title: A gene-specific role for the Ssu72 RNAPII CTD phosphatase in HIV-1 Tat transactivation.
abstract: HIV-1 Tat stimulates transcription elongation by recruiting the P-TEFb (positive transcription elongation factor-b) (CycT1:CDK9) C-terminal domain (CTD) kinase to the HIV-1 promoter. Here we show that Tat transactivation also requires the Ssu72 CTD Ser5P (S5P)-specific phosphatase, which mediates transcription termination and intragenic looping at eukaryotic genes. Importantly, HIV-1 Tat interacts directly with Ssu72 and strongly stimulates its CTD phosphatase activity. We found that Ssu72 is essential for Tat:P-TEFb-mediated phosphorylation of the S5P-CTD in vitro. Interestingly, Ssu72 also stimulates nascent HIV-1 transcription in a phosphatase-dependent manner in vivo. Chromatin immunoprecipitation (ChIP) experiments reveal that Ssu72, like P-TEFb and AFF4, is recruited by Tat to the integrated HIV-1 proviral promoter in TNF-α signaling 2D10 T cells and leaves the elongation complex prior to the termination site. ChIP-seq (ChIP combined with deep sequencing) and GRO-seq (genome-wide nuclear run-on [GRO] combined with deep sequencing) analysis further reveals that Ssu72 predominantly colocalizes with S5P-RNAPII (RNA polymerase II) at promoters in human embryonic stem cells, with a minor peak in the terminator region. A few genes, like NANOG, also have high Ssu72 at the terminator. Ssu72 is not required for transcription at most cellular genes but has a modest effect on cotranscriptional termination. We conclude that Tat alters the cellular function of Ssu72 to stimulate viral gene expression and facilitate the early S5P-S2P transition at the integrated HIV-1 promoter.
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Yupeng
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Lirong
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Conchi
authors.initials: C
authors.lastname: Estarás
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Seung H
authors.initials: SH
authors.lastname: Choi
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Luis
authors.initials: L
authors.lastname: Moreno
authors.affiliation: Department of Molecular Biology and Microbiology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA;
authors.forename: Jonathan
authors.initials: J
authors.lastname: Karn
authors.affiliation: Department of Chemical Physiology and Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
authors.forename: James J
authors.initials: JJ
authors.lastname: Moresco
authors.affiliation: Department of Chemical Physiology and Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Yates
authors.affiliation: Regulatory Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA; jones@salk.edu.
authors.forename: Katherine A
authors.initials: KA
authors.lastname: Jones
mesh_terms: D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D010749:Phosphoprotein Phosphatases; D011401:Promoter Regions, Genetic; D013601:T-Lymphocytes / Q000378:metabolism; D015533:Transcriptional Activation; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000235:genetics* / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D010749:Phosphoprotein Phosphatases; C496300:SSU72 protein, human
keywords: CTD phosphatase; HIV-1 Tat; Ssu72; elongation