Vanderkerken K, De Greef C, Asosingh K, Straetmans N, Van Camp B and Van Riet I

    Department of Hematology and Immunology, Free University Brussels, Laarbeeklaan 101, Brussels, B-1090, Belgium.

    The 67 kD laminin receptor (67LR) binds laminin-1 (LN), major component of the basement membrane, with high affinity. In this study, we demonstrated that human multiple myeloma cell lines (HMCL) and murine 5T2MM cells express 67LR. CD38(bright+) plasma cells in fresh multiple myeloma (MM) bone marrow (BM) samples showed weaker 67LR expression, but expression increased after direct exposure to a BM endothelial cell line (4LHBMEC). LN stimulated the in vitro migration of 3 HMCL (MM5.1, U266 and MMS.1), primary MM cells and the murine 5T2MM cells. 67LR has been shown to mediate the actions of LN through binding to CDPGYIGSR, a 9 amino acid sequence from the B1 chain of LN. MM cell migration was partially blocked by peptide 11, a synthetic nonapeptide derived from this amino sequence and also by a blocking antiserum against 67LR. Co-injection of peptide 11 with 5T2MM cells in a murine in vivo model of MM resulted in a decreased homing of 5T2MM cells to the BM compartment. In conclusion, LN acts as a chemoattractant for MM cells by interaction with 67LR. This interaction might be important during extravasation of circulating MM cells.

    British journal of cancer 2001;85;9;1387-95

  • Detection of the 67-kD laminin receptor in prostate cancer biopsies as a predictor of recurrence after radical prostatectomy.

    Waltregny D, de Leval L, Coppens L, Youssef E, de Leval J and Castronovo V

    Metastasis Research Laboratory, University of Liège, Belgium. David_Waltregny@dfci.harvard.edu

    Objectives: Reliable prognostic indicators are needed for a better pretherapeutic assessment of the agressiveness of organ-confined prostate cancer (PC) lesions. The 67-kD laminin receptor (67LR) is a cell-surface-associated protein involved in the acquisition of the invasive and metastatic phenotype of a variety of human cancer cell types. We have previously shown that 67LR detection in PC tissues from radical prostatectomy (RP) specimens is an independent predictor of biochemical (PSA) relapse in patients with clinically localized PC. In this study, we assessed 67LR detection in diagnostic PC biopsies as a predictor of biochemical relapse after RP.

    Methods: Diagnostic biopsy and subsequent RP tissue specimens from 151 patients with clinically localized PC were immunohistochemically analyzed for 67LR expression. The level of 67LR expression was evaluated by both intensity and extent of the staining. Clinicopathological preoperative and postoperative parameters, including 67LR expression, were correlated with each other and tested as predictors of biochemical relapse.

    Results: 67LR was detected in 67.5 and 68.2% of biopsies and RPs, respectively. 67LR detection in RP specimens was an independent predictor of relapse. The level of 67LR expression in the biopsy was significantly associated with the biopsy Gleason score (p<0.05) but failed to predict the pathological stage (p>0.1). Biochemical progression-free estimates for patients whose biopsy did or did not express the protein differed with only borderline statistical significance (p = 0.05). Multivariate analysis identified biopsy Gleason score as the only independent preoperative predictor of recurrence. Significant discrepancies in levels of 67LR expression were found between matched biopsy and RP specimens (p<0.05), with exact agreement rates <40%.

    Conclusions: 67LR detection in PC biopsies was not a significant preoperative predictor of outcome after RP. Heterogeneity of 67LR expression and biopsy sampling errors most likely represented the main reasons for discordant results between biopsy and RP specimens.

    European urology 2001;40;5;495-503

  • Monocytic cells synthesize, adhere to, and migrate on laminin-8 (alpha 4 beta 1 gamma 1).

    Pedraza C, Geberhiwot T, Ingerpuu S, Assefa D, Wondimu Z, Kortesmaa J, Tryggvason K, Virtanen I and Patarroyo M

    Microbiology and Tumor Biology Center, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.

