TY - JOUR
T1 - SHMT2 and the BRCC36/BRISC deubiquitinase regulate HIV-1 Tat K63-ubiquitylation and destruction by autophagy
AU - Xu, Muyu
AU - Moresco, James J.
AU - Chang, Max
AU - Mukim, Amey
AU - Smith, Davey
AU - Diedrich, Jolene K.
AU - Yates, John R.
AU - Jones, Katherine A.
N1 - Funding Information:
MX gratefully acknowledges postdoctoral fellowship support from the Margaret T. Morris Foundation. This work was funded from grants to KAJ from the California HIV/AIDS Research Program and the NIH (AI044615). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We thank Dr. Jonathan Karn (Case Western Reserve) for 2D10 cells, The Salk Next-Generation Sequencing and Mass Spectrometry core facilities for assistance with RNA-seq and proteomics analyses, Gemma Caballero (UCSD) for help with the HIV infectivity (blue cell) assays, and Dr. Alan Frankel (UCSF) for the STREP-Tat plasmids (WT, ∆K and ∆K (+K41)).
Publisher Copyright:
© 2018 Xu et al. http://creativecommons.org/licenses/by/4.0/
PY - 2018/5
Y1 - 2018/5
N2 - HIV-1 Tat is a key regulator of viral transcription, however little is known about the mechanisms that control its turnover in T cells. Here we use a novel proteomics technique, called DiffPOP, to identify the molecular target of JIB-04, a small molecule compound that potently and selectively blocks HIV-1 Tat expression, transactivation, and virus replication in T cell lines. