G2Cdb::Gene report

Gene id
Gene symbol
Rpsa (MGI)
Mus musculus
ribosomal protein SA
G00002468 (Homo sapiens)

Databases (9)

ENSMUSG00000032518 (Ensembl mouse gene)
16785 (Entrez Gene)
905 (G2Cdb plasticity & disease)
Gene Expression
NM_011029 (Allen Brain Atlas)
g02415 (BGEM)
rpsa (gensat)
150370 (OMIM)
Marker Symbol
MGI:105381 (MGI)
Protein Sequence
P14206 (UniProt)

Synonyms (6)

  • 67lr
  • Lamr
  • Lamr1
  • Lamrl1
  • P40-3
  • P40-8

Literature (44)

Pubmed - other

  • Green tea polyphenol epigallocatechin-3-gallate inhibits TLR2 signaling induced by peptidoglycan through the polyphenol sensing molecule 67-kDa laminin receptor.

    Byun EH, Omura T, Yamada K and Tachibana H

    Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, Fukuoka 812-8581, Japan.

    Here we show the molecular basis for the inhibition of peptidoglycan (PGN)-induced TLR2 signaling by a major green tea polyphenol epigallocatechin-3-gallate (EGCG). Recently, we identified the 67-kDa laminin receptor (67LR) as the cell-surface EGCG receptor. Anti-67LR antibody treatment or silencing of 67LR resulted in abrogation of the inhibitory action of EGCG on PGN-induced production of pro-inflammatory mediators and activation of mitogen-activated protein kinases. Silencing of Toll-interacting protein (Tollip), a negative regulator of TLR signaling impaired the TLR2 signaling inhibitory activity of EGCG, suggesting that TLR2 response could be inhibited by EGCG via 67LR and Tollip.

    FEBS letters 2011;585;5;814-20

  • TGF-beta1 improves cardiac performance via up-regulation of laminin receptor 37/67 in adult ventricular cardiomyocytes.

    Wenzel S, Henning K, Habbig A, Forst S, Schreckenberg R, Heger J, Maxeiner H and Schlüter KD

    Institute of Physiology, Justus-Liebig-University, Giessen, Germany. sibylle.wenzel@ugcvr.de

    TGF-beta1 plays an important role in cardiac fibrosis, apoptosis, induction of hypertrophy and contractile dysfunction. This study investigates whether TGF-beta1 plays a role in laminin receptor 37/67 (37/67 LR)-dependent regulation of cardiac performance. Therefore, isolated adult cardiomyocytes were stimulated with TGF-beta1, the expression of the 37/67 LR was determined and cell shortening was investigated on cells attached to a non-specific, serum-based attachment substrate or to specific, laminin-coated dishes. The role of the MAP kinases in TGF-beta1-dependent induction of the 37/67 LR was examined by addition of PD98059, SB202190 and SP600125. Finally, the expression of receptor mRNA was investigated in transgenic mice constitutively over-expressing TGF-beta1 and the relationship to distress score and lung wet weight-to-body weight was analysed. TGF-beta1 induced a significant increase of the 37/67 LR mRNA and protein expression. The cytokine induced p38 MAP kinase and JNK, but not ERK. Inhibition of either p38 MAP kinase or JNK attenuated the TGF-beta1-dependent increase in 37/67 LR expression. TGF-beta1 induced a loss of cell shortening in cells attached to a non-specific substrate, but not in cells on a pre-coated laminin matrix. Inhibition of JNK attenuated the protective effect of laminin receptor up-regulation on cardiac performance. Inhibition of p38 MAP kinase attenuated the depressive effect of TGF-beta1 on basal cell shortening. In transgenic mice over-expressing TGF-beta1 a strong induction of laminin receptor expression attenuated the severeness of the mice' symptoms. This study shows a new and protective role of TGF-beta1-dependent up-regulation of the 37/67 LR in cardiomyocytes in cardiac remodelling with increased laminin expression.

    Basic research in cardiology 2010;105;5;621-9

  • TLR4 signaling inhibitory pathway induced by green tea polyphenol epigallocatechin-3-gallate through 67-kDa laminin receptor.

    Hong Byun E, Fujimura Y, 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, Japan.

    Epigallocatechin-3-gallate (EGCG), a major active polyphenol of green tea, has been shown to downregulate inflammatory responses in macrophages; however, the underlying mechanism has not been understood. Recently, we identified the 67-kDa laminin receptor (67LR) as a cell-surface EGCG receptor that mediates the anticancer action of EGCG at physiologically relevant concentrations (0.1-1 microM). In this study, we show the molecular basis for the downregulation of TLR4 signal transduction by EGCG at 1 microM in macrophages. Anti-67LR Ab treatment or RNA interference-mediated silencing of 67LR resulted in abrogation of the inhibitory action of EGCG on LPS-induced activation of downstream signaling pathways and target gene expressions. Additionally, we found that EGCG reduced the TLR4 expression through 67LR. Interestingly, EGCG induced a rapid upregulation of Toll-interacting protein (Tollip), a negative regulator of TLR signaling, and this EGCG action was prevented by 67LR silencing or anti-67LR Ab treatment. RNA interference-mediated silencing of Tollip impaired the TLR4 signaling inhibitory activity of EGCG. Taken together, these findings demonstrate that 67LR plays a critical role in mediating anti-inflammatory action of a physiologically relevant EGCG, and Tollip expression could be modulated through 67LR. These results provide a new insight into the understanding of negative regulatory mechanisms for the TLR4 signaling pathway and consequent inflammatory responses that are implicated in the development and progression of many chronic diseases.

    Journal of immunology (Baltimore, Md. : 1950) 2010;185;1;33-45

  • Laminin receptor activation inhibits endothelial tissue factor expression.

    Holy EW, Stämpfli SF, Akhmedov A, Holm N, Camici GG, Lüscher TF and Tanner FC

    Cardiovascular Research, Physiology Institute, University of Zurich, Zurich, Switzerland.

    Tissue factor (TF) is an important trigger of arterial thrombosis. The green tea catechin epigallocatechin-3-gallate (EGCG) is a ligand of the 67-kDa laminin receptor (67LR) and exhibits cardioprotective effects. This study investigates whether 67LR regulates TF expression in human endothelial cells. Immunofluorescence demonstrated that human aortic endothelial cells expressed 67LR. Cells grown on laminin expressed 35% less TF in response to TNF-alpha (TNF-alpha) than those grown on fibronectin (n=6; p<0.001). EGCG (1-30 microM) inhibited TNF-alpha and histamine induced endothelial TF expression and activity in a concentration dependent manner resulting in 87% reduction of TF expression (n=5; p<0.001); in contrast, expression of tissue factor pathway inhibitor was not affected (n=4; p=NS). In vivo administration of EGCG (30 mg/kg/day) inhibited TF activity in carotid arteries of C57BL6 mice. Real-time PCR and promoter studies revealed that EGCG decreased TF expression at the transcriptional level and impaired activation of the mitogen activated protein (MAP) kinase JNK 1/2, but not ERK or p38. Similarly, the JNK 1/2 inhibitor SP600125 (1 microM) impaired TF promoter activity (n=4; p<0.001) and protein expression (n=4; p<0.001). 67LR blocking antibodies blunted the inhibitory effect of EGCG on both TF protein expression and JNK activation. In contrast, vascular cell adhesion molecule 1 (VCAM-1) was not affected by laminin nor EGCG, and its expression was not regulated by JNK. EGCG did not affect TNF-alpha stimulated NFkB activation. Laminin receptor activation inhibits endothelial TF expression by impairing JNK phosphorylation. Thus, 67LR may be a potential target for the development of novel anti-thrombotic therapies.

    Journal of molecular and cellular cardiology 2010;48;6;1138-45

  • Relationships between hematopoiesis and hepatogenesis in the midtrimester fetal liver characterized by dynamic transcriptomic and proteomic profiles.

    Guo Y, Zhang X, Huang J, Zeng Y, Liu W, Geng C, Li KW, Yang D, Wu S, Wei H, Han Z, Qian X, Jiang Y and He F

    State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing, China.

    In fetal hematopoietic organs, the switch from hematopoiesis is hypothesized to be a critical time point for organogenesis, but it is not yet evidenced. The transient coexistence of hematopoiesis will be useful to understand the development of fetal liver (FL) around this time and its relationship to hematopoiesis. Here, the temporal and the comparative transcriptomic and proteomic profiles were observed during the critical time points corresponding to the initiation (E11.5), peak (E14.5), recession (E15.5), and disappearance (3 ddp) of mouse FL hematopoiesis. We found that E11.5-E14.5 corresponds to a FL hematopoietic expansion phase with distinct molecular features, including the expression of new transcription factors, many of which are novel KRAB (Kruppel-associated box)-containing zinc finger proteins. This time period is also characterized by extensive depression of some liver functions, especially catabolism/utilization, immune and defense, classical complement cascades, and intrinsic blood coagulation. Instead, the other liver functions increased, such as xenobiotic and sterol metabolism, synthesis of carbohydrate and glycan, the alternate and lectin complement cascades and extrinsic blood coagulation, and etc. Strikingly, all of the liver functions were significantly increased at E14.5-E15.5 and thereafter, and the depression of the key pathways attributes to build the hematopoietic microenvironment. These findings signal hematopoiesis emigration is the key to open the door of liver maturation.

    PloS one 2009;4;10;e7641

  • Green tea (-)-epigallocatechin gallate inhibits insulin stimulation of 3T3-L1 preadipocyte mitogenesis via the 67-kDa laminin receptor pathway.

    Ku HC, Chang HH, Liu HC, Hsiao CH, Lee MJ, Hu YJ, Hung PF, Liu CW and Kao YH

    Dept. of Life Science, College of Science, National Central Univ., Jhong-li, Taiwan 32054.

    Insulin and (-)-epigallocatechin gallate (EGCG) have been reported to regulate fat cell mitogenesis and adipogenesis, respectively. This study investigated the pathways involved in EGCG modulation of insulin-stimulated mitogenesis in 3T3-L1 preadipocytes. EGCG inhibited insulin stimulation of preadipocyte proliferation in a dose- and time-dependent manner. EGCG also suppressed insulin-stimulated phosphorylation of the insulin receptor-beta, insulin receptor (IR) substrates 1 and 2 (IRS1 and IRS2), and mitogen-activated protein kinase pathway proteins, RAF1, MEK1/2, and ERK1/2, but not JNK. Furthermore, EGCG inhibited the association of IR with the IRS1 and IRS2 proteins, but not with the IRS4 protein. These data suggest that EGCG selectively affects particular types of IRS and MAPK family members. Generally, EGCG was more effective than epicatechin, epicatechin gallate, and epigallocatechin in modulating insulin-stimulated mitogenic signaling. We identified the EGCG receptor [also known as the 67-kDa laminin receptor (67LR)] in fat cells and found that its expression was sensitive to growth phase, tissue type, and differentiation state. Pretreatment of preadipocytes with 67LR antiserum prevented the effects of EGCG on insulin-stimulated phosphorylation of IRS2, RAF1, and ERK1/2 and insulin-stimulated preadipocyte proliferation (cell number and bromodeoxyuridine incorporation). Moreover, EGCG tended to increase insulin-stimulated associations between the 67LR and IR, IRS1, IRS2, and IRS4 proteins. These data suggest that EGCG mediates anti-insulin signaling in preadipocyte mitogenesis via the 67LR pathway.

    American journal of physiology. Cell physiology 2009;297;1;C121-32

  • Laminin receptor initiates bacterial contact with the blood brain barrier in experimental meningitis models.

    Orihuela CJ, Mahdavi J, Thornton J, Mann B, Wooldridge KG, Abouseada N, Oldfield NJ, Self T, Ala'Aldeen DA and Tuomanen EI

    Department of Infectious Diseases, St. Jude Children's Research Hospital, Memphis, Tennessee, USA.

    A diverse array of infectious agents, including prions and certain neurotropic viruses, bind to the laminin receptor (LR), and this determines tropism to the CNS. Bacterial meningitis in childhood is almost exclusively caused by the respiratory tract pathogens Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae, but the mechanism by which they initiate contact with the vascular endothelium of the blood brain barrier (BBB) is unknown. We hypothesized that an interaction with LR might underlie their CNS tropism. Using affinity chromatography, coimmunoprecipitation, retagging, and in vivo imaging approaches, we identified 37/67-kDa LR as a common receptor for all 3 bacteria on the surface of rodent and human brain microvascular endothelial cells. Mutagenesis studies indicated that the corresponding bacterial LR-binding adhesins were pneumococcal CbpA, meningococcal PilQ and PorA, and OmpP2 of H. influenzae. The results of competitive binding experiments suggest that a common adhesin recognition site is present in the carboxyl terminus of LR. Together, these findings suggest that disruption or modulation of the interaction of bacterial adhesins with LR might engender unexpectedly broad protection against bacterial meningitis and may provide a therapeutic target for the prevention and treatment of disease.

    Funded by: NCI NIH HHS: CA21765, P30 CA021765; NIAID NIH HHS: R01 AI027913, R01 AI27913

    The Journal of clinical investigation 2009;119;6;1638-46

  • Subcellular localization of prion proteins and the 37 kDa/67 kDa laminin receptor fused to fluorescent proteins.

    Nikles D, Vana K, Gauczynski S, Knetsch H, Ludewigs H and Weiss S

    Laboratorium für Molekulare Biologie - Genzentrum-Institut für Biochemie der LMU München, Feodor-Lynen-Str. 25, 81377 München, Germany.

    The 37 kDa/67 kDa laminin receptor LRP/LR acts as a receptor for both PrPc and PrPSc at the cell surface. Here, we further analyzed the subcellular localization of fluorescent labeled prion protein (PrP) and laminin receptor (LRP/LR) molecules. We show that EGFP-PrP is localized at the cell surface and in a perinuclear compartment, respectively. In contrast, a DsRed-DeltaSP-PrP mutant lacking the signal peptide is almost exclusively found in the nucleus but does not colocalize with heterochromatin. Interestingly, LRP-DsRed efficiently colocalizes with EGFP-PrP in the perinuclear compartment and LRP-ECFP partly colocalizes with DsRed-DeltaSP-PrP in the nucleus, respectively. We conclude that the interactions of PrP and LRP/LR are not restricted to the cell surface but occur also in intracellular compartments suggesting a putative role of LRP/LR in the trafficking of PrP molecules.

    Biochimica et biophysica acta 2008;1782;5;335-40

  • Ciz, a transcription factor with a nucleocytoplasmic shuttling activity, interacts with C-propeptides of type I collagen.

    Hayata T, Nakamoto T, Ezura Y and Noda M

    Department of Molecular Pharmacology, Medical Research Institute, Tokyo Medical and Dental University, 3-10 Kanda-Surugadai 2-Chome, Chiyoda-ku, Tokyo 101-0062, Japan.

    Ciz is a zinc finger transcription factor with nucleocytoplasmic shuttling activity. Ciz-deficient mice show high bone mass phenotype. As a first step to address how Ciz suppresses bone formation, we examined the binding partners of Ciz based on a yeast two-hybrid screening. While Ciz is an intracellular protein, 47% of the positive clones were genes encoding extracellular matrix proteins, including Col1a1, Col1a2, Fbln2, and Rpsa. In vitro coimmunoprecipitation experiments using in vitro translated proteins revealed direct binding of Ciz-DeltaZF (zinc finger) to C-propeptides of Col1a1 and Col1a2. In vivo association of the transfected Ciz and C-propeptide of Col1a1 was observed in COS-7 cells based on immunoprecipitation. In terms of intracellular localization, overexpressed C-propeptides of Col1 and Ciz were co-localized in nuclei. These results revealed that Ciz interacts with C-propeptides of type I collagen and this association takes place in nuclei.

    Biochemical and biophysical research communications 2008;368;2;205-10

  • 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

  • MARCKS-like protein, a membrane protein identified for its expression in developing neural retina, plays a role in regulating retinal cell proliferation.

    Zhao J, Izumi T, Nunomura K, Satoh S and Watanabe S

    Department of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, 108-8639, Tokyo, Japan.

    Membrane proteins are expressed in a specific manner in developing tissues, and characterization of these proteins is valuable because it allows them to be used as cell surface markers. Furthermore, they are potentially important for the regulation of organogenesis because some may participate in signal transduction. In the present study, we used proteomics to examine the comprehensive protein expression profile of the membrane fraction in the embryonic and adult mouse retina. We purified the retinal membrane fraction by sucrose-density-gradient centrifugation and analysed total proteins using shotgun analysis on a nanoflow LC-MS/MS (liquid chromatography tandem MS) system. Approximately half of the 326 proteins from the adult retina and a quarter of the 310 proteins from the embryonic retina (day 17) appeared to be membrane-associated proteins. Among these, MLP [MARCKS (myristoylated alanine-rich C-kinase substrate)-like protein], which shares approx. 50% amino acid identity with MARCKS, was selected for further characterization. The mRNA and surface protein expression of MLP decreased as retinal development progressed. Overexpression of MLP by retrovirus-mediated gene transfer enhanced the proliferation of retinal progenitor cells without affecting differentiation or cell migration in a retinal explant culture system. In contrast, MLP overexpression did not promote proliferation in fibroblasts (NIH 3T3 cells). Mutation analysis of MLP demonstrated that myristoylation was necessary to promote proliferation and that phosphorylation inhibited proliferation, indicating the functional importance of membrane localization.

    The Biochemical journal 2007;408;1;51-9

  • The metastasis-associated 67-kDa laminin receptor is involved in G-CSF-induced hematopoietic stem cell mobilization.

    Selleri C, Ragno P, Ricci P, Visconte V, Scarpato N, Carriero MV, Rotoli B, Rossi G and Montuori N

    Department of Biochemistry and Medical Biotechnology, Institute of Experimental Endocrinology and Oncology (IEOS), National Research Council (CNR), Via S Pansini 5, 80131, Naples, Italy.

    The 67-kDa laminin receptor (67LR) is a nonintegrin cell-surface receptor with high affinity for laminin, which plays a key role in tumor invasion and metastasis. We investigated the role of 67LR in granulocyte colony-stimulating factor (G-CSF)-induced mobilization of CD34+ hematopoietic stem cells (HSCs) from 35 healthy donors. G-CSF-mobilized HSCs, including CD34+/CD38- cells, showed increased 67LR expression as compared with unstimulated marrow HSCs; noteworthy, also, is the fact that the level of 67LR expression in G-CSF-mobilized HSCs correlated significantly with mobilization efficiency. During G-CSF-induced HSC mobilization, the expression of laminin receptors switched from alpha6 integrins, which mediated laminin-dependent adhesion of steady-state human marrow HSCs, to 67LR, responsible for G-CSF-mobilized HSC adhesion and migration toward laminin. In vitro G-CSF treatment, alone or combined with exposure to marrow-derived endothelial cells, induced 67LR up-regulation in marrow HSCs; moreover, anti-67LR antibodies significantly inhibited transendothelial migration of G-CSF-stimulated marrow HSCs. Finally, G-CSF-induced mobilization in mice was associated with 67LR up-regulation both in circulating and marrow CD34+ cells, and anti-67LR antibodies significantly reduced HSC mobilization, providing the first in vivo evidence for 67LR involvement in stem-cell egress from bone marrow after G-CSF administration. In conclusion, 67LR up-regulation in G-CSF-mobilized HSCs correlates with their successful mobilization and reflects its increase in marrow HSCs, which contributes to the egress from bone marrow by mediating laminin-dependent cell adhesion and transendothelial migration.

    Blood 2006;108;7;2476-84

  • 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

  • A trans-dominant negative 37kDa/67kDa laminin receptor mutant impairs PrP(Sc) propagation in scrapie-infected neuronal cells.

    Vana K and Weiss S

    Laboratorium für Molekulare Biologie-Genzentrum-Institut für Biochemie der LMU München, Feodor-Lynen Strasse 25, D-81377 Munich, Germany.

    The 37kDa/67kDa laminin receptor (LRP/LR) has been identified as a cell surface receptor for cellular and infectious prion proteins. Here, we show that an N-terminally truncated LRP mutant encompassing the extracellular domain of the LRP/LR (LRP102-295::FLAG) reduces the binding of recombinant cellular huPrP to mouse neuroblastoma cells, and infectious moPrP27-30 to BHK cells, and interferes with the PrP(Sc) propagation in scrapie-infected neuroblastoma cells (N2aSc(+)). A cell-free binding assay demonstrated the direct binding of the LRP102-295::FLAG mutant to both PrP(c) and PrP(Sc). These results, together with the finding that endogenous LRP levels remain unaffected by the expression of the mutant, indicate that the secreted LRP102-295::FLAG mutant may act in a trans-dominant negative manner as a decoy by trapping PrP molecules. The LRP mutant might represent a potential therapeutic tool for the treatment of TSEs.

    Journal of molecular biology 2006;358;1;57-66

  • BGEM: an in situ hybridization database of gene expression in the embryonic and adult mouse nervous system.

    Magdaleno S, Jensen P, Brumwell CL, Seal A, Lehman K, Asbury A, Cheung T, Cornelius T, Batten DM, Eden C, Norland SM, Rice DS, Dosooye N, Shakya S, Mehta P and Curran T

    Department of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, Tennessee, United States.

    Funded by: NINDS NIH HHS: 5R37NS036558, N01-NS-0-2331, R37 NS036558

    PLoS biology 2006;4;4;e86

  • 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

  • Libraries enriched for alternatively spliced exons reveal splicing patterns in melanocytes and melanomas.

    Watahiki A, Waki K, Hayatsu N, Shiraki T, Kondo S, Nakamura M, Sasaki D, Arakawa T, Kawai J, Harbers M, Hayashizaki Y and Carninci P

    Genome Science Laboratory, RIKEN, Wako main campus, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.

    It is becoming increasingly clear that alternative splicing enables the complex development and homeostasis of higher organisms. To gain a better understanding of how splicing contributes to regulatory pathways, we have developed an alternative splicing library approach for the identification of alternatively spliced exons and their flanking regions by alternative splicing sequence enriched tags sequencing. Here, we have applied our approach to mouse melan-c melanocyte and B16-F10Y melanoma cell lines, in which 5,401 genes were found to be alternatively spliced. These genes include those encoding important regulatory factors such as cyclin D2, Ilk, MAPK12, MAPK14, RAB4, melastatin 1 and previously unidentified splicing events for 436 genes. Real-time PCR further identified cell line-specific exons for Tmc6, Abi1, Sorbs1, Ndel1 and Snx16. Thus, the ASL approach proved effective in identifying splicing events, which suggest that alternative splicing is important in melanoma development.

    Nature methods 2004;1;3;233-9

  • Genomic analysis of mouse retinal development.

    Blackshaw S, Harpavat S, Trimarchi J, Cai L, Huang H, Kuo WP, Weber G, Lee K, Fraioli RE, Cho SH, Yung R, Asch E, Ohno-Machado L, Wong WH and Cepko CL

    Department of Genetics and Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts, USA.

    The vertebrate retina is comprised of seven major cell types that are generated in overlapping but well-defined intervals. To identify genes that might regulate retinal development, gene expression in the developing retina was profiled at multiple time points using serial analysis of gene expression (SAGE). The expression patterns of 1,051 genes that showed developmentally dynamic expression by SAGE were investigated using in situ hybridization. A molecular atlas of gene expression in the developing and mature retina was thereby constructed, along with a taxonomic classification of developmental gene expression patterns. Genes were identified that label both temporal and spatial subsets of mitotic progenitor cells. For each developing and mature major retinal cell type, genes selectively expressed in that cell type were identified. The gene expression profiles of retinal Müller glia and mitotic progenitor cells were found to be highly similar, suggesting that Müller glia might serve to produce multiple retinal cell types under the right conditions. In addition, multiple transcripts that were evolutionarily conserved that did not appear to encode open reading frames of more than 100 amino acids in length ("noncoding RNAs") were found to be dynamically and specifically expressed in developing and mature retinal cell types. Finally, many photoreceptor-enriched genes that mapped to chromosomal intervals containing retinal disease genes were identified. These data serve as a starting point for functional investigations of the roles of these genes in retinal development and physiology.

    Funded by: NCI NIH HHS: P20 CA096470, P20 CA96470; NEI NIH HHS: EY08064, R01 EY008064

    PLoS biology 2004;2;9;E247

  • 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

  • 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

  • Wnk1 kinase deficiency lowers blood pressure in mice: a gene-trap screen to identify potential targets for therapeutic intervention.

    Zambrowicz BP, Abuin A, Ramirez-Solis R, Richter LJ, Piggott J, BeltrandelRio H, Buxton EC, Edwards J, Finch RA, Friddle CJ, Gupta A, Hansen G, Hu Y, Huang W, Jaing C, Key BW, Kipp P, Kohlhauff B, Ma ZQ, Markesich D, Payne R, Potter DG, Qian N, Shaw J, Schrick J, Shi ZZ, Sparks MJ, Van Sligtenhorst I, Vogel P, Walke W, Xu N, Zhu Q, Person C and Sands AT

    Lexicon Genetics, 8800 Technology Forest Place, The Woodlands, TX 77381, USA. brian@lexgen.com

    The availability of both the mouse and human genome sequences allows for the systematic discovery of human gene function through the use of the mouse as a model system. To accelerate the genetic determination of gene function, we have developed a sequence-tagged gene-trap library of >270,000 mouse embryonic stem cell clones representing mutations in approximately 60% of mammalian genes. Through the generation and phenotypic analysis of knockout mice from this resource, we are undertaking a functional screen to identify genes regulating physiological parameters such as blood pressure. As part of this screen, mice deficient for the Wnk1 kinase gene were generated and analyzed. Genetic studies in humans have shown that large intronic deletions in WNK1 lead to its overexpression and are responsible for pseudohypoaldosteronism type II, an autosomal dominant disorder characterized by hypertension, increased renal salt reabsorption, and impaired K+ and H+ excretion. Consistent with the human genetic studies, Wnk1 heterozygous mice displayed a significant decrease in blood pressure. Mice homozygous for the Wnk1 mutation died during embryonic development before day 13 of gestation. These results demonstrate that Wnk1 is a regulator of blood pressure critical for development and illustrate the utility of a functional screen driven by a sequence-based mutagenesis approach.

    Proceedings of the National Academy of Sciences of the United States of America 2003;100;24;14109-14

  • A large-scale, gene-driven mutagenesis approach for the functional analysis of the mouse genome.

    Hansen J, Floss T, Van Sloun P, Füchtbauer EM, Vauti F, Arnold HH, Schnütgen F, Wurst W, von Melchner H and Ruiz P

    Institute of Developmental Genetics, GSF-National Research Center for Environment and Health, D-85764 Neuherberg, Germany.

    A major challenge of the postgenomic era is the functional characterization of every single gene within the mammalian genome. In an effort to address this challenge, we assembled a collection of mutations in mouse embryonic stem (ES) cells, which is the largest publicly accessible collection of such mutations to date. Using four different gene-trap vectors, we generated 5,142 sequences adjacent to the gene-trap integration sites (gene-trap sequence tags; http://genetrap.de) from >11,000 ES cell clones. Although most of the gene-trap vector insertions occurred randomly throughout the genome, we found both vector-independent and vector-specific integration "hot spots." Because >50% of the hot spots were vector-specific, we conclude that the most effective way to saturate the mouse genome with gene-trap insertions is by using a combination of gene-trap vectors. When a random sample of gene-trap integrations was passaged to the germ line, 59% (17 of 29) produced an observable phenotype in transgenic mice, a frequency similar to that achieved by conventional gene targeting. Thus, gene trapping allows a large-scale and cost-effective production of ES cell clones with mutations distributed throughout the genome, a resource likely to accelerate genome annotation and the in vivo modeling of human disease.

    Proceedings of the National Academy of Sciences of the United States of America 2003;100;17;9918-22

  • Genetic imbalances in preleukemic thymuses.

    Verlaet M, Deregowski V, Denis G, Humblet C, Stalmans MT, Bours V, Castronovo V, Boniver J and Defresne MP

    Laboratory of Pathological Anatomy and Cytology, University of Liège, CHU, Liège, B-4000, Belgium. M.Verlaet@ulg.ac.be

    To understand the molecular mechanisms involved in preleukemia, the suppression subtractive hybridization method was used in a murine radiation-induced thymic lymphoma model. Seventeen mRNAs overexpressed in preleukemic thymuses were identified: mouse laminin binding protein (p40/37LBP), E25 protein, Rattus norvegicus clone BB.1.4.1, profilin, poly(A) binding protein (PABP), mouse high mobility group protein 1, topoisomerase I, clusterin, proteasome RC1 subunit, rat prostatein C3 and C1 subunits; two ESTs and four unknown genes. The overexpression of PABP, clusterin, profilin, and the p40/37LBP mRNAs was confirmed in preleukemic thymuses and can be related to some cellular events observed during the preleukemic period, i.e., alterations of cell cycle and apoptosis properties. The p40/37LBP and 67-kDa laminin receptor proteins were upregulated during the preleukemic period. The data suggest that additional studies on p40/37LBP and 67-kDa laminin receptor regulation are required to evaluate their potential role in the lymphoma prevention by TNF-alpha and IFN-gamma.

    Biochemical and biophysical research communications 2001;283;1;12-8

  • Large-scale screen for genes controlling mammalian embryogenesis, using high-throughput gene expression analysis in mouse embryos.

    Neidhardt L, Gasca S, Wertz K, Obermayr F, Worpenberg S, Lehrach H and Herrmann BG

    Max-Planck-Institut für Immunbiologie, Abt. Entwicklungsbiologie, Stübeweg 51, 79108, Freiburg, Germany.

    We have adapted the whole-mount in situ hybridization technique to perform high-throughput gene expression analysis in mouse embryos. A large-scale screen for genes showing specific expression patterns in the mid-gestation embryo was carried out, and a large number of genes controlling development were isolated. From 35760 clones of a 9.5 d.p.c. cDNA library, a total of 5348 cDNAs, enriched for rare transcripts, were selected and analyzed by whole-mount in situ hybridization. Four hundred and twenty-eight clones revealed specific expression patterns in the 9.5 d.p.c. embryo. Of 361 tag-sequenced clones, 198 (55%) represent 154 known mouse genes. Thirty-nine (25%) of the known genes are involved in transcriptional regulation and 33 (21%) in inter- or intracellular signaling. A large number of these genes have been shown to play an important role in embryogenesis. Furthermore, 24 (16%) of the known genes are implicated in human disorders and three others altered in classical mouse mutations. Similar proportions of regulators of embryonic development and candidates for human disorders or mouse mutations are expected among the 163 new mouse genes isolated. Thus, high-throughput gene expression analysis is suitable for isolating regulators of embryonic development on a large-scale, and in the long term, for determining the molecular anatomy of the mouse embryo. This knowledge will provide a basis for the systematic investigation of pattern formation, tissue differentiation and organogenesis in mammals.

    Mechanisms of development 2000;98;1-2;77-94

  • Genome-wide expression profiling of mid-gestation placenta and embryo using a 15,000 mouse developmental cDNA microarray.

    Tanaka TS, Jaradat SA, Lim MK, Kargul GJ, Wang X, Grahovac MJ, Pantano S, Sano Y, Piao Y, Nagaraja R, Doi H, Wood WH, Becker KG and Ko MS

    Laboratory of Genetics and DNA Array Unit, National Institute on Aging, National Institutes of Health, Baltimore, MD 21224-6820, USA.

    cDNA microarray technology has been increasingly used to monitor global gene expression patterns in various tissues and cell types. However, applications to mammalian development have been hampered by the lack of appropriate cDNA collections, particularly for early developmental stages. To overcome this problem, a PCR-based cDNA library construction method was used to derive 52,374 expressed sequence tags from pre- and peri-implantation embryos, embryonic day (E) 12.5 female gonad/mesonephros, and newborn ovary. From these cDNA collections, a microarray representing 15,264 unique genes (78% novel and 22% known) was assembled. In initial applications, the divergence of placental and embryonic gene expression profiles was assessed. At stage E12.5 of development, based on triplicate experiments, 720 genes (6.5%) displayed statistically significant differences in expression between placenta and embryo. Among 289 more highly expressed in placenta, 61 placenta-specific genes encoded, for example, a novel prolactin-like protein. The number of genes highly expressed (and frequently specific) for placenta has thereby been increased 5-fold over the total previously reported, illustrating the potential of the microarrays for tissue-specific gene discovery and analysis of mammalian developmental programs.

    Proceedings of the National Academy of Sciences of the United States of America 2000;97;16;9127-32

  • Identification of novel genes expressed during metanephric induction through single-cell library screening.

    Abidari JM, Gonzales ET, Inoue K, Lupski JR, Karsenty G and Katsanis N

    Scott Department of Urology, Department of Molecular and Human Genetics, and Department of Pediatrics, Baylor College of Medicine, and Texas Children's Hospital, Houston, Texas 77030, USA.

    Background: Development of the mature kidney is dependent on a series of inductive events between a portion of the epithelial bud at the distal end of the nephric duct and a neighboring domain of committed metanephric mesenchyme. Several genes have been identified to date that are critical in the inductive process. For example, the deletion of Bmp7 from the mouse genome results in dysgenesis or agenesis of the kidney. These findings suggest that Bmp7 controls the expression of genes important for nephrogenesis, but the identity of these genes has remained largely undetermined.

    Methods: Single cells were isolated from mouse metanephric mesenchyme during the time of induction (between E11.0 and E11.5) and cDNA libraries constructed from induced and uninduced tissue. Subtractive hybridization was performed to isolate genes that were expressed during E11.5 but not E11.0.

    Results: Using this approach, we identified eight previously known genes, three of which were known to be involved in metanephric induction, thus validating our approach, and nine novel genes. Eight of these genes were completely novel, whereas one was similar to a member of the yeast Anaphase Promoting Complex.

    Conclusions: Through subtractive hybridization of mouse E11.0 and E11.5 metanephric mesenchyme single-cell cDNA libraries, we have identified novel genes that are candidates for involvement in nephrogenesis through their up-regulation during the inductive process.

    Funded by: NIAMS NIH HHS: AR45548

    Kidney international 2000;57;6;2221-8

  • Genome-wide mapping of unselected transcripts from extraembryonic tissue of 7.5-day mouse embryos reveals enrichment in the t-complex and under-representation on the X chromosome.

    Ko MS, Threat TA, Wang X, Horton JH, Cui Y, Wang X, Pryor E, Paris J, Wells-Smith J, Kitchen JR, Rowe LB, Eppig J, Satoh T, Brant L, Fujiwara H, Yotsumoto S and Nakashima H

    Center for Molecular Medicine and Genetics and Department of Internal Medicine, Wayne State University School of Medicine, 5047 Gullen Mall, Detroit, MI 48202, USA. msko@cmb.biosci.wayne.edu

    Mammalian embryos can only survive if they attach to the uterus (implantation) and establish proper maternal-fetal interactions. To understand this complex implantation pathway, we have initiated genomic analysis with a systematic study of the cohort of genes expressed in extraembryonic cells that are derived from the conceptus and play a major role in this process. A total of 2103 cDNAs from the extraembryonic portion of 7.5-day post-conception mouse embryos yielded 3186 expressed sequence tags, approximately 40% of which were novel to the sequence databases. Furthermore, when 155 of the cDNA clones with no homology to previously detected genes were genetically mapped, apparent clustering of these expressed genes was detected in subregions of chromosomes 2, 7, 9 and 17, with 6.5% of the observed genes localized in the t-complex region of chromosome 17, which represents only approximately 1.5% of the mouse genome. In contrast, X-linked genes were under-represented. Semi-quantitative RT-PCR analyses of the mapped genes demonstrated that one third of the genes were expressed solely in extraembryonic tissue and an additional one third of the genes were expressed predominantly in the extraembryonic tissues. The over-representation of extraembryonic-expressed genes in dosage-sensitive autosomal imprinted regions and under-representation on the dosage-compensated X chromosome may reflect a need for tight quantitative control of expression during development.

    Funded by: NHGRI NIH HHS: HG00941; NICHD NIH HHS: HD32243

    Human molecular genetics 1998;7;12;1967-78

  • The 67-kd laminin receptor is preferentially expressed by proliferating retinal vessels in a murine model of ischemic retinopathy.

    Stitt AW, McKenna D, Simpson DA, Gardiner TA, Harriott P, Archer DB and Nelson J

    Department of Ophthalmology, The Queen's University of Belfast, United Kingdom. a.stitt@qub.ac.uk

    Endothelial cell association with vascular basement membranes is complex and plays a critical role in regulation of cell adhesion and proliferation. The interaction between the membrane-associated 67-kd receptor (67LR) and the basement membrane protein laminin has been studied in several cell systems where it was shown to be crucial for adhesion and attachment during angiogenesis. As angiogenesis in the pathological setting of proliferative retinopathy is a major cause of blindness in the Western world we examined the expression of 67LR in a murine model of hyperoxia-induced retinopathy that exhibits retinal neovascularization. Mice exposed to hyperoxia for 5 days starting at postnatal day 7 (P7) and returned to room air (at P12) showed closure of the central retinal vasculature. In response to the ensuing retinal ischemia, there was consistent preretinal neovascularization starting around P17, which persisted until P21, after which the new vessels regressed. Immunohistochemistry was performed on these retinas using an antibody specific for 67LR. At P12, immunoreactivity for 67LR was absent in the retina, but by P17 it was observed in preretinal proliferating vessels and also within the adjacent intraretinal vasculature. Intraretinal 67LR immunoreactivity diminished beyond P17 until by P21 immunoreactivity was almost completely absent, although it persisted in the preretinal vasculature. Control P17 mice (not exposed to hyperoxia) failed to demonstrate any 67LR immunoreactivity in their retinas. Parallel in situ hybridization studies demonstrated 67LR gene expression in the retinal ganglion cells of control and hyperoxia-exposed mice. In addition, the neovascular intra- and preretinal vessels of hyperoxia-treated P17 and P21 mice labeled strongly for 67LR mRNA. This study has characterized 67LR immunolocalization and gene expression in a murine model of ischemic retinopathy. Results suggest that, although the 67LR gene is expressed at high levels in the retinal ganglion cells, the mature receptor protein is preferentially localized to the proliferating retinal vasculature and is almost completely absent from quiescent vessels. The differential expression of 67LR between proliferating and quiescent retinal vessels suggests that this laminin receptor is an important and novel target for future chemotherapeutic intervention during proliferative vasculopathies.

