title: Interactome of the inhibitory isoform of the nuclear transporter Importin 13.
abstract: Importin 13 (Imp13) is a bidirectional nuclear transporter of proteins involved in a range of important cellular processes, with an N-terminally truncated inhibitory isoform (tImp13) specifically expressed in testis. To gain insight into tImp13 function, we performed a yeast-2-hybrid screen from a human testis cDNA library, identifying for the first time a suite of interactors with roles in diverse cellular process. We validated the interaction of tImp13 with Eukaryotic translation initiation factor 4γ2 (EIF4G2) and High mobility group containing protein 20A (HMG20A), benchmarking that with glucocorticoid receptor (GR), a known Imp13 interactor expressed in testis. Coimmunoprecipitation assays indicated association of both tImp13 and Imp13 with EIF4G2, HMG20A and GR. Quantitative confocal microscopic analysis revealed the ability of tImp13 to inhibit the nuclear localisation of EIF4G2, HMG20A and GR, as well as that of Imp13 to act as a nuclear exporter for both EIF4G2 and HMG20A, and as a nuclear importer for GR. The physiological relevance of these results was highlighted by the cytoplasmic localisation of EIF4G2, HMG20A and GR in pachytene spermatocytes/round spermatids in the murine testis where tImp13 is present at high levels, in contrast to the nuclear localisation of HMG20A and GR in spermatogonia, where tImp13 is largely absent. Interestingly, Imp13, EIF4G2, HMG20A and GR were found together in the acrosome vesicle of murine epididymal spermatozoa. Collectively, our findings show, for the first time, that tImp13 may have a functional role in the mature spermatozoa, in addition to that in the meiotic germ cells of the testis.
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Shadma
authors.initials: S
authors.lastname: Fatima
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Kylie M
authors.initials: KM
authors.lastname: Wagstaff
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Kim G
authors.initials: KG
authors.lastname: Lieu
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Rebecca G
authors.initials: RG
authors.lastname: Davies
authors.affiliation: Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan; Embryology Unit, Children's Medical Research Institute, Westmead, Australia.
authors.forename: Satomi S
authors.initials: SS
authors.lastname: Tanaka
authors.affiliation: Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan; Embryology Unit, Children's Medical Research Institute, Westmead, Australia.
authors.forename: Yasuka L
authors.initials: YL
authors.lastname: Yamaguchi
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia; Department of Anatomy and Developmental Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Kate L
authors.initials: KL
authors.lastname: Loveland
authors.affiliation: Embryology Unit, Children's Medical Research Institute, Westmead, Australia; Discipline of Medicine, Sydney Medical School, University of Sydney, Sydney, Australia.
authors.forename: Patrick P L
authors.initials: PP
authors.lastname: Tam
authors.affiliation: Department of Biochemistry & Molecular Biology, Monash University, Clayton, Victoria, Australia. Electronic address: david.jans@monash.edu.
authors.forename: David A
authors.initials: DA
authors.lastname: Jans
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D039603:Eukaryotic Initiation Factor-4G / Q000235:genetics / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D015723:Gene Library; D006609:High Mobility Group Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D028884:Karyopherins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D051379:Mice; D011485:Protein Binding; D025941:Protein Interaction Mapping; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013087:Spermatids / Q000254:growth & development / Q000378:metabolism* / Q000648:ultrastructure; D013090:Spermatocytes / Q000254:growth & development / Q000378:metabolism* / Q000648:ultrastructure; D013091:Spermatogenesis / Q000235:genetics*; D013737:Testis / Q000166:cytology / Q000254:growth & development / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: C104828:Eif4g2 protein, mouse; D039603:Eukaryotic Initiation Factor-4G; D006609:High Mobility Group Proteins; C507322:Hmg20a protein, mouse; D028884:Karyopherins; D020033:Protein Isoforms; D011965:Receptors, Glucocorticoid; C514744:importin 13 protein, mouse
keywords: Imp13; NRNI; Nuclear import; Spermatogenesis; Testis; tImp13