Abstract
ATP13A4 is a member of the subfamily of P5-type ATPases. P5-type ATPases are the least studied of the P-type ATPase subfamilies with no ion specificities assigned to them. In order to elucidate ATP13A4 function, we studied the protein’s subcellular localization and tested whether it is involved in calcium regulation. The intracellular calcium concentration was measured in COS-7 cells over-expressing mouse ATP13A4 using ratiometric calcium imaging with fura-2 AM as a calcium indicator. The results of this study show that ATP13A4 is localized to the endoplasmic reticulum (ER). Furthermore, we demonstrate that over-expression of ATP13A4 in COS-7 cells caused a significant increase in the intracellular calcium level. Interestingly, over-expression of the sequence variant containing a substitution of aspartic acid for a glutamic acid (E646D), previously found in patients with autism spectrum disorder (ASD), did not increase the free cellular calcium likely due to the mutation. In this study, we also describe the expression of ATP13A4 during mouse embryonic development. Quantitative real-time PCR revealed that ATP13A4 was highly expressed at embryonic days 15–17, when neurogenesis takes place. The present study is the first to provide further insights into the biological role of a P5-type ATPase. Our results demonstrate that ATP13A4 may be involved in calcium regulation and that its expression is developmentally regulated. Overall, this study provides support for the hypothesis that ATP13A4 may play a vital role in the developing nervous system and its impairment can contribute to the symptoms seen in ASD.
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References
Augustine GJ (2001) How does calcium trigger neurotransmitter release? Curr Opin Neurobiol 11:320–326
Auranen M, Vanhala R, Varilo T, Ayers K, Kempas E, Ylisaukko-Oja T, Sinsheimer JS, Peltonen L, Jarvela I (2002) A genomewide screen for autism-spectrum disorders: evidence for a major susceptibility locus on chromosome 3q25–27. Am J Hum Genet 71:777–790
Auranen M, Varilo T, Alen R, Vanhala R, Ayers K, Kempas E, Ylisaukko-Oja T, Peltonen L, Jarvela I (2003) Evidence for allelic association on chromosome 3q25–27 in families with autism spectrum disorders originating from a subisolate of Finland. Mol Psychiatry 8:879–884
Axelsen KB, Palmgren MG (1998) Evolution of substrate specificities in the P-type ATPase superfamily. J Mol Evol 46:84–101
Bardo S, Cavazzini MG, Emptage N (2006) The role of the endoplasmic reticulum Ca2+ store in the plasticity of central neurons. Trends Pharmacol Sci 27:78–84
Barnby G, Abbott A, Sykes N, Morris A, Weeks DE, Mott R, Lamb J, Bailey AJ, Monaco AP (2005) Candidate-gene screening and association analysis at the autism-susceptibility locus on chromosome 16p: evidence of association at GRIN2A and ABAT. Am J Hum Genet 76:950–966
Bezprozvanny I, Hayden MR (2004) Deranged neuronal calcium signaling and Huntington disease. Biochem Biophys Res Commun 322:1310–1317
Blaustein MP, Golovina VA (2001) Structural complexity and functional diversity of endoplasmic reticulum Ca(2+) stores. Trends Neurosci 24:602–608
Burette A, Rockwood JM, Strehler EE, Weinberg RJ (2003) Isoform-specific distribution of the plasma membrane Ca2+ ATPase in the rat brain. J Comp Neurol 467:464–476
Cox B, Emili A (2006) Tissue subcellular fractionation and protein extraction for use in mass-spectrometry-based proteomics. Nat Protoc 1:1872–1878
Cronin SR, Khoury A, Ferry DK, Hampton RY (2000) Regulation of HMG-CoA reductase degradation requires the P-type ATPase Cod1p/Spf1p. J Cell Biol 148:915–924
Cronin SR, Rao R, Hampton RY (2002) Cod1p/Spf1p is a P-type ATPase involved in ER function and Ca2+ homeostasis. J Cell Biol 157:1017–1028
Ding J, Wu Z, Crider BP, Ma Y, Li X, Slaughter C, Gong L, Xie XS (2000) Identification and functional expression of four isoforms of ATPase II, the putative aminophospholipid translocase. Effect of isoform variation on the ATPase activity and phospholipid specificity. J Biol Chem 275:23378–23386
El Meskini R, Cline LB, Eipper BA, Ronnett GV (2005) The developmentally regulated expression of Menkes protein ATP7A suggests a role in axon extension and synaptogenesis. Dev Neurosci 27:333–348