    The American journal of pathology 1998;152;5;1359-65

  • Cloning, chromosomal mapping and expression pattern of the mouse Brca2 gene.

    Connor F, Smith A, Wooster R, Stratton M, Dixon A, Campbell E, Tait TM, Freeman T and Ashworth A

    CRC Centre for Cell and Molecular Biology, Chester Beatty Laboratories, Institute of Cancer Research, London, UK.

    A proportion of human breast cancers result from an inherited predisposition to the disease. Mutations in the BRCA2 gene confer a high risk of breast cancer and are responsible for almost half of these cases. The recent cloning of the human BRCA2 gene has revealed that it encodes a large protein having little significant homology to known proteins. Here we describe the mouse Brca2 gene. The gene maps to mouse chromosome 5, consistent with its location on human chromosome 13q12. We have sequenced cDNA for the entire 3329 amino acid Brca2 protein and this has revealed that, like Brca1, Brca2 is relatively poorly conserved between humans and mice. Brca2 is transcribed in a diverse range of mouse tissues, and the pattern of expression is strikingly similar to that of Brca1. Taken together, our data highlight some intriguing similarities between two genes involved in inherited breast cancer susceptibility.

    Funded by: Wellcome Trust

    Human molecular genetics 1997;6;2;291-300

  • 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

  • Identification of genes differentially expressed in B16 murine melanoma sublines with different metastatic potentials.

    Ishiguro T, Nakajima M, Naito M, Muto T and Tsuruo T

    Laboratory of Biomedical Research, University of Tokyo, Japan.

    B16-F10 and B16-BL6 are B16 mouse melanoma sublines that preferentially metastasize to the lung following i.v. and s.c. injections, respectively. To study molecular mechanisms underlying the different metastatic behaviors exhibited by the B16 melanoma sublines, we performed differential hybridization of the genes transcribed in these cells and compared their expression levels. We isolated four genes that were highly expressed in B16-F10 cells but not in B16-BL6 cells: TI-225 (polyubiquitin), TI-229 (pyruvate kinase), TI-241 (LRF-1 homologue), and TI-227 (novel gene). Triosephosphate isomerase, 10-formyltetrahydrofolate dehydrogenase, tyrosinase-related protein 2, cytochrome c oxidase, ATP synthetase alpha subunit, RNA helicase, and ribosomal protein (L37, J1, acidic phosphoprotein), however, showed higher expression in B16-BL6 cells than in B16-F10 cells. Among these clones, transfection of TI-241 into the low metastatic clone F1 converted the parental cells from low- into high-metastatic cells. TI-241 may regulate the expression of various genes as a transcription factor in the complex process of metastasis.

    Cancer research 1996;56;4;875-9

  • Expression patterns of laminin receptor splice variants alpha 6A beta 1 and alpha 6B beta 1 suggest different roles in mouse development.

    Thorsteinsdóttir S, Roelen BA, Freund E, Gaspar AC, Sonnenberg A and Mummery CL

    Department of Zoology, Faculty of Sciences, University of Lisbon, Portugal.

    The alpha 6 beta 1 integrin is a receptor for laminins and is present from early stages of mouse embryogenesis. In the present study we determined the temporal and spatial expression of the two cytoplasmic splice variants of the alpha 6 integrin subunit, alpha 6A and alpha 6B, in the early- and mid-gestation mouse postimplantation embryo using RT-PCR, in situ hybridization, and immunofluorescence. Our results show that alpha 6B is present in the embryo at all stages studied and is expressed before alpha 6A. alpha 6A expression begins in 8.5 day p.c. embryos and is initially exclusively localized to the developing heart. In 8.5 (and 9.5) day p.c. embryos alpha 6A mRNA and protein are present in a gradient in the myocardium of the heart tube from strongest expression in the sinus venosus and in the common atrial chamber to a weakening expression along the ventricle and bulbus cordis. In 10.5 day p.c. embryos this gradient is less evident and in 12.5 day p.c. embryos alpha 6A mRNA and protein are present in comparable amounts between atria and ventricles. Neither alpha 6A nor alpha 6B is present in endocardial cushion tissue. By day 12.5 p.c. alpha 6A expression is also present in the developing epidermis, dental primordia, lens, gonads, and in a few epithelia such as those of the digestive tract. alpha 6B expression is always much more widespread than alpha 6A expression. For example, only alpha 6B is present in the myotome of the somites of 9.5 day p.c. embryos, in the developing central and peripheral nervous systems, and in the nephrogenic system at all stages studied, except after the differentiation of the gonads when alpha 6A is also present. Furthermore, alpha 6B is the only splice variant present on endothelial cells. We also examined the distribution of the beta 4 integrin subunit to determine whether the alpha 6 beta 4 integrin was present during these stages of development. Beta 4 protein was absent in early postimplantation stages but was present in the epidermis and digestive tract of 12.5 day p.c. embryos. These results show a differential distribution of alpha 6A and alpha 6B during mouse development and thus strongly suggest a different function of these splice variants during embryogenesis. Our results point to a possible role for the alpha 6A beta 1 integrin in the development of the myocardium of the developing heart, but not in the migration of endocardial cushion cells, while alpha 6B beta 1 could be important in the developing nephrogenic and nervous systems.

    Developmental dynamics : an official publication of the American Association of Anatomists 1995;204;3;240-58

  • Isolation of novel tissue-specific genes from cDNA libraries representing the individual tissue constituents of the gastrulating mouse embryo.

    Harrison SM, Dunwoodie SL, Arkell RM, Lehrach H and Beddington RS

    National Institute for Medical Research, London, UK.

    A total of 5 conventional, directionally cloned plasmid cDNA libraries have been constructed from the entire embryonic region of the mid-gastrulation mouse embryo and from its four principal tissue constituents (ectoderm, mesoderm, endoderm and primitive streak). These libraries have been validated with respect to the number of independent clones, insert-size and appropriate representation of diagnostic marker genes. Subtractive hybridisation has been used to remove clones common to the Endoderm and Mesoderm cDNA libraries resulting in an Endoderm minus Mesoderm subtracted library. Probe prepared from this subtracted library has been hybridised to a grid containing approximately 18,500 Embryonic Region library clones. Three novel clones have been recovered as well as expected genes already known to be highly expressed in the primitive endoderm lineage at this stage of development. In situ hybridisation to early postimplantation embryos has revealed the expression patterns of these novel genes. One is highly expressed exclusively in visceral endoderm, one is expressed in ectodermal and endodermal tissues, and the third proves to be an early marker of prospective and differentiated surface ectoderm as well as being expressed in endoderm and its derivatives.

    Development (Cambridge, England) 1995;121;8;2479-89

  • Isolation of nine gene sequences induced by silica in murine macrophages.

    Segade F, Claudio E, Wrobel K, Ramos S and Lazo PS

    Department of Functional Biology, Oviedo University, Spain.

    Macrophage activation by silica is the initial step in the development of silicosis. To identify genes that might be involved in silica-mediated activation, RAW 264.7 mouse macrophages were treated with silica for 48 h, and a subtracted cDNA library enriched for silica-induced genes (SIG) was constructed and differentially screened. Nine cDNA clones (designated SIG-12, -14, -20, -41, -61, -81, -91, -92, and -111) were partially sequenced and compared with sequences in GenBank/EMBL databases. SIG-12, -14, and -20 corresponded to the genes for ribosomal proteins L13a, L32, and L26, respectively. SIG-61 is the mouse homologue of p21 RhoC. SIG-91 is identical to the 67-kDa high-affinity laminin receptor. Four genes were not identified and are novel. All of the mRNAs corresponding to the nine cloned cDNAs were inducible by silica. Steady-state levels of mRNAs in RAW 264.7 cells treated with various macrophage activators and inducers of signal transduction pathways were determined. A complex pattern of induction and repression was found, indicating that upon phagocytosis of silica particles, many regulatory mechanisms of gene expression are simultaneously triggered.

    Journal of immunology (Baltimore, Md. : 1950) 1995;154;5;2384-92

  • Localization of the 110 kDa receptor for laminin in brains of embryonic and postnatal mice.

    Luckenbill-Edds L, Kaiser CA, Rodgers TR and Powell DD

    Ohio University College of Osteopathic Medicine, Department of Biological Sciences, Athens 45701.

    Laminin, a large glycoprotein of the basement membrane that promotes the growth of nerve cell processes in vitro has also been detected in the brains of developing embryos in situ where it is postulated to promote or guide neural outgrowth. We have investigated the histological and developmental patterns of a receptor to a specific pentapeptide sequence in the A chain of the laminin molecule (PA22-2 or IKVAV) that has been identified as a neuron growth-promoting sequence. Standard immunocytochemical procedures were used to localize the receptor by means of a polyclonal antibody to affinity-purified receptor (MR = 110 kDa) from mouse brains. Results for postnatal stages (P) stages (P 1,7,8,25,30,and adult) show that the 110 kDa receptor is localized in fibers in the cortex and hippocampus, in astroglial cells at the surface of the cortex, and in neuronal cell bodies in the hippocampus. In contrast, the A-chain ligand is localized in cell bodies in the same regions at P stages. For embryonic stages (E) (E 14 and E 16) the receptor is localized in bundles of fibers in the superficial and deep cortical layers, and in cell bodies in these regions at E 14 only. Staining for the A chain ligand of the receptor was first seen postnatally. We speculate that the inverse histological pattern of receptor and ligand with respect to cell bodies and fibers may reflect a role in controlling axon guidance during development or repair during regeneration.

    Cell and tissue research 1995;279;2;371-7

  • Covalently immobilized laminin peptide Tyr-Ile-Gly-Ser-Arg (YIGSR) supports cell spreading and co-localization of the 67-kilodalton laminin receptor with alpha-actinin and vinculin.

    Massia SP, Rao SS and Hubbell JA

    Department of Chemical Engineering, University of Texas, Austin 78712-1062.

    The laminin-based nonapeptide Cys-Asp-Pro-Gly-Tyr-Ile-Gly-Ser-Arg (CDPGYIGSR) and pentapeptide Tyr-Ile-Gly-Ser-Arg (YIGSR) have been previously demonstrated to support the attachment of several cell types and to competitively bind to the 67-kDa high affinity laminin receptor. Cell attachment, but not spreading, on substrates containing adsorbed CDPGYIGSR or YIGSR was observed. In this report we describe YIGSR-mediated attachment and spreading of a wide variety of cell types. GYIGSRY promoted cell spreading and stress fiber formation when it was covalently immobilized through the amino-terminal Gly residue, used as a spacer arm. Spreading was not observed when adsorbed YIGSR peptide was used. Functionally blocking antiserum directed against the 67-kDa and related laminin-binding proteins blocked human foreskin fibroblast (HFF) spreading, but not attachment, on covalently grafted GYIGSRY substrates. However, functionally blocking antisera directed against the vitronectin receptor, integrin alpha v beta 3, and the fibronectin receptor, integrin alpha 5 beta 1, did not affect HFF spreading on these substrates. When HFFs spread on these substrates, the 67-kDa laminin receptor co-localized with the cytoplasmic proteins alpha-actinin and vinculin into discrete structures. These results suggest that the adhesion ligand YIGSR is solely sufficient for cell spreading when it is conformationally constrained by covalent attachment to a solid substrate, at least when attached via its amino terminus. Furthermore, the role of the 67-kDa laminin receptor in recognition of this ligand and mediating cell attachment is confirmed in this study. This report also provides the first evidence for direct or indirect association of this receptor with vinculin and alpha-actinin when YIGSR-mediated cell spreading occurs.

    The Journal of biological chemistry 1993;268;11;8053-9

  • Characterization of laminin receptor messenger ribonucleic acid and protein expression in mouse spermatogenic cells.

    Fulcher KD, Welch JE, Davis CM, O'Brien DA and Eddy EM

    Gamete Biology Section, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709.

    A cDNA proposed to encode the mouse laminin receptor (MLR) was isolated from a mouse round spermatid expression library. The cDNA contained complete coding and 3' untranslated regions but was missing the first 42 bases from the 5' untranslated region. Northern blot analysis using a 3' EcoRI fragment of the cDNA identified a 1.2-kb transcript in mouse testes and all somatic tissues tested. Additional transcripts of 1.3 and 0.9 kb were present in round spermatids isolated from mouse testes. Northern blots using ribonuclease (RNase) H-treated poly(A)+ RNA indicated that the difference in the size of 1.3-kb round spermatid transcripts and 1.2-kb transcripts was due to differing poly(A)+ tail lengths. The 0.9-kb round spermatid transcript hybridized to all but the 5' end of the 1.2-kb MLR cDNA, suggesting that an alternate start site is used or that transcript processing occurs in these cells. Immunoblot analysis identified proteins in spermatogenic cells corresponding to the 67-70-kDa MLR and its 43-kDa precursor. In addition, ligand binding studies and affinity chromatography procedures indicated that spermatogenic cell proteins of these sizes bind laminin. However, spermatocytes and spermatids are spatially isolated from laminin in the testes, and MLR may have other functions in these cells.

    Funded by: NICHD NIH HHS: HD-26485, P30-HD-18968

    Biology of reproduction 1993;48;3;674-82

  • Genetic linkage analysis in recombinant inbred mice of P40, a putative clone for the high-affinity laminin receptor.

    Douville PJ and Carbonetto S

    Centre for Research in Neurosciences, Montreal General Hospital, Quebec, Canada.

    Recombinant inbred (RI) mice were used to genetically map sequences of a 43-kDa protein, P40, that was originally identified as a high-affinity laminin receptor. More recent data have implicated this protein in development of the retina (Rabacchi et al. Development 109: 521-531, 1990), possibly via its proposed function in protein translation (G. Brawerman, personal communication). We have identified, in Southern blots, a set of P40-related sequences in BXD recombinant inbred mouse DNA. Ten restriction fragment length polymorphisms (RFLPs) segregate among the RI strains and display strain distribution patterns (SDPs) which are linked in varying degrees to previously typed loci on Chromosomes (Chrs) 1, 3, 6, 9, 11, 14, and 19. An intronic DNA probe from an incompletely processed P40 mRNA transcript overlaps with two of these loci mapping near the cholecystokinin gene locus on the distal arm of Chr 9 and to another site on the distal arm of Chr 6, suggesting that functional genes probably reside at least at these two sites. These P40 loci comprise part of a multimember gene family that is well dispersed in the mouse genome.

    Mammalian genome : official journal of the International Mammalian Genome Society 1992;3;8;438-46

  • Functional domains of the 67-kDa laminin receptor precursor.

    Castronovo V, Taraboletti G and Sobel ME

    Tumor Invasion and Metastasis Section, National Cancer Institute, Bethesda, Maryland 20892.

