title: Dystonia type 6 gene product Thap1: identification of a 50 kDa DNA-binding species in neuronal nuclear fractions.
abstract: Mutations in THAP1 result in dystonia type 6, with partial penetrance and variable phenotype. The goal of this study was to examine the nature and expression pattern of the protein product(s) of the Thap1 transcription factor (DYT6 gene) in mouse neurons, and to study the regional and developmental distribution, and subcellular localization of Thap1 protein. The goal was accomplished via overexpression and knock-down of Thap1 in the HEK293T cell line and in mouse striatal primary cultures and western blotting of embryonic Thap1-null tissue. The endogenous and transduced Thap1 isoforms were characterized using three different commercially available anti-Thap1 antibodies and validated by immunoprecipitation and DNA oligonucleotide affinity chromatography. We identified multiple, novel Thap1 species of apparent Mr 32 kDa, 47 kDa, and 50-52 kDa in vitro and in vivo, and verified the previously identified species at 29-30 kDa in neurons. The Thap1 species at the 50 kDa size range was exclusively detected in murine brain and testes and were located in the nuclear compartment. Thus, in addition to the predicted 25 kDa apparent Mr, we identified Thap1 species with greater apparent Mr that we speculate may be a result of posttranslational modifications. The neural localization of the 50 kDa species and its nuclear compartmentalization suggests that these may be key Thap1 species controlling neuronal gene transcription. Dysfunction of the neuronal 50 kDa species may therefore be implicated in the pathogenesis of DYT6.
authors.forename: Maitane
authors.initials: M
authors.lastname: Ortiz-Virumbrales
authors.forename: Marta
authors.initials: M
authors.lastname: Ruiz
authors.forename: Eugene
authors.initials: E
authors.lastname: Hone
authors.forename: Georgia
authors.initials: G
authors.lastname: Dolios
authors.forename: Rong
authors.initials: R
authors.lastname: Wang
authors.forename: Andrika
authors.initials: A
authors.lastname: Morant
authors.forename: Jessica
authors.initials: J
authors.lastname: Kottwitz
authors.forename: Laurie J
authors.initials: LJ
authors.lastname: Ozelius
authors.forename: Sam
authors.initials: S
authors.lastname: Gandy
authors.forename: Michelle E
authors.initials: ME
authors.lastname: Ehrlich
mesh_terms: D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000235:genetics* / Q000378:metabolism*; D001921:Brain / Q000166:cytology / Q000378:metabolism; D002466:Cell Nucleolus / Q000378:metabolism*; D002478:Cells, Cultured; D003342:Corpus Striatum / Q000166:cytology; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D004622:Embryo, Mammalian; D005786:Gene Expression Regulation / Q000235:genetics; D006801:Humans; D047468:Immunoprecipitation; D051379:Mice; D018345:Mice, Knockout; D018832:Molecular Chaperones / Q000378:metabolism; D008970:Molecular Weight; D009417:Nerve Tissue / Q000378:metabolism / Q000473:pathology; D009474:Neurons / Q000378:metabolism* / Q000648:ultrastructure*; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D012262:Ribonucleoside Diphosphate Reductase; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; D004268:DNA-Binding Proteins; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; C474080:THAP1 protein, human; C108175:TOR1A protein, human; D025521:Tumor Suppressor Proteins; C475286:RRM1 protein, human; D012262:Ribonucleoside Diphosphate Reductase