    Laminins, a growing family of large heterotrimeric proteins with cell adhesive and signaling properties, are major components of vascular and other basement membranes. Expression, recognition, and use of laminin isoforms by leukocytes are poorly understood. In monoblastic THP-1 cells, transcripts for laminin gamma(1)-, beta(1)-, and alpha(4)-chains were detected by RT-PCR. Following immunoaffinity purification on a laminin beta(1) Ab-Sepharose column, laminin beta(1)- (220 kDa), gamma(1)- (200 kDa), and alpha(4)- (180/200 kDa) chains were detected by Western blotting in THP-1 cells and in two other monoblastic cell lines, U-937 and Mono Mac 6. After cell permeabilization, a mAb to laminin gamma(1)-chain reacted with practically all blood monocytes by immunofluorescence flow cytometry, and laminin-8 (alpha(4)beta(1)gamma(1)) could be isolated also from these cells. Monoblastic JOSK-I cells adhered constitutively to immobilized recombinant laminin-8, less than to laminin-10/11 (alpha(5)beta(1)gamma(1)/alpha(5)beta(2)gamma(1)) but to a higher level than to laminin-1 (alpha(1)beta(1)gamma(1)). Compared with the other laminin isoforms, adhesion to laminin-8 was preferentially mediated by alpha(6)beta(1) and beta(2) integrins. Laminin-8 and, to a lower extent, laminin-1 promoted spontaneous and chemokine-induced migration of blood monocytes, whereas laminin-10/11 was inhibitory. Altogether, the results indicate that leukocytes, as other cell types, are able to synthesize complete laminin molecules. Expression, recognition, and use of laminin-8 by leukocytes suggest a major role of this laminin isoform in leukocyte physiology.

    Journal of immunology (Baltimore, Md. : 1950) 2000;165;10;5831-8

  • Integrins as receptors for laminins.

    Belkin AM and Stepp MA

    Department of Biochemistry, The Holland Laboratory, American Red Cross, Rockville, Maryland 20855, USA.

    Laminins are a family of trimeric glycoproteins present in the extracellular matrix and the major constituents of basement membranes. Integrins are alpha beta transmembrane receptors that play critical roles in both cell-matrix and cell-cell adhesion. Several members of the integrin family, including alpha 1 beta 1, alpha 2 beta 1, alpha 3 beta 1, alpha 6 beta 1, alpha 7 beta 1 and alpha 6 beta 4 heterodimers serve as laminin receptors on a variety of cell types. This review summarizes recent advances in understanding the involvement of individual integrins in cell interactions with laminins and the roles of laminin-binding integrins in adhesion-mediated events in vertebrates, including embryonic development, cell migration and tumor cell invasiveness, cell proliferation and differentiation, as well as basement membrane assembly. We discuss the regulation of integrin function via alternative splicing of cytoplasmic domains of alpha and beta subunits of the integrin receptors for laminins and present examples of functional collaboration between laminin-binding integrins and non-integrin laminin receptors. Advances in our understanding of the laminin-binding integrins continue to demonstrate the essential roles these receptors play in maintaining cell polarity and tissue architecture.

    Funded by: NCI NIH HHS: R29 CA77697; NEI NIH HHS: R01 EY08512

    Microscopy research and technique 2000;51;3;280-301

  • The expression of membrane-associated 67-kDa laminin receptor (67LR) is modulated in vitro by cell-contact inhibition.

    Donaldson EA, McKenna DJ, McMullen CB, Scott WN, Stitt AW and Nelson J

    School of Biology and Biochemistry, Queen's University of Belfast, Belfast, BT9 7BL, Northern Ireland.

    The interaction of cells with their substratum is an important determinant of cell behaviour, influencing attachment, proliferation, and motility. Such interactions are mediated by cell surface receptors which bind to attachment factors, like the glycoprotein laminin in basement membranes. We have previously shown that expression of the 67-kDa laminin receptor (67LR) is elevated in proliferating retinal microvasculature compared with mature, quiescent vessels. Here, we examined 67LR mRNA and protein expression in primary cultures of retinal microvascular endothelial cells (RMEC) and in the breast cancer cell-line T47D during stages of contact inhibition. In both cell types, the expression levels of 67LR mRNA and membrane-associated 67LR protein were significantly increased during the proliferative phases of monolayer formation. As the cells achieved contact inhibition, 67LR expression was reduced to comparatively low levels. Thus, the differential expression of 67LR between dividing and contact-inhibited cells may indicate a role for this receptor during proliferative processes.

