G2Cdb::Gene report

Gene id
G00000298
Gene symbol
Atp6v1b2 (MGI)
Species
Mus musculus
Description
ATPase, H+ transporting, lysosomal V1 subunit B2
Orthologue
G00001547 (Homo sapiens)

Databases (9)

Gene
ENSMUSG00000006273 (Ensembl mouse gene)
11966 (Entrez Gene)
634 (G2Cdb plasticity & disease)
Gene Expression
NM_007509 (Allen Brain Atlas)
11966 (Genepaint)
atp6v1b2 (gensat)
Literature
606939 (OMIM)
Marker Symbol
MGI:109618 (MGI)
Protein Sequence
P62814 (UniProt)

Synonyms (2)

  • Atp6b2
  • HO57

Literature (22)

Pubmed - other

  • A high-resolution anatomical atlas of the transcriptome in the mouse embryo.

    Diez-Roux G, Banfi S, Sultan M, Geffers L, Anand S, Rozado D, Magen A, Canidio E, Pagani M, Peluso I, Lin-Marq N, Koch M, Bilio M, Cantiello I, Verde R, De Masi C, Bianchi SA, Cicchini J, Perroud E, Mehmeti S, Dagand E, Schrinner S, Nürnberger A, Schmidt K, Metz K, Zwingmann C, Brieske N, Springer C, Hernandez AM, Herzog S, Grabbe F, Sieverding C, Fischer B, Schrader K, Brockmeyer M, Dettmer S, Helbig C, Alunni V, Battaini MA, Mura C, Henrichsen CN, Garcia-Lopez R, Echevarria D, Puelles E, Garcia-Calero E, Kruse S, Uhr M, Kauck C, Feng G, Milyaev N, Ong CK, Kumar L, Lam M, Semple CA, Gyenesei A, Mundlos S, Radelof U, Lehrach H, Sarmientos P, Reymond A, Davidson DR, Dollé P, Antonarakis SE, Yaspo ML, Martinez S, Baldock RA, Eichele G and Ballabio A

    Telethon Institute of Genetics and Medicine, Naples, Italy.

    Ascertaining when and where genes are expressed is of crucial importance to understanding or predicting the physiological role of genes and proteins and how they interact to form the complex networks that underlie organ development and function. It is, therefore, crucial to determine on a genome-wide level, the spatio-temporal gene expression profiles at cellular resolution. This information is provided by colorimetric RNA in situ hybridization that can elucidate expression of genes in their native context and does so at cellular resolution. We generated what is to our knowledge the first genome-wide transcriptome atlas by RNA in situ hybridization of an entire mammalian organism, the developing mouse at embryonic day 14.5. This digital transcriptome atlas, the Eurexpress atlas (http://www.eurexpress.org), consists of a searchable database of annotated images that can be interactively viewed. We generated anatomy-based expression profiles for over 18,000 coding genes and over 400 microRNAs. We identified 1,002 tissue-specific genes that are a source of novel tissue-specific markers for 37 different anatomical structures. The quality and the resolution of the data revealed novel molecular domains for several developing structures, such as the telencephalon, a novel organization for the hypothalamus, and insight on the Wnt network involved in renal epithelial differentiation during kidney development. The digital transcriptome atlas is a powerful resource to determine co-expression of genes, to identify cell populations and lineages, and to identify functional associations between genes relevant to development and disease.

    Funded by: Medical Research Council: MC_U127527203; Telethon: TGM11S03

    PLoS biology 2011;9;1;e1000582

  • Inhibition of osteoclast bone resorption by disrupting vacuolar H+-ATPase a3-B2 subunit interaction.

    Kartner N, Yao Y, Li K, Crasto GJ, Datti A and Manolson MF

    Dental Research Institute, Faculty of Dentistry, University of Toronto, Toronto, Ontario M5G 1G6 Canada.

