A human splicing factor, SKIP, associates with P-TEFb and enhances transcription elongation by HIV-1 Tat.
HIV-1 Tat binds human CyclinT1 and recruits the CDK9/P-TEFb complex to the viral TAR RNA in a step that links RNA polymerase II (RNAPII) C-terminal domain (CTD) Ser 2 phosphorylation with transcription elongation. Previous studies have suggested a connection between Tat and pre-mRNA splicing factors. Here we show that the splicing-associated c-Ski-interacting protein, SKIP, is required for Tat transactivation in vivo and stimulates HIV-1 transcription elongation, but not initiation, in vitro. SKIP associates with CycT1:CDK9/P-TEFb and Tat:P-TEFb complexes in nuclear extracts and interacts with recombinant Tat:P-TEFb:TAR RNA complexes in vitro, indicating that it may act through nascent RNA to overcome pausing by RNAPII. SKIP also associates with U5snRNP proteins and tri-snRNP110K in nuclear extracts, and facilitates recognition of an alternative Tat-specific splice site in vivo. The effects of SKIP on transcription elongation, binding to P-TEFb, and splicing are mediated through the SNW domain. HIV-1 Tat transactivation is accompanied by the recruitment of P-TEFb, SKIP, and tri-snRNP110K to the integrated HIV-1 promoter in vivo, whereas the U5snRNPs associate only with the transcribed coding region. These findings suggest that SKIP plays independent roles in transcription elongation and pre-mRNA splicing.
Brès
Vanessa
V
Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA.
Gomes
Nathan
N
Pickle
Loni
L
Jones
Katherine A
KA
D017398:Alternative Splicing / Q000502:physiology; D015696:Gene Products, tat / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D046912:Multiprotein Complexes / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D056918:Nuclear Receptor Coactivators; D016366:Open Reading Frames / Q000502:physiology; D042925:Positive Transcriptional Elongation Factor B / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D012319:RNA Polymerase II / Q000378:metabolism; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology*; D054322:tat Gene Products, Human Immunodeficiency Virus
D015696:Gene Products, tat; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D056918:Nuclear Receptor Coactivators; D017415:Ribonucleoprotein, U5 Small Nuclear; C111791:SNW1 protein, human; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus; D042925:Positive Transcriptional Elongation Factor B; D012319:RNA Polymerase II