    Molecular cell biology research communications : MCBRC 2000;3;1;53-9

  • The nonintegrin laminin binding protein (p67 LBP) is expressed on a subset of activated human T lymphocytes and, together with the integrin very late activation antigen-6, mediates avid cellular adherence to laminin.

    Canfield SM and Khakoo AY

    Laboratory of Molecular Immunology, Department of Medicine, Columbia University, New York 10032, USA. smc12@columbia.edu

    A search for genes expressed in activated T cells revealed that the nonintegrin, 67-kDa laminin binding protein (p67 LBP) is expressed on the surface of a subset (10-15%) of activated peripheral blood T cells. Surface p67 LBP expression is detectable by FACS using the anti-p67 LBP mAb, MLuC5, within 6 h of T cell activation with phorbol dibutyrate and ionomycin, peaks 18-36 h postactivation, and persists for 7-10 days. The subset of T cells expressing p67 LBP is composed of mature, single-positive cells (85% CD4+8-, 15% CD4-8+) of memory cell phenotype (100% CD45 RO+/CD45 RA-). The p67 LBP+ T cells also express the integrin alpha6 chain (CD49f), which is known to associate with p67 LBP on tumor cells. In addition, the p67 LBP+ T cells express the integrin beta1, which associates with alpha6 in the laminin-specific integrin receptor very late activation Ag (VLA)-6 (alpha6beta1). Expression of an exogenous cDNA encoding the 37-kDa LBP precursor (p37 LBPP) confers p67 LBP surface expression on a p67 LBP-negative Jurkat T cell line (B2.7). Expression of p67 LBP induces B2.7 transfectants to adhere to laminin, but avid laminin binding depends on coexpression of VLA-6. Taken together, these data indicate that p67 LBP is an activation-induced surface structure on memory T cells that, together with VLA-6, mediates cellular adherence to laminin.

    Funded by: NCI NIH HHS: R-01 CA 55713

    Journal of immunology (Baltimore, Md. : 1950) 1999;163;6;3430-40

  • Ribosome-associated protein LBP/p40 binds to S21 protein of 40S ribosome: analysis using a yeast two-hybrid system.

    Sato M, Saeki Y, Tanaka K and Kaneda Y

    Division of Gene Therapy Science, Osaka University School of Medicine, 2-2 Yamada-oka, Osaka, Suita City, 565-0871, Japan.

    The ribosome-associated protein LBP/p40, which was originally named after "laminin binding protein precursor p40," is distributed on the cell surface as laminin binding protein p67 (LBP/p67), in the nucleus, and on 40S ribosomes. In a broad range of eukaryotes, the localization of LBP/p40 on the 40S ribosome is well conserved. Two yeast homologs of LBP/p40 are believed to be essential for cell viability and each gene product probably corresponds to the assembly and/or stability of the 40S ribosomal subunit. The precise role of LBP/p40 in translation, however, remains to be elucidated, especially in higher eukaryotes. In this report, we used a yeast two-hybrid screening method to isolate molecules associated with human LBP/p40 protein on ribosomes. We found that the 40S ribosomal protein S21 was tightly bound with LBP/p40 in this yeast two-hybrid system and in in vitro analysis. Further, we discovered that the association required a broad region of the LBP/p40 amino acid sequence, which corresponds to the highly conserved region of LBP/p40 homologs among eukaryotes.

    Biochemical and biophysical research communications 1999;256;2;385-90

  • Prognostic significance of 67-kDa laminin receptor expression in advanced gastric cancer.

    de Manzoni G, Guglielmi A, Verlato G, Tomezzoli A, Pelosi G, Schiavon I and Cordiano C

    Istituto di Semeiotica Chirurgica, Università di Verona, Verona, Italia.