    Vacuolar H(+)-ATPases (V-ATPases) are highly expressed in ruffled borders of bone-resorbing osteoclasts, where they play a crucial role in skeletal remodeling. To discover protein-protein interactions with the a subunit in mammalian V-ATPases, a GAL4 activation domain fusion library was constructed from an in vitro osteoclast model, receptor activator of NF-κB ligand-differentiated RAW 264.7 cells. This library was screened with a bait construct consisting of a GAL4 binding domain fused to the N-terminal domain of V-ATPase a3 subunit (NTa3), the a subunit isoform that is highly expressed in osteoclasts (a1 and a2 are also expressed, to a lesser degree, whereas a4 is kidney-specific). One of the prey proteins identified was the V-ATPase B2 subunit, which is also highly expressed in osteoclasts (B1 is not expressed). Further characterization, using pulldown and solid-phase binding assays, revealed an interaction between NTa3 and the C-terminal domains of both B1 and B2 subunits. Dual B binding domains of equal affinity were observed in NTa, suggesting a possible model for interaction between these subunits in the V-ATPase complex. Furthermore, the a3-B2 interaction appeared to be moderately favored over a1, a2, and a4 interactions with B2, suggesting a mechanism for the specific subunit assembly of plasma membrane V-ATPase in osteoclasts. Solid-phase binding assays were subsequently used to screen a chemical library for inhibitors of the a3-B2 interaction. A small molecule benzohydrazide derivative was found to inhibit osteoclast resorption with an IC(50) of ∼1.2 μm on both synthetic hydroxyapatite surfaces and dentin slices, without significantly affecting RAW 264.7 cell viability or receptor activator of NF-κB ligand-mediated osteoclast differentiation. Further understanding of these interactions and inhibitors may contribute to the design of novel therapeutics for bone loss disorders, such as osteoporosis and rheumatoid arthritis.

    Funded by: Canadian Institutes of Health Research: MOP-79322, PDD-86132

    The Journal of biological chemistry 2010;285;48;37476-90

  • Optic nerve compression and retinal degeneration in Tcirg1 mutant mice lacking the vacuolar-type H-ATPase a3 subunit.

    Kawamura N, Tabata H, Sun-Wada GH and Wada Y

    Department of Biochemistry, Faculty of Pharmaceutical Sciences, Doshisha Women's College, Kyotanabe, Japan.

    Background: Vacuolar-type proton transporting ATPase (V-ATPase) is involved in the proper development of visual function. Mutations in the Tcirg1 (also known as Atp6V0a3) locus, which encodes the a3 subunit of V-ATPase, cause severe autosomal recessive osteopetrosis (ARO) in humans. ARO is often associated with impaired vision most likely because of nerve compression at the optic canal. We examined the ocular phenotype of mice deficient in Tcirg1 function.

    X-ray microtomography showed narrowed foramina in the skull, suggesting that optic nerve compression occurred in the a3-deficient (Tcirg1-/-) mice. The retina of the mutant mice had normal architecture, but the number of apoptotic cells was increased at 2-3 wks after birth. In the ocular system, the a3 subunit accumulated in the choriocapillary meshwork in uveal tissues. Two other subunit isoforms a1 and a2 accumulated in the retinal photoreceptor layer. We found that the a4 subunit, whose expression has previously been shown to be restricted to several transporting epithelia, was enriched in pigmented epithelial cells of the retina and ciliary bodies. The expression of a4 in the uveal tissue was below the level of detection in wild-type mice, but it was increased in the mutant choriocapillary meshwork, suggesting that compensation may have occurred among the a subunit isoforms in the mutant tissues.

    Conclusions: Our findings suggest that a similar etiology of visual impairment is involved in both humans and mice; thus, a3-deficient mice may provide a suitable model for clinical and diagnostic purposes in cases of ARO.

    PloS one 2010;5;8;e12086

  • Lack of periostin leads to suppression of Notch1 signaling and calcific aortic valve disease.

    Tkatchenko TV, Moreno-Rodriguez RA, Conway SJ, Molkentin JD, Markwald RR and Tkatchenko AV

    Department of Anatomy and Cell Biology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.

    The Postn gene encodes protein periostin. During embryonic development, it is highly expressed in the outflow tract (OFT) endocardial cushions of the developing heart, which give rise to several structures of the mature heart including the aortic valve. Periostin was previously implicated in osteoblast differentiation, cancer metastasis, and tooth and bone development, but its role in cardiac OFT development is unclear. To elucidate the role that periostin plays in the developing heart we analyzed cardiac OFT phenotype in mice after deletion of the Postn gene. We found that lack of periostin in the embryonic OFT leads to ectopic expression of the proosteogenic growth factor pleiotrophin (Ptn) and overexpression of delta-like 1 homolog (Dlk1), a negative regulator of Notch1, in the distal (prevalvular) cushions of the OFT. This resulted in suppression of Notch1 signaling, strong induction of the central transcriptional regulator of osteoblast cell fate Runx2, upregulation of osteopontin and osteocalcin expression, and subsequent calcification of the aortic valve. Our data suggest that periostin represses a default osteogenic program in the OFT cushion mesenchyme and promotes differentiation along a fibrogenic lineage. Lack of periostin causes derepression of the osteogenic potential of OFT mesenchymal cells, calcium deposition, and calcific aortic valve disease. These results establish periostin as a key regulator of OFT endocardial cushion mesenchymal cell fate during embryonic development.

