Phosphorylation of HEXIM1 at Tyr271 and Tyr274 Promotes Release of P‐TEFb from the 7SK snRNP Complex and Enhances Proviral HIV Gene Expression. Issue 12 (15th May 2015)

• Record Type: Journal Article
• Authors: Mbonye, Uri R.; Wang, Benlian; Gokulrangan, Giridharan; Chance, Mark R.; Karn, Jonathan; Cristea, Ileana M.; Graham, David
• Is Part Of: Proteomics. Volume 15:Issue 12(2015:Jun.)
• Page Start: 2078
• Page End: 2086
• Publication Date: 2015-05-15
• Subjects: Proteins -- Separation -- Periodicals; Bioinformatics -- Periodicals; Proteomics -- Periodicals; Genomes -- Periodicals; Molecular genetics -- Periodicals; 572.605
• DOI: 10.1002/pmic.201500038
• Languages: English
• ISSNs: 1615-9853

Abstract

<abstract abstract-type="main"> <title> <x xml:space="preserve">Abstract</x> </title> <p>Efficient HIV transcription requires P‐TEFb, an essential co‐factor for Tat. In actively replicating cells, P‐TEFb is incorporated into the 7SK snRNP complex together with the repressor protein HEXIM1. Using an affinity purification‐tandem mass spectrometry approach to identify modification sites on HEXIM1 that regulate the sequestration of P‐TEFb by 7SK snRNP, we found that HEXIM1 can be phosphorylated on adjacent residues in a region immediately upstream of the coiled‐coil dimerization domain (Ser268, Thr270, Tyr271, and Tyr274). Phosphomimetic mutations of Tyr271 and Tyr274 disrupted the assembly of P‐TEFb and HEXIM1 into the 7SK snRNP complex. Although Y271E/Y274E did not adversely affect the nuclear localization pattern of HEXIM1, it induced the redistribution of the CDK9 subunit of P‐TEFb into the cytoplasm. By contrast, the Y271F/Y274F HEXIM1 mutant assembled normally with P‐TEFb within the 7SK snRNP complex but severely reduced proviral gene expression in T cells in response to activation signals and caused a severe growth defect of Jurkat T cells. Thus, Y271F/Y274F, which cannot be phosphorylated on these residues, appears to block the exchange of active P‐TEFb from the 7SK complex, thereby limiting the level of P‐TEFb below the threshold required to support transcription elongation of the HIV provirus and cellular genes.</p> </abstract>