    The ability of cancer cells to attach to laminin has been correlated with their metastatic potential and highly metastatic cancer cells seem to express on their surface significantly more laminin receptors than do their much less metastatic or benign counterparts. The expression of 67-kDa laminin receptors (LR) was investigated in a group of 75 patients who underwent gastrectomy for advanced gastric cancer, with special reference to the possible role in the tumor progression and in the overall survival. The tumor LR expression was immunohistochemically determined in paraffin-embedded sections using the MLuC5 monoclonal antibody which recognizes the 67-kDa LR and the avidin-biotin immunoperoxidase method. Of the 75 cases analyzed, 43 cases (57.3%) displayed a positive reaction. The cumulative 5-year survival rate was 72.6% (95% CI 52.5-85.3) for patients without expression of LR and 46.6% (29.8-61.8) for those with positive LR expression. A significant association between LR expression and depth of tumor invasion (0.022) was found. By univariate analysis the presence of laminin receptors seemed to be associated with a higher risk of death [RR 1.72 (95% CI 0.71-4.20], but this effect disappeared after controlling for depth of tumor invasion. In conclusion, these results suggest that tumor expression of laminin receptors could be correlated with tumor aggressiveness. However, the prognostic significance of laminin receptor expression is already provided by the depth of tumor invasion.

    Oncology 1998;55;5;456-60

  • A map of 75 human ribosomal protein genes.

    Kenmochi N, Kawaguchi T, Rozen S, Davis E, Goodman N, Hudson TJ, Tanaka T and Page DC

    Howard Hughes Medical Institute, Whitehead Institute and Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02142, USA. kenmochi@med.u-ryuku.ac.jp

    We mapped 75 genes that collectively encode >90% of the proteins found in human ribosomes. Because localization of ribosomal protein genes (rp genes) is complicated by the existence of processed pseudogenes, multiple strategies were devised to identify PCR-detectable sequence-tagged sites (STSs) at introns. In some cases we exploited specific, pre-existing information about the intron/exon structure of a given human rp gene or its homolog in another vertebrate. When such information was unavailable, selection of PCR primer pairs was guided by general insights gleaned from analysis of all mammalian rp genes whose intron/exon structures have been published. For many genes, PCR amplification of introns was facilitated by use of YAC pool DNAs rather than total human genomic DNA as templates. We then assigned the rp gene STSs to individual human chromosomes by typing human-rodent hybrid cell lines. The genes were placed more precisely on the physical map of the human genome by typing of radiation hybrids or screening YAC libraries. Fifty-one previously unmapped rp genes were localized, and 24 previously reported rp gene localizations were confirmed, refined, or corrected. Though functionally related and coordinately expressed, the 75 mapped genes are widely dispersed: Both sex chromosomes and at least 20 of the 22 autosomes carry one or more rp genes. Chromosome 19, known to have a high gene density, contains an unusually large number of rp genes (12). This map provides a foundation for the study of the possible roles of ribosomal protein deficiencies in chromosomal and Mendelian disorders.

    Genome research 1998;8;5;509-23

  • Expression of high-affinity 67-kDa laminin receptors in primary breast cancers and metachronous metastatic lesions or contralateral cancers.

    Daidone MG, Silvestrini R, Benini E, Grigioni WF and D'Errico A

    Oncologia Sperimentale C, Istituto Nazionale per lo Studio e la Cura del Tumori, Milano, Italy.

    The presence of high-affinity 67-kDa laminin receptors, detected immunohistochemically, was determined on 63 primary breast cancers and on metachronous metastatic lesions or contralateral cancers from the same patients. A disagreement was observed in two-thirds of the cases. In particular, laminin receptor content was significantly lower (P = 0.02) in local recurrences and slightly higher in lymph node metastasis than in the corresponding primary tumours.

    British journal of cancer 1997;76;1;52-3

  • Identification of the active gene coding for the metastasis-associated 37LRP/p40 multifunctional protein.

    Clausse N, Jackers P, Jarès P, Joris B, Sobel ME and Castronovo V

    Metastasis Research Laboratory, University of Liège, Belgium.