    Funded by: NCRR NIH HHS: 2P20-RR-016434; NHLBI NIH HHS: HL-077342, HL-33756, R01 HL092508, R01 HL092508-01A1

    Physiological genomics 2009;39;3;160-8

  • Circadian clock and output genes are rhythmically expressed in extratesticular ducts and accessory organs of mice.

    Bebas P, Goodall CP, Majewska M, Neumann A, Giebultowicz JM and Chappell PE

    Department of Animal Physiology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

    Circadian clocks regulate multiple rhythms in mammalian tissues. In most organs core clock gene expression is oscillatory, with negative components Per and Cry peaking in antiphase to Bmal1. A notable exception is the testis, where clock genes seem nonrhythmic. Earlier mammalian studies, however, did not examine clock expression patterns in accessory ductal tissue required for sperm maturation and transport. Previous studies in insects demonstrated control of sperm maturation in vas deferens by a local circadian system. Sperm ducts express clock genes and display circadian pH changes controlled by vacuolar-type H(+)-ATPase and carbonic anhydrase (CA-II). It is unknown whether sperm-processing rhythms are conserved beyond insects. To address this question in mice housed in a light-dark environment, we examined temporal patterns of mPer1 and Bmal1 gene expression and protein abundance in epididymis, vas deferens, seminal vesicles, and prostate. Results demonstrate variable tissue-specific patterns of expression of the two genes, with variations in levels of clock proteins and their nucleo-cytoplasmic cycling observed among examined tissues. Strikingly, mPer1 and Bmal1 mRNA and proteins oscillate in antiphase in the prostate, with similar peak-trough patterns as observed in the suprachiasmatic nuclei, the brain's central clock. Genes encoding CA and a V-ATPase subunit, which are rhythmically expressed in sperm ducts of moths, are also rhythmic in some segments of murine sperm ducts. Our data suggest that some sperm duct segments may contain peripheral circadian systems whereas others may express clock genes in a pleiotropic manner.

    Funded by: NIDDK NIH HHS: K01 DK064919, R01 DK075357; NIGMS NIH HHS: GM073792, R01 GM073792

    FASEB journal : official publication of the Federation of American Societies for Experimental Biology 2009;23;2;523-33

  • Proteomics analysis identifies phosphorylation-dependent alpha-synuclein protein interactions.

    McFarland MA, Ellis CE, Markey SP and Nussbaum RL

    National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland 20891, USA.

    Mutations and copy number variation in the SNCA gene encoding the neuronal protein alpha-synuclein have been linked to familial Parkinson disease (Thomas, B., and Beal, M. F. (2007) Parkinson's disease. Hum. Mol. Genet. 16, R183-R194). The carboxyl terminus of alpha-synuclein can be phosphorylated at tyrosine 125 and serine 129, although only a small fraction of the protein is phosphorylated under normal conditions (Okochi, M., Walter, J., Koyama, A., Nakajo, S., Baba, M., Iwatsubo, T., Meijer, L., Kahle, P. J., and Haass, C. (2000) Constitutive phosphorylation of the Parkinson's disease associated alpha-synuclein. J. Biol. Chem. 275, 390-397). Under pathological conditions, such as in Parkinson disease, alpha-synuclein is a major component of Lewy bodies, a pathological hallmark of Parkinson disease, and is mostly phosphorylated at Ser-129 (Anderson, J. P., Walker, D. E., Goldstein, J. M., de Laat, R., Banducci, K., Caccavello, R. J., Barbour, R., Huang, J. P., Kling, K., Lee, M., Diep, L., Keim, P. S., Shen, X. F., Chataway, T., Schlossmacher, M. G., Seubert, P., Schenk, D., Sinha, S., Gai, W. P., and Chilcote, T. J. (2006) Phosphorylation of Ser-129 is the dominant pathological modification of alpha-synuclein in familial and sporadic Lewy body disease. J. Biol. Chem. 281, 29739-29752). Controversy exists over the extent to which phosphorylation of alpha-synuclein and/or the visible protein aggregation in Lewy bodies are steps in disease pathogenesis, are protective, or are neutral markers for the disease process. Here we used the combination of peptide pulldown assays and mass spectrometry to identify and compare protein-protein interactions of phosphorylated and non-phosphorylated alpha-synuclein. We showed that non-phosphorylated alpha-synuclein carboxyl terminus pulled down protein complexes that were highly enriched for mitochondrial electron transport proteins, whereas alpha-synuclein carboxyl terminus phosphorylated on either Ser-129 or Tyr-125 did not. Instead the set of proteins pulled down by phosphorylated alpha-synuclein was highly enriched in certain cytoskeletal proteins, in vesicular trafficking proteins, and in a small number of enzymes involved in protein serine phosphorylation. This targeted comparative proteomics approach for unbiased identification of protein-protein interactions suggests that there are functional consequences when alpha-synuclein is phosphorylated.

    Funded by: Intramural NIH HHS; NIMH NIH HHS: Z01 MH000279

    Molecular & cellular proteomics : MCP 2008;7;11;2123-37

  • V-ATPase expression in the mouse olfactory epithelium.

    Paunescu TG, Jones AC, Tyszkowski R and Brown D

    Center for Systems Biology, Program in Membrane Biology and Division of Nephrology, Massachusetts General Hospital, Massachusetts 02114, USA. paunescu@receptor.mgh.harvard.edu

    The vacuolar proton-pumping ATPase (V-ATPase) is responsible for the acidification of intracellular organelles and for the pH regulation of extracellular compartments. Because of the potential role of the latter process in olfaction, we examined the expression of V-ATPase in mouse olfactory epithelial (OE) cells. We report that V-ATPase is present in this epithelium, where we detected subunits ATP6V1A (the 70-kDa "A" subunit) and ATP6V1E1 (the ubiquitous 31-kDa "E" subunit isoform) in epithelial cells, nerve fiber cells, and Bowman's glands by immunocytochemistry. We also located both isoforms of the 56-kDa B subunit, ATP6V1B1 ("B1," typically expressed in epithelia specialized in regulated transepithelial proton transport) and ATP6V1B2 ("B2") in the OE. B1 localizes to the microvilli of the apical plasma membrane of sustentacular cells and to the lateral membrane in a subset of olfactory sensory cells, which also express carbonic anhydrase type IV, whereas B2 expression is stronger in the subapical domain of sustentacular cells. V-ATPase expression in mouse OE was further confirmed by immunoblotting. These findings suggest that V-ATPase may be involved in proton secretion in the OE and, as such, may be important for the pH homeostasis of the neuroepithelial mucous layer and/or for signal transduction in CO(2) detection.

    Funded by: NIDDK NIH HHS: DK-42956, DK-43351, DK-57521, DK-73266; PHS HHS: 5U54A1057159-03

    American journal of physiology. Cell physiology 2008;295;4;C923-30

  • New role of bone morphogenetic protein 7 in brown adipogenesis and energy expenditure.

    Tseng YH, Kokkotou E, Schulz TJ, Huang TL, Winnay JN, Taniguchi CM, Tran TT, Suzuki R, Espinoza DO, Yamamoto Y, Ahrens MJ, Dudley AT, Norris AW, Kulkarni RN and Kahn CR

    Section on Obesity and Hormone Action, Joslin Diabetes Center, Harvard Medical School, Boston, Massachusetts 02215, USA. yu-hua.tseng@joslin.harvard.edu

    Adipose tissue is central to the regulation of energy balance. Two functionally different types of fat are present in mammals: white adipose tissue, the primary site of triglyceride storage, and brown adipose tissue, which is specialized in energy expenditure and can counteract obesity. Factors that specify the developmental fate and function of white and brown adipose tissue remain poorly understood. Here we demonstrate that whereas some members of the family of bone morphogenetic proteins (BMPs) support white adipocyte differentiation, BMP7 singularly promotes differentiation of brown preadipocytes even in the absence of the normally required hormonal induction cocktail. BMP7 activates a full program of brown adipogenesis including induction of early regulators of brown fat fate PRDM16 (PR-domain-containing 16; ref. 4) and PGC-1alpha (peroxisome proliferator-activated receptor-gamma (PPARgamma) coactivator-1alpha; ref. 5), increased expression of the brown-fat-defining marker uncoupling protein 1 (UCP1) and adipogenic transcription factors PPARgamma and CCAAT/enhancer-binding proteins (C/EBPs), and induction of mitochondrial biogenesis via p38 mitogen-activated protein (MAP) kinase-(also known as Mapk14) and PGC-1-dependent pathways. Moreover, BMP7 triggers commitment of mesenchymal progenitor cells to a brown adipocyte lineage, and implantation of these cells into nude mice results in development of adipose tissue containing mostly brown adipocytes. Bmp7 knockout embryos show a marked paucity of brown fat and an almost complete absence of UCP1. Adenoviral-mediated expression of BMP7 in mice results in a significant increase in brown, but not white, fat mass and leads to an increase in energy expenditure and a reduction in weight gain. These data reveal an important role of BMP7 in promoting brown adipocyte differentiation and thermogenesis in vivo and in vitro, and provide a potential new therapeutic approach for the treatment of obesity.