    We report the characterization of two functional domains of the metastasis-associated 67-kDa laminin receptor (67-LR). Using synthetic peptides deduced from the cDNA sequence of the 37-kDa precursor of the laminin receptor (37-LRP) as well as their corresponding affinity-purified polyclonal antibodies, we identified a unique laminin binding site as well as a membrane-associated domain of the receptor. In laminin dot blot and solid phase radioligand assays, a 20 amino acid synthetic peptide (IPCNNKGAHSVGLMWWMLAR, amino acid residues 161-180, designated peptide G) specifically bound to laminin with high affinity (Kd = 5 x 10(-8) M). Peptide G also specifically eluted the 67-LR from a laminin affinity column. Peptide G and laminin reacted with a 1:1 stoichiometry, suggesting that there is one recognition site on laminin for the peptide G domain. Immunofluorescence studies, performed on permeabilized and nonpermeabilized human A2058 melanoma cells using 10 different affinity-purified antibodies to distinct regions of the 37-LRP, identified an unusually short membrane-associated domain that was consistent with a computer predicted transmembrane domain (residues 86-101). Our data demonstrate for the first time that the 37-LRP has two functional domains consistent with the characteristics of the mature 67-LR. Furthermore, we propose peptide G as a potential inhibitor of tumor cell interactions with laminin.

    The Journal of biological chemistry 1991;266;30;20440-6

  • The high affinity murine laminin receptor is a member of a multicopy gene family.

    Fernández MT, Castronovo V, Rao CN and Sobel ME

    Tumor Invasion and Metastasis Section, National Cancer Institute, Bethesda, Maryland 20892.

    The high affinity laminin receptor is differentially expressed in metastasis. We now report that there are multiple copies (6 +/- 1) of the laminin receptor gene in the murine genome of normal diploid cells as well as in cell lines derived from cancer cells. We have analyzed three distinct cDNA clones isolated from an Okayama-Berg cDNA library of transformed mouse fibroblasts that may represent transcripts of three different laminin receptor genes. Polymorphic changes include insertion of bases at the 5' terminus, a base substitution within the coding region resulting in an amino acid change from phenylalanine to leucine, a base substitution obliterating a polyadenylation signal, as well as changes in the length of the 3' untranslated domains. The discovery of multiple transcripts of laminin receptor genes suggests that there is a strong selective pressure to maintain laminin receptor expression in murine cells.

    Biochemical and biophysical research communications 1991;175;1;84-90

  • A dorso-ventral asymmetry in the embryonic retina defined by protein conformation.

    McCaffery P, Neve RL and Dräger UC

    Department of Neurobiology, Harvard Medical School, Boston, MA 02115.

    In a search for determinants of retinotopic specification we previously identified an antigen in the dorsal embryonic retina as a protein called the 68-kDa laminin receptor. A dorso-ventral asymmetry in a laminin receptor seemed consistent with the known responsiveness of embryonic optic axons to laminin, but there were three peculiar points. (i) The molecular mass of this presumed laminin receptor in immunoblots is not 68 kDa but 43 kDa, and the molecular mass of the protein deduced from the mRNA is only 33 kDa. (ii) The antigen does not have the localization expected of a receptor for the extracellular matrix: the antibodies label mainly a granular cytoplasmic antigen in dorsal retina; an additional sparse cell-surface antigen present on a few cells does not show a dorso-ventral asymmetry. (iii) Despite the pronounced dorso-ventral difference seen immunohistochemically, in immunoblots the 43-kDa protein (p40) is evenly distributed throughout the retina. Here we show that (i) native p40 and in vitro-translated gene product are indistinguishable and their anomalous migration in denaturing gels probably is due to low pI; (ii) p40 is bound in a Mg2(+)-dependent manner to large cytoplasmic complexes that appear to include ribosomes; and (iii) there is a labile conformational difference in p40 between dorsal and ventral retina: dorsally it is more accessible to proteolysis, suggesting a more open conformation. In conjunction with the recent hypothesis that p40 constitutes a translation initiation factor (D. Auth and G. Brawerman, personal communication), these observations point to a dorso-ventral asymmetry in some aspect of protein translation, which in turn may set up differences in recognition factors on retinal growth cones.

    Proceedings of the National Academy of Sciences of the United States of America 1990;87;21;8570-4

  • A positional marker for the dorsal embryonic retina is homologous to the high-affinity laminin receptor.

    Rabacchi SA, Neve RL and Dräger UC

    Department of Neurobiology, Harvard Medical School, Boston, MA 02115.

    In a search for determinants of positional information in the embryonic eye, we isolated two monoclonal antibodies that label strongly the dorsal part of the undifferentiated embryonic retina in mammals, bird and cold-blooded vertebrates. In the chick, the optic tectum is labeled in a corresponding fashion, the ventral tectum more heavily than the dorsal tectum. Through biochemical and molecular analysis both antibodies were found to recognize a protein that has been cloned repeatedly, first in a screen with antibodies to the '68K-laminin receptor' (Wewer et al. (1986) Cancer Res. 47, 5691-5698), a name that may not exhaustively describe its function. Western blots show the protein to be present in most or all tissues, and Western and Southern blots reveal a high degree of conservation in the detected signals up to invertebrates and bacteria. Despite the very strong and selective labeling of the dorsal retina in conventional immunohistochemical preparations, the protein and its mRNA are present in even amounts throughout the embryonic retina, as demonstrated by Western and Northern blots of bisected retinas, and immunohistochemically in retinas fixed with ethylene glycole bissuccinimide (EGS), an NH2-group crosslinker with very long spacer arm. This indicates that the dorsoventral asymmetry in the embryonic retina is not in the amount but in the configuration of this protein; whether this difference relates to laminin binding is not known.

    Development (Cambridge, England) 1990;109;3;521-31

  • Evidence for a precursor of the high-affinity metastasis-associated murine laminin receptor.

    Rao CN, Castronovo V, Schmitt MC, Wewer UM, Claysmith AP, Liotta LA and Sobel ME

    Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland 20892.

    The high-affinity cellular receptor for the basement membrane component laminin is differentially expressed during tumor invasion and metastasis. A cDNA clone encoding the murine laminin receptor was isolated and identified on the basis of sequence homology to the human laminin receptor [Wewer et al. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 7137-7141]. Primer extension experiments demonstrated that the clone contained the complete 5' sequence of the murine laminin receptor mRNA. RNA blot data demonstrated a single-sized laminin receptor mRNA, approximately 1400 bases long, in human, mouse, and rat. The nascent laminin receptor predicted from the cDNA sequence is 295 amino acids long, with a molecular weight of 33,000, and contains one intradisulfide bridge, a short putative transmembrane domain, and an extracellular carboxy-terminal region which has abundant glutamic acid residues and multiple repeat sequences. The precursor of the laminin receptor is apparently smaller than the 67-kilodalton protein isolated from tissue. The apparent molecular weight on SDS-polyacrylamide gels of the rabbit reticulocyte cell-free translation product of selectively hybridized laminin receptor mRNA is 37,000. Antisera to three different domains of the cDNA-predicted receptor were used to study the relationship between the 37- and 67-kilodalton polypeptides. Antisera to cDNA-deduced synthetic peptides of the receptor immunoprecipitated a 37-kilodalton band both from cell-free translation products and from pulse-labeled cell extracts. On immunoblots of cell extracts, one antisynthetic peptide antiserum recognized only the 67-kilodalton receptor, while another antiserum identified both 37- and 67-kilodalton polypeptides, suggesting a precursor-product relationship between the two polypeptides.

    Biochemistry 1989;28;18;7476-86

  • Nucleotide sequence for a major messenger RNA for a 40 kilodalton polypeptide that is under translational control in mouse tumor cells.

    Makrides S, Chitpatima ST, Bandyopadhyay R and Brawerman G

    Department of Biochemistry, Tufts University Health Sciences Schools, Boston, MA 02111.

    Funded by: NIGMS NIH HHS: GM17973

    Nucleic acids research 1988;16;5;2349

Gene lists (3)

Gene List Source Species Name Description Gene count
L00000001 G2C Mus musculus Mouse PSD Mouse PSD adapted from Collins et al (2006) 1080
L00000008 G2C Mus musculus Mouse PSP Mouse PSP adapted from Collins et al (2006) 1121
L00000070 G2C Mus musculus BAYES-COLLINS-HUMAN-PSD-FULL Human cortex biopsy PSD full list (ortho) 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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