Empson RM, Garside ML, Knopfel T (2007) Plasma membrane Ca2+ ATPase 2 contributes to short-term synapse plasticity at the parallel fiber to Purkinje neuron synapse. J Neurosci 27:3753–3758
Fitzjohn SM, Collingridge GL (2002) Calcium stores and synaptic plasticity. Cell Calcium 32:405–411
Gillies K, Price DJ (1993a) Cell migration and subplate loss in explant cultures of murine cerebral cortex. NeuroReport 4:911–914
Gillies K, Price DJ (1993b) The fates of cells in the developing cerebral cortex of normal and methylazoxymethanol acetate-lesioned mice. Eur J Neurosci 5:73–84
Gomes E, Jakobsen MK, Axelsen KB, Geisler M, Palmgren MG (2000) Chilling tolerance in Arabidopsis involves ALA1, a member of a new family of putative aminophospholipid translocases. Plant Cell 12:2441–2454
Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260:3440–3450
Hardingham GE, Bading H (1999) Calcium as a versatile second messenger in the control of gene expression. Microsc Res Tech 46:348–355
Hensley K, Carney JM, Mattson MP, Aksenova M, Harris M, Wu JF, Floyd RA, Butterfield DA (1994) A model for beta-amyloid aggregation and neurotoxicity based on free radical generation by the peptide: relevance to Alzheimer disease. Proc Natl Acad Sci USA 91:3270–3274
Jamain S, Betancur C, Quach H, Philippe A, Fellous M, Giros B, Gillberg C, Leboyer M, Bourgeron T (2002) Linkage and association of the glutamate receptor 6 gene with autism. Mol Psychiatry 7:302–310
Jensen TP, Filoteo AG, Knopfel T, Empson RM (2007) Presynaptic plasma membrane Ca2+ ATPase isoform 2a regulates excitatory synaptic transmission in rat hippocampal CA3. J Physiol 579:85–99
Jorgensen PL, Hakansson KO, Karlish SJ (2003) Structure and mechanism of Na, K-ATPase: functional sites and their interactions. Annu Rev Physiol 65:817–849
Kann O, Kovacs R (2007) Mitochondria and neuronal activity. Am J Physiol Cell Physiol 292:C641–C657
Konur S, Ghosh A (2005) Calcium signaling and the control of dendritic development. Neuron 46:401–405
Kwasnicka-Crawford DA, Carson AR, Roberts W, Summers AM, Rehnstrom K, Jarvela I, Scherer SW (2005) Characterization of a novel cation transporter ATPase gene (ATP13A4) interrupted by 3q25–q29 inversion in an individual with language delay. Genomics 86:182–194
Levers TE, Edgar JM, Price DJ (2001) The fates of cells generated at the end of neurogenesis in developing mouse cortex. J Neurobiol 48:265–277
Martonosi AN, Pikula S (2003) The network of calcium regulation in muscle. Acta Biochim Pol 50:1–30
Mata AM, Sepulveda MR (2005) Calcium pumps in the central nervous system. Brain Res Brain Res Rev 49:398–405
Moller JV, Juul B, le Maire M (1996) Structural organization, ion transport, and energy transduction of P-type ATPases. Biochim Biophys Acta 1286:1–51
Moller AB, Asp T, Holm PB, Palmgren MG (2008) Phylogenetic analysis of P5 P-type ATPases, a eukaryotic lineage of secretory pathway pumps. Mol Phylogenet Evol 46:619–634
Monteith GR, Roufogalis BD (1995) The plasma membrane calcium pump—a physiological perspective on its regulation. Cell Calcium 18:459–470
Noji T, Yamamoto T, Saito K, Fujimura-Kamada K, Kondo S, Tanaka K (2006) Mutational analysis of the Lem3p-Dnf1p putative phospholipid-translocating P-type ATPase reveals novel regulatory roles for Lem3p and a carboxyl-terminal region of Dnf1p independent of the phospholipid-translocating activity of Dnf1p in yeast. Biochem Biophys Res Commun 344:323–331
Paulusma CC, Folmer DE, Ho-Mok KS, de Waart DR, Hilarius PM, Verhoeven AJ, Oude Elferink RP (2008) ATP8B1 requires an accessory protein for endoplasmic reticulum exit and plasma membrane lipid flippase activity. Hepatology 47:268–278
Periasamy M, Huke S (2001) SERCA pump level is a critical determinant of Ca(2+)homeostasis and cardiac contractility. J Mol Cell Cardiol 33:1053–1063
Pomorski T, Lombardi R, Riezman H, Devaux PF, van Meer G, Holthuis JC (2003) Drs2p-related P-type ATPases Dnf1p and Dnf2p are required for phospholipid translocation across the yeast plasma membrane and serve a role in endocytosis. Mol Biol Cell 14:1240–1254
Ramirez A, Heimbach A, Grundemann J, Stiller B, Hampshire D, Cid LP, Goebel I, Mubaidin AF, Wriekat AL, Roeper J, Al-Din A, Hillmer AM, Karsak M, Liss B, Woods CG, Behrens MI, Kubisch C (2006) Hereditary parkinsonism with dementia is caused by mutations in ATP13A2, encoding a lysosomal type 5 P-type ATPase. Nat Genet 38:1184–1191