    Funded by: NIDDK NIH HHS: K08 DK064906, K08 DK64906, P30 DK040561, P30 DK040561-13, P30 DK046200, P30 DK46200, R01 DK 060837, R01 DK060837, R01 DK067536, R01 DK077097, R01 DK077097-01A1, R01 DK077097-02, R01 DK67536, R21 DK070722, R21 DK070722-01, R21 DK070722-02

    Nature 2008;454;7207;1000-4

  • Tissue specific expression of the splice variants of the mouse vacuolar proton-translocating ATPase a4 subunit.

    Kawasaki-Nishi S, Yamaguchi A, Forgac M and Nishi T

    Department of Cell Membrane Biology, ISIR, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka, Japan.

    We have identified splicing variants of the mouse a4 subunit which have the same open reading frame but have a different 5'-noncoding sequence. Further determination of the 5'-upstream region of the a4 gene in mouse indicated the presence of two first exons (exon 1a and exon 1b) which include the 5'-noncoding sequence of each variant. The mRNAs of both splicing variants (a4-I and a4-II) show a similar expression pattern in mouse kidney by in situ hybridization. However, tissue and developmental expression patterns of the variants are different. In addition to strong expression in kidney, a4-I expression was detected in heart, lung, skeletal muscle, and testis, whereas a4-II is expressed in lung, liver, and testis. During development, a4-I was expressed beginning with the early embryonic stage, but a4-II mRNA was detected from day 17. These results suggest that each a4 variant has both a tissue and developmental stage specific function.

    Funded by: NIGMS NIH HHS: GM34478, R01 GM034478, R01 GM034478-15, R01 GM034478-16, R37 GM034478, R37 GM034478-17, R37 GM034478-18, R37 GM034478-19

    Biochemical and biophysical research communications 2007;364;4;1032-6

  • Compensatory membrane expression of the V-ATPase B2 subunit isoform in renal medullary intercalated cells of B1-deficient mice.

    Paunescu TG, Russo LM, Da Silva N, Kovacikova J, Mohebbi N, Van Hoek AN, McKee M, Wagner CA, Breton S and Brown D

    Div. of Nephrology, Massachusetts General Hospital, 185 Cambridge St., CPZN 8150, Boston, MA 02114, USA. paunescu@receptor.mgh.harvard.edu

    Mice deficient in the ATP6V1B1 ("B1") subunit of the vacuolar proton-pumping ATPase (V-ATPase) maintain body acid-base homeostasis under normal conditions, but not when exposed to an acid load. Here, compensatory mechanisms involving the alternate ATP6V1B2 ("B2") isoform were examined to explain the persistence of baseline pH regulation in these animals. By immunocytochemistry, the mean pixel intensity of apical B2 immunostaining in medullary A intercalated cells (A-ICs) was twofold greater in B1-/- mice than in B1+/+ animals, and B2 was colocalized with other V-ATPase subunits. No significant upregulation of B2 mRNA or protein expression was detected in B1-/- mice compared with wild-type controls. We conclude that increased apical B2 staining is due to relocalization of B2-containing V-ATPase complexes from the cytosol to the plasma membrane. Recycling of B2-containing holoenzymes between these domains was confirmed by the intracellular accumulation of B1-deficient V-ATPases in response to the microtubule-disrupting drug colchicine. V-ATPase membrane expression is further supported by the presence of "rod-shaped" intramembranous particles seen by freeze fracture microscopy in apical membranes of normal and B1-deficient A-ICs. Intracellular pH recovery assays show that significant (28-40% of normal) V-ATPase function is preserved in medullary ICs from B1-/- mice. We conclude that the activity of apical B2-containing V-ATPase holoenzymes in A-ICs is sufficient to maintain baseline acid-base homeostasis in B1-deficient mice. However, our results show no increase in cell surface V-ATPase activity in response to metabolic acidosis in ICs from these animals, consistent with their inability to appropriately acidify their urine under these conditions.

    Funded by: NICHD NIH HHS: HD-40793; NIDDK NIH HHS: DK-38452, DK-42956, DK-43266

    American journal of physiology. Renal physiology 2007;293;6;F1915-26

  • Relocalization of the V-ATPase B2 subunit to the apical membrane of epididymal clear cells of mice deficient in the B1 subunit.

