title: Manipulation of P-TEFb control machinery by HIV: recruitment of P-TEFb from the large form by Tat and binding of HEXIM1 to TAR.
abstract: Basal transcription of the HIV LTR is highly repressed and requires Tat to recruit the positive transcription elongation factor, P-TEFb, which functions to promote the transition of RNA polymerase II from abortive to productive elongation. P-TEFb is found in two forms in cells, a free, active form and a large, inactive complex that also contains 7SK RNA and HEXIM1 or HEXIM2. Here we show that HIV infection of cells led to the release of P-TEFb from the large form. Consistent with Tat being the cause of this effect, transfection of a FLAG-tagged Tat in 293T cells caused a dramatic shift of P-TEFb out of the large form to a smaller form containing Tat. In vitro, Tat competed with HEXIM1 for binding to 7SK, blocked the formation of the P-TEFb-HEXIM1-7SK complex, and caused the release P-TEFb from a pre-formed P-TEFb-HEXIM1-7SK complex. These findings indicate that Tat can acquire P-TEFb from the large form. In addition, we found that HEXIM1 binds tightly to the HIV 5' UTR containing TAR and recruits and inhibits P-TEFb activity. This suggests that in the absence of Tat, HEXIM1 may bind to TAR and repress transcription elongation of the HIV LTR.
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, IA, USA.
authors.forename: Stanley C
authors.initials: SC
authors.lastname: Sedore
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Byers
authors.forename: Sebastian
authors.initials: S
authors.lastname: Biglione
authors.forename: Jason P
authors.initials: JP
authors.lastname: Price
authors.forename: Wendy J
authors.initials: WJ
authors.lastname: Maury
authors.forename: David H
authors.initials: DH
authors.lastname: Price
mesh_terms: D001665:Binding Sites; D001667:Binding, Competitive; D002460:Cell Line; D015696:Gene Products, tat / Q000378:metabolism*; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D042925:Positive Transcriptional Elongation Factor B / Q000737:chemistry / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000378:metabolism; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus
chemical_list: D015696:Gene Products, tat; C476123:HEXIM1 protein, human; D012342:RNA, Small Nuclear; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus; D042925:Positive Transcriptional Elongation Factor B