    A 37LRP/p40 polypeptide is of major interest because it is consistently up-regulated in cancer cells in correlation with their invasive and metastatic phenotype. Furthermore, this polypeptide presents intriguing multifunctional properties because it has been characterized as the precursor of the metastasis-associated 67-kD laminin receptor (67LR) and as a cytoplasmic ribosomal-associated protein. The isolation of the 37LRP/p40 gene is a prerequisite for identifying the molecular mechanisms responsible for the constant up-regulation of the 67LR expression in cancer cells. To date, the active 37LRP/p40 gene has never been identified in any species due to the existence of multiple pseudogenes in most vertebrates genomes. In this study, we report for the first time the gene structure and potential regulatory sequences of the 37 LRP/p40 gene. The chicken genome was selected to undergo this characterization because it is the only known vertebrate that bears a single 37 LRP/p40 gene copy. The 37 LRP/p40 active gene is composed of 7 exons and 6 introns and bears features characteristic of a ribosomal protein gene. It does not bear a classical TATA box and it exhibits several transcription initiation sites as demonstrated by RNase protection assay and primer extension. Analysis of potential regulatory regions suggests that gene expression is driven not only by the 5' genomic region but also by the 5' untranslated and intron 1 sequences. On the basis of gene structure and extensive protein evolutionary study, we found that the carboxyterminal domain of the protein is a conserved lock-and-key structure/function domain that could be involved in the biosynthesis of the higher-molecular-weight 67-kD laminin receptor in vertebrates, whereas the central core of the protein would be responsible for the ribosome associated function. The first identification of the active 37LRP/p40 gene presented in this study is a critical step toward the isolation of the corresponding human gene and the understanding of the molecular mechanisms involved in the up-regulation of its expression during tumor invasion and metastasis.

    DNA and cell biology 1996;15;12;1009-23

  • Isolation from a multigene family of the active human gene of the metastasis-associated multifunctional protein 37LRP/p40 at chromosome 3p21.3.

    Jackers P, Minoletti F, Belotti D, Clausse N, Sozzi G, Sobel ME and Castronovo V

    Metastasis Research Laboratory, University of Liège, Belgium.

    The 37 kD precursor of the 67 kD laminin receptor (37LRP) is a polypeptide whose expression is consistently upregulated in aggressive carcinoma. Interestingly, the 37LRP appears to be a multifunctional protein involved in the translational machinery and has also been identified as p40 ribosome-associated protein. Although highly conserved cDNAs corresponding to this polypeptide have been isolated from several species including vertebrates, invertebrates, plants and prokaryotes, characterization of any of the corresponding active genes has never been reported. In this study, we have cloned an intron-containing fragment which permitted us to isolate the active 37LRP/p40 human gene. This gene contains seven exons and six introns. Ribonuclease protection experiments suggest multiple transcription start sites. The promoter area does not bear a TATA box but contains four Sp1 sites. The first intron is also GC rich containing five Sp1 sites. Intron 4 contains the full sequence of the small nuclear RNA E2 and two Alu sequences are found in intron 3. Fluorescent in situ hybridization localized the 37LRP/p40 active gene on chromosome 3 in the locus 3p21.3 which, interestingly, is a hot spot for genetic alterations in several cancers and particularly in small cell lung carcinoma.

    Oncogene 1996;13;3;495-503

  • Characterization of the human small-ribosomal-subunit proteins by N-terminal and internal sequencing, and mass spectrometry.

    Vladimirov SN, Ivanov AV, Karpova GG, Musolyamov AK, Egorov TA, Thiede B, Wittmann-Liebold B and Otto A

    Novosibirsk Institute of Bioorganic Chemistry, Siberian Division, Russian Academy of Sciences, Russian Federation.

    Reverse-phase HPLC was used to fractionate 40S ribosomal proteins from human placenta. Application of a C4 reverse-phase column allowed us to obtain 27 well-resolved peaks. The protein composition of each chromatographic fraction was established by two-dimensional polyacrylamide gel electrophoresis and N-terminal sequencing. N-terminally blocked proteins were cleaved with endoproteinase Lys-C, and suitable peptides were sequenced. All sequences were compared with those of ribosomal proteins available from data bases. This allowed us to identify all proteins from the 40S human ribosomal subunit in the HPLC elution profile. By matrix-assisted laser-desorption ionization mass spectrometry the masses of the 40S proteins were determined and checked for the presence of post-translational modifications. For several proteins differences to the deduced sequences and the calculated masses were found to be due to post-translational modifications.