    Da Silva N, Shum WW, El-Annan J, Păunescu TG, McKee M, Smith PJ, Brown D and Breton S

    Massachusetts General Hospital, Harvard Medical School, Program in Membrane Biology, Nephrology Div., 185 Cambridge St., CPZN 8150, Boston, MA 02114-2790, USA. ndasilva@partners.org

    An acidic luminal pH in the epididymis contributes to maintaining sperm quiescent during their maturation and storage. The vacuolar H(+)ATPase (V-ATPase), located in narrow and clear cells, is a major contributor to luminal acidification. Mutations in one of the V-ATPase subunits, ATP6v1B1 (B1), cause distal renal tubular acidosis in humans but surprisingly, B1(-/-) mice do not develop metabolic acidosis and are fertile. While B1 is located in the apical membrane of narrow and clear cells, the B2 subunit localizes to subapical vesicles in wild-type mouse, rat and human epididymis. However, a marked increase (84%) in the mean pixel intensity of B2 staining was observed in the apical pole of clear cells by conventional immunofluorescence, and relocalization into their apical membrane was detected by confocal microscopy in B1(-/-) mice compared with B1(+/+). Immunogold electron microscopy showed abundant B2 in the apical microvilli of clear cells in B1(-/-) mice. B2 mRNA expression, determined by real time RT-PCR using laser-microdissected epithelial cells, was identical in both groups. Semiquantitative Western blots from whole epididymis and cauda epididymidis showed no variation of B2 expression. Finally, the luminal pH of the cauda epididymidis was the same in B1(-/-) mice as in B1(+/+) (pH 6.7). These data indicate that whereas overall expression of B2 is not affected in B1(-/-) mice, significant redistribution of B2-containing complexes occurs from intracellular compartments into the apical membrane of clear cells in B1(-/-) mice. This relocation compensates for the absence of functional B1 and maintains the luminal pH in an acidic range that is compatible with fertility.

    Funded by: NICHD NIH HHS: HD-40793; NIDDK NIH HHS: DK-38452, DK-43351, DK-57521, DK-73266

    American journal of physiology. Cell physiology 2007;293;1;C199-210

  • Qualitative and quantitative analyses of protein phosphorylation in naive and stimulated mouse synaptosomal preparations.

    Munton RP, Tweedie-Cullen R, Livingstone-Zatchej M, Weinandy F, Waidelich M, Longo D, Gehrig P, Potthast F, Rutishauser D, Gerrits B, Panse C, Schlapbach R and Mansuy IM

    Brain Research Institute, Medical Faculty of the University of Zürich, Switzerland.

    Activity-dependent protein phosphorylation is a highly dynamic yet tightly regulated process essential for cellular signaling. Although recognized as critical for neuronal functions, the extent and stoichiometry of phosphorylation in brain cells remain undetermined. In this study, we resolved activity-dependent changes in phosphorylation stoichiometry at specific sites in distinct subcellular compartments of brain cells. Following highly sensitive phosphopeptide enrichment using immobilized metal affinity chromatography and mass spectrometry, we isolated and identified 974 unique phosphorylation sites on 499 proteins, many of which are novel. To further explore the significance of specific phosphorylation sites, we used isobaric peptide labels and determined the absolute quantity of both phosphorylated and non-phosphorylated peptides of candidate phosphoproteins and estimated phosphorylation stoichiometry. The analyses of phosphorylation dynamics using differentially stimulated synaptic terminal preparations revealed activity-dependent changes in phosphorylation stoichiometry of target proteins. Using this method, we were able to differentiate between distinct isoforms of Ca2+/calmodulin-dependent protein kinase (CaMKII) and identify a novel activity-regulated phosphorylation site on the glutamate receptor subunit GluR1. Together these data illustrate that mass spectrometry-based methods can be used to determine activity-dependent changes in phosphorylation stoichiometry on candidate phosphopeptides following large scale phosphoproteome analysis of brain tissue.

    Molecular & cellular proteomics : MCP 2007;6;2;283-93

  • Ubiquitous and kidney-specific subunits of vacuolar H+-ATPase are differentially expressed during nephrogenesis.

    Jouret F, Auzanneau C, Debaix H, Wada GH, Pretto C, Marbaix E, Karet FE, Courtoy PJ and Devuyst O

    Division of Nephrology, Université catholique de Louvain, 10 Avenue Hippocrate, Brussels, Belgium B-1200.

    The vacuolar H(+)-ATPase (V-ATPase) is a ubiquitous multisubunit pump that is responsible for acidification of intracellular organelles. In the kidney, a particular form of V-ATPase, made of specific subunits isoforms, has been located at the plasma membrane of intercalated cells (IC). Mutations in genes encoding IC-specific subunits cause infant distal renal tubular acidosis (dRTA), suggesting that the segmental distribution of these subunits is acquired at birth or during early infancy. However, the comparative ontogeny of the IC-specific versus the ubiquitous subunits of V-ATPase and the mechanisms involved in their segmental expression remain unknown. Real-time reverse transcription-PCR, in situ hybridization, immunoblotting, immunostaining, and subcellular fractionation analyses characterized the expression and distribution of V-ATPase subunits, transcription factors, and differentiation markers during mouse nephrogenesis. Ubiquitous A, E1, B2, G1, and C1 subunits showed an early (embryonic day 13.5 [E13.5]) and stable expression throughout nephrogenesis, followed by a slight increase around birth. The developmental pattern of a1 was bimodal, with early induction, gradual decrease during organogenesis, and neonatal increase. These patterns contrasted with the later (from E15.5) and progressive expression of IC-specific a4, B1, G3, and C2 subunits, after the induction of the forkhead transcription factor Foxi1. From E15.5, Foxi1 mRNA was detected in IC, where it co-distributed with B1 in late nephrogenesis. Immunostaining showed that the distribution of ubiquitous E1 and B2 was acquired from E15.5, whereas a4 was located in IC during late nephrogenesis. Subcellular fractionation showed that in both fetal and mature (cortex and medulla) kidneys, E1 and a4 were located in endosomes. These data demonstrate a differential expression and a coordinate regulation of IC-specific versus ubiquitous V-ATPase subunits during nephrogenesis. They provide new insights into the complex regulation of V-ATPase subunits, the maturation of IC along the nephron, and the pathophysiology of hereditary dRTA.