Roy J, Minotti S, Dong L, Figlewicz DA, Durham HD (1998) Glutamate potentiates the toxicity of mutant Cu/Zn-superoxide dismutase in motor neurons by postsynaptic calcium-dependent mechanisms. J Neurosci 18:9673–9684
Saito K, Fujimura-Kamada K, Furuta N, Kato U, Umeda M, Tanaka K (2004) Cdc50p, a protein required for polarized growth, associates with the Drs2p P-type ATPase implicated in phospholipid translocation in Saccharomyces cerevisiae. Mol Biol Cell 15:3418–3432
Santos AR, Duarte CB (2008) Validation of internal control genes for expression studies: effects of the neurotrophin BDNF on hippocampal neurons. J Neurosci Res 86:3684–3692
Schultheis PJ, Hagen TT, O’Toole KK, Tachibana A, Burke CR, McGill DL, Okunade GW, Shull GE (2004) Characterization of the P5 subfamily of P-type transport ATPases in mice. Biochem Biophys Res Commun 323:731–738
Sepulveda MR, Marcos D, Berrocal M, Raeymaekers L, Mata AM, Wuytack F (2008) Activity and localization of the Secretory Pathway Ca2+ -ATPase isoform 1 (SPCA1) in different areas of the mouse brain during postnatal development. Mol Cell Neurosci 38:461–473
Serajee FJ, Zhong H, Nabi R, Huq AH (2003) The metabotropic glutamate receptor 8 gene at 7q31: partial duplication and possible association with autism. J Med Genet 40:e42
Sheng ZH, Westenbroek RE, Catterall WA (1998) Physical link and functional coupling of presynaptic calcium channels and the synaptic vesicle docking/fusion machinery. J Bioenerg Biomembr 30:335–345
Splawski I, Timothy KW, Sharpe LM, Decher N, Kumar P, Bloise R, Napolitano C, Schwartz PJ, Joseph RM, Condouris K, Tager-Flusberg H, Priori SG, Sanguinetti MC, Keating MT (2004) Ca(V)1.2 calcium channel dysfunction causes a multisystem disorder including arrhythmia and autism. Cell 119:19–31
Splawski I, Timothy KW, Decher N, Kumar P, Sachse FB, Beggs AH, Sanguinetti MC, Keating MT (2005) Severe arrhythmia disorder caused by cardiac L-type calcium channel mutations. Proc Natl Acad Sci USA 102:8089–8096 discussion 8086–8
Splawski I, Yoo DS, Stotz SC, Cherry A, Clapham DE, Keating MT (2006) CACNA1H mutations in autism spectrum disorders. J Biol Chem 281:22085–22091
Stanley EF (1997) The calcium channel and the organization of the presynaptic transmitter release face. Trends Neurosci 20:404–409
Surmeier DJ (2007) Calcium, ageing, and neuronal vulnerability in Parkinson’s disease. Lancet Neurol 6:933–938
Suzuki C (2001) Immunochemical and mutational analyses of P-type ATPase Spf1p involved in the yeast secretory pathway. Biosci Biotechnol Biochem 65:2405–2411
Suzuki C, Shimma YI (1999) P-type ATPase spf1 mutants show a novel resistance mechanism for the killer toxin SMKT. Mol Microbiol 32:813–823
Takuma H, Kwak S, Yoshizawa T, Kanazawa I (1999) Reduction of GluR2 RNA editing, a molecular change that increases calcium influx through AMPA receptors, selective in the spinal ventral gray of patients with amyotrophic lateral sclerosis. Ann Neurol 46:806–815
Tipper DJ, Harley CA (2002) Yeast genes controlling responses to topogenic signals in a model transmembrane protein. Mol Biol Cell 13:1158–1174
Vashist S, Frank CG, Jakob CA, Ng DT (2002) Two distinctly localized p-type ATPases collaborate to maintain organelle homeostasis required for glycoprotein processing and quality control. Mol Biol Cell 13:3955–3966
Wuytack F, Raeymaekers L, Missiaen L (2003) PMR1/SPCA Ca2+ pumps and the role of the Golgi apparatus as a Ca2+ store. Pflugers Arch 446:148–153
Xiang M, Mohamalawari D, Rao R (2005) A novel isoform of the secretory pathway Ca2+, Mn(2+)-ATPase, hSPCA2, has unusual properties and is expressed in the brain. J Biol Chem 280:11608–11614
Acknowledgments
This study was supported by the Natural Sciences and Engineering Research Council of Canada (NSERC) and Canada Foundation for Innovation (CFI). We thank Dr. Thomas Hawke for providing mouse brain samples.
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Vallipuram, J., Grenville, J. & Crawford, D.A. The E646D-ATP13A4 Mutation Associated with Autism Reveals a Defect in Calcium Regulation. Cell Mol Neurobiol 30, 233–246 (2010). https://doi.org/10.1007/s10571-009-9445-8
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DOI: https://doi.org/10.1007/s10571-009-9445-8