    European journal of biochemistry 1996;239;1;144-9

  • The gene for human E2 small nucleolar RNA resides in an intron of a laminin-binding protein gene.

    Selvamurugan N and Eliceiri GL

    Department of Pathology, Saint Louis University School of Medicine, St. Louis, Missouri 63104-1028, USA.

    Genomics 1995;30;2;400-1

  • Cloning of 67-kDa laminin receptor cDNA and gene expression in normal and malignant cell lines of the human lung.

    Satoh K, Narumi K, Sakai T, Abe T, Kikuchi T, Matsushima K, Sindoh S and Motomiya M

    Department of Internal Medicine, Tohoku University, Sendai, Japan.

    Cell-adhesive protein laminin and its specific receptor play an important role in the processes of cancer proliferation, invasion and metastasis. In the present study, we cloned the cDNAs of the 67-kDa laminin receptor both from a human lung cell line (IMR90) and from a human lung cancer cell line (SBC3), and determined the nucleotide sequences. In comparison with both cDNA sequences of the protein-coding region, three nucleotide differences were found. These differences in the secondary structure of the protein, however, were caused by nucleotide substitutions. It was also demonstrated that the level of 67-kDa-laminin receptor mRNA was higher in SBC3 than in IMR90.

    Cancer letters 1992;62;3;199-203

  • Characteristics of a multicopy gene family predominantly consisting of processed pseudogenes.

    Van den Ouweland AM, Van Duijnhoven HL, Deichmann KA, Van Groningen JJ, de Leij L and Van de Ven WJ

    Department of Biochemistry, University of Nijmegen, The Netherlands.

    The monoclonal antibody MOC-32 detected a 40 kDa protein in Western blot analysis. Immunological screening of an expression library of human SCLC cells with MOC-32 led to the isolation of overlapping cDNA clones. One of these, cHD4, was 1.0 kbp long and of about the same size as its corresponding mRNA. Preceded by an in phase stop codon, an open reading frame of 885 bp was present in cHD4 and a translational product of only 33 kDa could be calculated. Biochemical and immunological analysis established the relationship between the 40 kDa antigen and the isolated coding sequences and resolved the apparent discrepancy between the calculated molecular weight and the observed electrophoretic mobility. Nucleotide sequence comparison of cHD4 to the EMBL database revealed that cHD4 was nearly identical to a sequence claimed to encode a laminin binding protein. Southern blot and nucleotide sequence analysis indicated the presence of multiple copies of the gene in the human genome. At least five of these appeared to represent processed pseudogenes.

    Nucleic acids research 1989;17;10;3829-43

  • The human laminin receptor is a member of the integrin family of cell adhesion receptors.

    Gehlsen KR, Dillner L, Engvall E and Ruoslahti E

    Cancer Research Center, La Jolla Cancer Research Foundation, CA 92037.

    A receptor for the adhesive basement membrane protein, laminin, was isolated from human glioblastoma cells by affinity chromatography on laminin. This receptor has a heterodimeric structure similar to that of receptors for other extracellular matrix proteins such as fibronectin and vitronectin. Incorporation of the laminin receptor into liposomal membranes makes it possible for liposomes to attach to surfaces coated with laminin. The receptor liposomes also attached to some extent to surfaces coated with fibronectin, but not with other matrix proteins. These properties identify the laminin receptor as a member of the integrin family of cell adhesion receptors.

    Funded by: NCI NIH HHS: CA28896, CA42507; NIDDK NIH HHS: DK30051; ...

    Science (New York, N.Y.) 1988;241;4870;1228-9

  • Increased mRNA expression of a laminin-binding protein in human colon carcinoma: complete sequence of a full-length cDNA encoding the protein.

    Yow HK, Wong JM, Chen HS, Lee CG, Davis S, Steele GD and Chen LB

    Dana-Farber Cancer Institute, New England Deaconess Hospital, Boston, MA.