    Journal of the American Society of Nephrology : JASN 2005;16;11;3235-46

  • 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

  • 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

  • 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

  • Expression of the 56-kDa B2 subunit isoform of the vacuolar H(+)-ATPase in proton-secreting cells of the kidney and epididymis.

    Paunescu TG, Da Silva N, Marshansky V, McKee M, Breton S and Brown D

    Program in Membrane Biology/Renal Unit, Massachusetts General Hospital, Charlestown, MA 02129, USA. paunescu@receptor.mgh.harvard.edu

    B1 and B2 are two highly homologous isoforms of the vacuolar H(+)-ATPase (V-ATPase) 56-kDa B subunit. We investigated whether the B2 subunit is expressed alongside B1 in proton-secreting cells of the rodent kidney collecting duct (intercalated cells, IC) and epididymis (clear cells) by using antibodies against distinct COOH-terminal peptides from the two B isoforms. B2 was detected not only in the kidney proximal tubule, thick ascending limb, distal convoluted tubule, and connecting segment but also in A- and B-type IC of collecting ducts (CD) in both rat and mouse. B2 had a predominant cytoplasmic localization in most IC but was clearly located in a tighter apical band together with the V-ATPase 31-kDa E subunit in some A-IC, especially in the medulla. Apical membrane staining was confirmed by immunogold electron microscopy. B2 was very weakly expressed on the basolateral membranes of B-IC in control kidney CD, but some connecting segment B-IC had more distinct basolateral staining. In response to chronic carbonic anhydrase inhibition by acetazolamide, many A-IC showed a strong apical membrane localization of B2, where it colocalized with E and B1. In rat and mouse epididymis, B2 isoform expression was detected in clear cells, where it was concentrated in subapical vesicles. Unlike B1, B2 did not colocalize with the E subunit in the apical microvilli. These findings indicate that in addition to its role in the acidification of intracellular organelles, the B2 isoform could also contribute to transepithelial proton secretion and the maintenance of acid-base homeostasis.

    Funded by: NICHD NIH HHS: HD-40793; NIDDK NIH HHS: DK-38452, DK-42956, DK-43351, DK-57521, T32 DK-07540

    American journal of physiology. Cell physiology 2004;287;1;C149-62

  • Determination of downstream targets of FGF signalling using gene trap and cDNA subtractive approaches.

    Tateossian H, Powles N, Dickinson R, Ficker M and Maconochie M

    Mammalian Genetics Unit, Medical Research Council, Harwell, Oxon OX11 0RD, UK.

    Signalling through the fibroblast growth factor family (FGF) of ligands is essential for normal mammalian embryonic development. At a cellular level, many details of the molecular basis of the signal transduction process have been uncovered, but our knowledge of the identity of the downstream effectors of the FGF signal in the developing embryo remains limited. We have used two independent approaches to begin to identify downstream targets of FGF signalling in the embryo: (1). a gene trap approach and (2). cDNA subtraction, using mouse embryonic stem (ES) cells as a cellular system representative of an early window on the developing embryo. Both approaches led to the identification of a number of targets of FGF signalling, and we provide data to show that the chaperone Mrj, the tumour antigen Tum, collapsin mediator response protein Crmp, a novel transcriptional repressor Nac1 and ribophorin are all differentially regulated following FGF signalling. Independent gene trapping of Mrj previously indicated a role for the gene in embryogenesis [Development 126 (1999) 1247], and we present transcript data implicating a number of the newly isolated FGF target genes in different embryonic processes.

    Experimental cell research 2004;292;1;101-14

  • 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

  • Osteoclasts express the B2 isoform of vacuolar H(+)-ATPase intracellularly and on their plasma membranes.

