Human immunodeficiency virus type 1 Vpr interacts with spliceosomal protein SAP145 to mediate cellular pre-mRNA splicing inhibition.
Vpr, an accessory gene product of human immunodeficiency virus type 1 (HIV-1), affects both viral and cellular proliferation by mediating long terminal repeat activation, cell cycle arrest at the G2 phase, and apoptosis. We previously found that Vpr plays a novel role as a regulator of pre-mRNA splicing both in vivo and in vitro. However, the cellular target of Vpr, as well as the mechanism of cellular pre-mRNA splicing inhibition by Vpr, is unknown. Here, we show clearly that Vpr inhibits the splicing of cellular pre-mRNA, such as beta-globin pre-mRNA and immunoglobulin (Ig) M pre-mRNA and that the third alpha-helical domain and arginine-rich region are important its ability to inhibit splicing. Additionally, using mutants with specific substitutions in two domains of Vpr, we demonstrated that the interaction between Vpr and SAP145, an essential splicing factor, was indispensable for splicing inhibition. Finally, co-immunoprecipitation and in vitro competitive binding assays indicated that Vpr associates with SAP145 and interferes with SAP145-SAP49 complex formation. Thus, these results suggest that cellular expression of Vpr may block spliceosome assembly by interfering with the function of the SAP145-SAP49 complex in host cells.
Hashizume
Chieko
C
Retrovirus Research Unit, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Kuramitsu
Madoka
M
Zhang
Xianfeng
X
Kurosawa
Terue
T
Kamata
Masakazu
M
Aida
Yoko
Y
D000595:Amino Acid Sequence; D001609:Beta-Globulins / Q000235:genetics; D001667:Binding, Competitive; D016336:Gene Products, vpr / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007075:Immunoglobulin M / Q000235:genetics; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D011485:Protein Binding; D012322:RNA Precursors / Q000235:genetics*; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes / Q000235:genetics / Q000378:metabolism*
D001609:Beta-Globulins; D016336:Gene Products, vpr; D007075:Immunoglobulin M; D012322:RNA Precursors; D000072260:RNA Splicing Factors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C486348:SF3B2 protein, human; C486347:SF3B4 protein, human