    Reliable markers to distinguish human colon carcinoma from normal colonic epithelium are needed particularly for poorly differentiated tumors where no useful marker is currently available. To search for markers we constructed cDNA libraries from human colon carcinoma cell lines and screened for clones that hybridize to a greater degree with mRNAs of colon carcinomas than with their normal counterparts. Here we report one such cDNA clone that hybridizes with a 1.2-kilobase (kb) mRNA, the level of which is approximately equal to 9-fold greater in colon carcinoma than in adjacent normal colonic epithelium. Blot hybridization of total RNA from a variety of human colon carcinoma cell lines shows that the level of this 1.2-kb mRNA in poorly differentiated colon carcinomas is as high as or higher than that in well-differentiated carcinomas. Molecular cloning and complete sequencing of cDNA corresponding to the full-length open reading frame of this 1.2-kb mRNA unexpectedly show it to contain all the partial cDNA sequence encoding 135 amino acid residues previously reported for a human laminin receptor. The deduced amino acid sequence suggests that this putative laminin-binding protein from human colon carcinomas consists of 295 amino acid residues with interesting features. Containing only two cysteine residues, the protein does not have consensus sequences for asparagine-linked glycosylation, amphipathic alpha-helix, or the N-terminal leader signal sequences for entry into endoplasmic reticulum, although hydrophobic segments for potential membrane associations exist. There is an unusual C-terminal 70-amino acid segment, which is trypsin-resistant (no lysine or arginine) and highly negatively charged (13 aspartic plus glutamic residues). Within this segment are five repeats of (Asp/Glu)-Trp-(Ser/Thr); two of these are nearly tandem repeats of Thr-Glu-Asp-Trp-Ser-Ala-Xaa-Pro.

    Funded by: NCI NIH HHS: P01 CA22427, P01 CA44704; NIGMS NIH HHS: R01 GM38318

    Proceedings of the National Academy of Sciences of the United States of America 1988;85;17;6394-8

  • Altered levels of laminin receptor mRNA in various human carcinoma cells that have different abilities to bind laminin.

    Wewer UM, Liotta LA, Jaye M, Ricca GA, Drohan WN, Claysmith AP, Rao CN, Wirth P, Coligan JE, Albrechtsen R et al.

    The human laminin receptor was purified and molecularly cloned to investigate its biosynthetic regulation. Laminin receptor from normal and neoplastic tissue was preparatively affinity purified to homogeneity based on the high affinity of the receptor for laminin. The apparent molecular weight of the receptor from different carcinoma sources and from normal placental tissue is in the range of 68-72 kDa. Isoelectric focusing and two-dimensional gel electrophoresis indicated that the receptor protein consists of one major polypeptide chain with a pI value of 6.4 +/- 0.2. Laminin receptor cDNA clones were isolated after screening a human endothelial lambda gt11 cDNA library with a monoclonal antibody directed against a domain of the laminin receptor involved in ligand binding. Definitive identification of the cDNA clones was based on comparison of cDNA sequence with the amino acid sequence of a cyanogen bromide-generated octapeptide of purified placental laminin receptor. The laminin receptor mRNA is approximately 1700 bases long. The level of laminin receptor mRNA in a variety of human carcinoma-derived cell lines correlated with the number of laminin receptors on the cell surfaces of those cells. This suggests that the amount of laminin receptor mRNA may be a rate-limiting control step in the biosynthesis of the laminin receptor, and hence in the regulation of cellular attachment to basement membranes via laminin.

    Proceedings of the National Academy of Sciences of the United States of America 1986;83;19;7137-41

Gene lists (3)

Gene List Source Species Name Description Gene count
L00000009 G2C Homo sapiens Human PSD Human orthologues of mouse PSD adapted from Collins et al (2006) 1080
L00000016 G2C Homo sapiens Human PSP Human orthologues of mouse PSP adapted from Collins et al (2006) 1121
L00000069 G2C Homo sapiens BAYES-COLLINS-HUMAN-PSD-FULL Human cortex biopsy PSD full list 1461
© G2C 2014. The Genes to Cognition Programme received funding from The Wellcome Trust and the EU FP7 Framework Programmes:
EUROSPIN (FP7-HEALTH-241498), SynSys (FP7-HEALTH-242167) and GENCODYS (FP7-HEALTH-241995).

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