    Lee BS, Holliday LS, Ojikutu B, Krits I and Gluck SL

    Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

    Osteoclasts express high levels of vacuolar H(+)-ATPase (V-ATPase) in their ruffled membranes, driving the secretion of H+ required for normal bone resorption. Previous reports have suggested that the B subunit of the osteoclast V-ATPase differs from those expressed in kidney and other tissues. In this study, B subunit isoform-specific antibodies and cDNA probes were used to examine which B subunit isoform is expressed in osteoclasts and osteoclast-like cells. Immunoblotting and RNA hybridization analysis were used to demonstrate that cells from an osteoclast-rich mouse bone marrow culture model express the B2 but not the B1 subunit isoform. Immunocytochemical staining of murine osteoclasts generated in vitro and of native rat osteoclasts in bone sections showed that the B2 but not the B1 isoform was expressed at high levels and was polarized to the ruffled membrane. Human marrow cultures and monocyte-derived macrophages, used as models for osteoclasts, also expressed the B2 but not the B1 subunit isoform. These results indicate that V-ATPases containing the B2 subunit isoform mediate osteoclast bone resorption.

    Funded by: NIAMS NIH HHS: AR-32087; NIDDK NIH HHS: DK-09976, DK-38848; ...

    The American journal of physiology 1996;270;1 Pt 1;C382-8

  • Resorption-cycle-dependent polarization of mRNAs for different subunits of V-ATPase in bone-resorbing osteoclasts.

    Laitala-Leinonen T, Howell ML, Dean GE and Väänänen HK

    Department of Anatomy, University of Oulu, Finland.

    Protein sorting in eukaryotic cells is mainly done by specific targeting of polypeptides. The present evidence from oocytes, neurons, and some other polarized cells suggests that protein sorting can be further facilitated by concentrating mRNAs to their corresponding subcellular areas. However, very little is known about the mechanism(s) involved in mRNA targeting, or how widespread and dynamic such mRNA sorting might be. In this study, we have used an in vitro cell culture system, where large multinucleated osteoclasts undergo continuous structural and functional changes from polarized (resorbing) to a nonpolarized (resting) stage. We demonstrate here, using a nonradioactive in situ hybridization technique and confocal microscopy, that mRNAs for several vacuolar H(+)-ATPase subunits change their localization and polarity in osteoclasts according to the resorption cycle, whereas mRNA for cytoplasmic carbonic anhydrase II is found diffusely located throughout the osteoclast during the whole resorption cycle. Antisense RNA against the 16-kDa or 60-kDa V-ATPase subunit inhibits polarization of the osteoclasts, as determined by cytoskeleton staining. Antisense RNA against carbonic anhydrase II, however, has no such effect.

    Funded by: NIGMS NIH HHS: GM-39555

    Molecular biology of the cell 1996;7;1;129-42

  • Heterogeneity of vacuolar H(+)-ATPase: differential expression of two human subunit B isoforms.

    van Hille B, Richener H, Schmid P, Puettner I, Green JR and Bilbe G

    CIBA-GEIGY A.G., Basel, Switzerland.

    The catalytic domain of the vacuolar proton ATPase is composed of a hexamer of three A subunits and three B subunits. Here we describe the cloning and characterization of a cDNA isoform of subunit B, HO57, from an osteoclastoma cDNA library. HO57 is represented by three species of mRNA of 1.6, 2.6 and 2.8 kb and is expressed at low levels in a range of human tissues, but at significantly higher levels in brain, kidney and osteoclastoma, and is probably an ubiquitously expressed isoform. In contrast, the kidney-specific isoform has an mRNA of 2 kb and is specifically expressed at high levels only in kidney and, at a lower level, in placenta. Thus the HO57 isoform is integral to the vacuolar ATPase found in the general secretory system of all cells as well as in vacuolar-ATPase-rich sources such as neurones and osteoclasts, whereas both the kidney-specific isoform and HO57 are highly expressed in the kidney. Furthermore, we show by in situ hybridization that HO57 is the only isoform that is exclusively and highly expressed by osteoclasts.

    The Biochemical journal 1994;303 ( Pt 1);191-8

Gene lists (6)

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
L00000060 G2C Mus musculus BAYES-COLLINS-HUMAN-PSD-CONSENSUS Human cortex PSD consensus (ortho) 748
L00000062 G2C Mus musculus BAYES-COLLINS-MOUSE-PSD-CONSENSUS Mouse cortex PSD consensus 984
L00000070 G2C Mus musculus BAYES-COLLINS-HUMAN-PSD-FULL Human cortex biopsy PSD full list (ortho) 1461
L00000072 G2C Mus musculus BAYES-COLLINS-MOUSE-PSD-FULL Mouse cortex PSD full list 1556
© 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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