title: The Purkinje cell degeneration 5J mutation is a single amino acid insertion that destabilizes Nna1 protein.
abstract: In the mouse, Purkinje cell degeneration (pcd) is a recessive mutation characterized by degeneration of cerebellar Purkinje cells, retinal photoreceptors, olfactory bulb mitral neurons, and certain thalamic neurons, and is accompanied by defective spermatogenesis. Previous studies of pcd have led to the identification of Nna1 as the causal gene; however, how loss of Nna1 function results in neurodegeneration remains unresolved. One useful approach for establishing which functional domains of a protein underlie a recessive phenotype has been to determine the genetic basis of the various alleles at the locus of interest. Because none of the pcd alleles analyzed at the time of the identification of Nna1 provided insight into the molecular basis of Nna1 loss-of-function, we obtained a recent pcd remutation--pcd5J, and after determining that its phenotype is comparable to existing pcd severe alleles, we sought its genetic basis by sequencing Nna1. In this article we report that pcd5J results from the insertion of a single GAC triplet encoding an aspartic acid residue at position 775 of Nna1. Although this insertion does not affect Nna1 expression at the RNA level, Nna1pcd-5J protein expression is markedly decreased. Pulse-chase experiments reveal that the aspartic acid insertion dramatically destabilizes Nna1pcd-5J protein, accounting for the observation that pcd5J is a severe allele. The presence of a readily detectable genetic mutation in pcd5J confirms that Nna1 loss-of-function alone underlies the broad pcd phenotype and will facilitate further studies of how Nna1 loss-of-function produces neurodegeneration and defective spermatogenesis in pcd mice.
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, Washington 98195-7110, USA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Chakrabarti
authors.forename: James T
authors.initials: JT
authors.lastname: Neal
authors.forename: Michael
authors.initials: M
authors.lastname: Miles
authors.forename: Refugio A
authors.initials: RA
authors.lastname: Martinez
authors.forename: Annette C
authors.initials: AC
authors.lastname: Smith
authors.forename: Bryce L
authors.initials: BL
authors.lastname: Sopher
authors.forename: Albert R
authors.initials: AR
authors.lastname: La Spada
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002531:Cerebellum / Q000378:metabolism / Q000473:pathology; D019204:GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009410:Nerve Degeneration / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D011689:Purkinje Cells / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D012160:Retina / Q000378:metabolism / Q000473:pathology; D046929:Serine-Type D-Ala-D-Ala Carboxypeptidase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D001224:Aspartic Acid; D046929:Serine-Type D-Ala-D-Ala Carboxypeptidase; C420042:Agtpbp1 protein, mouse; D019204:GTP-Binding Proteins