Abstract
We observed severe ataxia in mice homozygous for modification of the Pparg locus. Genetic analysis and nucleotide sequencing revealed that ataxia is caused by a T692K substitution in plasma membrane calcium ATPase 2 (Pmca2), which is tightly linked to Pparg, but not by modified PPARγ itself. We traced this mutation and found that it arose spontaneously during clonal expansion of the targeted embryonic stem (ES) cells. Consistent with the deafwaddler phenotype in other Pmca2 mutants, homozygous T692K Pmca2 mutants exhibit severe balance disorder, impaired neurologic reflexes, and motor coordination, and have profound hearing loss. Heterozygous mutants have normal movement and motor function but are severely deficient in hearing. Our findings represent a cautionary example since, although rare, spontaneous mutations do arise in ES cells during culture and hitchhike onto the targeted gene mutation.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Apicella S, Chen S, Bing R, Penniston JT, Llinas R, et al. (1997) Plasmalemmal ATPase calcium pump localizes to inner and outer hair bundles. Neuroscience 79:145–1151
Bronson SK, Smithies O (1994) Altering mice by homologous recombination using embryonic stem cells. J Biol Chem 269:7155–27158
Bunting M, Bernstein KE, Greer JM, Capecchi MR, Thomas KR (1999) Targeting genes for self-excision in the germ line. Genes Dev 13:524–1528
Chen CY, Chen TM, Shyu AB (1994) Interplay of two functionally and structurally distinct domains of the c-fos AU-rich element specifies its mRNA-destabilizing function. Mol Cell Biol 14:16–426
Crawley JN (2000) What’s wrong with my mouse? Behavioral phenotyping of transgenic and knockout mice. (New York: Wiley-Liss)
Doetschman T, Maeda N, Smithies O (1988) Targeted mutation of the Hprt gene in mouse embryonic stem cells. Proc Natl Acad Sci U S A 85:583–8587
Duncan GE, Moy SS, Perez A, Eddy DM, Zinzow WM, et al. (2004) Deficits in sensorimotor gating and tests of social behavior in a genetic model of reduced NMDA receptor function. Behav Brain Res 153:07–519
Kakoki M, Tsai YS, Kim HS, Hatada S, Ciavatta DJ, et al. (2004) Altering the expression in mice of genes by modifying their 3′ regions. Dev Cell 6:97–606
Kozel PJ, Friedman RA, Erway LC, Yamoah EN, Liu LH, et al. (1998) Balance and hearing deficits in mice with a null mutation in the gene encoding plasma membrane Ca2+-ATPase isoform 2. J Biol Chem 273:18693–18696
Maitra A, Arking DE, Shivapurkar N, Ikeda M, Stastny V, et al. (2005) Genomic alterations in cultured human embryonic stem cells. Nat Genet 37:1099–1103
McCullough BJ, Tempel BL (2004) Haplo-insufficiency revealed in deafwaddler mice when tested for hearing loss and ataxia. Hear Res 195:90–102
Moy SS, Nadler JJ, Perez A, Barbaro RP, Johns JM, et al. (2004) Sociability and preference for social novelty in five inbred strains: an approach to assess autistic-like behavior in mice. Genes Brain Behav 3:287–302
Muller U (1999) Ten years of gene targeting: targeted mouse mutants, from vector design to phenotype analysis. Mech Dev 82:3–21
Paylor R, Crawley JN (1997) Inbred strain differences in prepulse inhibition of the mouse startle response. Psychopharmacology (Berl) 132:169–180
Piedrahita JA, Zhang SH, Hagaman JR, Oliver PM, Maeda N (1992) Generation of mice carrying a mutant apolipoprotein E gene inactivated by gene targeting in embryonic stem cells. Proc Natl Acad Sci U S A 89:4471–4475
Reid LH, Shesely EG, Kim HS, Smithies O (1991) Cotransformation and gene targeting in mouse embryonic stem cells. Mol Cell Biol 11:2769–2777
Schmidt EE, Taylor DS, Prigge JR, Barnett S, Capecchi MR (2000) Illegitimate Cre-dependent chromosome rearrangements in transgenic mouse spermatids. Proc Natl Acad Sci U S A 97:13702–13707
Smithies O (2005) Many little things: one geneticist’s view of complex diseases. Nat Rev Genet 6:419–425
Smithies O, Maeda N (1995) Gene targeting approaches to complex genetic diseases: atherosclerosis and essential hypertension. Proc Natl Acad Sci U S A 92:5266–5272
Stahl WL, Eakin TJ, Owens JW Jr, Breininger JF, Filuk PE, et al. (1992) Plasma membrane Ca(2+)-ATPase isoforms: distribution of mRNAs in rat brain by in situ hybridization. Brain Res Mol Brain Res 16:223–231
Street VA, McKee-Johnson JW, Fonseca RC, Tempel BL, Noben-Trauth K (1998) Mutations in a plasma membrane Ca2+-ATPase gene cause deafness in deafwaddler mice. Nat Genet 19:390–394
Sweadner KJ, Donnet C (2001) Structural similarities of Na,K-ATPase and SERCA, the Ca(2+)-ATPase of the sarcoplasmic reticulum. Biochem J 356:685–704
Takahashi K, Kitamura K (1999) A point mutation in a plasma membrane Ca(2+)-ATPase gene causes deafness in Wriggle Mouse Sagami. Biochem Biophys Res Commun 261:773–778
Thomas KR, Deng C, Capecchi MR (1992) High-fidelity gene targeting in embryonic stem cells by using sequence replacement vectors. Mol Cell Biol 12:2919–2923
Toyoshima C, Mizutani T (2004) Crystal structure of the calcium pump with a bound ATP analogue. Nature 430:529–535
Zheng H, Hasty P, Brenneman MA, Grompe M, Gibbs RA, et al. (1991) Fidelity of targeted recombination in human fibroblasts and murine embryonic stem cells. Proc Natl Acad Sci U S A 88:8067–8071
Acknowledgments
The authors thank Dr. Kumar Pandya for discussion and critical reading of the manuscript. This work was supported by grants from the National Institutes of Health to N. Maeda (HL42630 and HL70523) and the American Heart Association to Y.S. Tsai (0215270U).
Author information
Authors and Affiliations
Corresponding author
Electronic Supplementary Material
Video supplement
Video contribution
Rights and permissions
About this article
Cite this article
Tsai, YS., Pendse, A., Moy, S.S. et al. A de novo deafwaddler mutation of Pmca2 arising in ES cells and hitchhiking with a targeted modification of the Pparg gene. Mamm Genome 17, 716–722 (2006). https://doi.org/10.1007/s00335-005-0191-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00335-005-0191-z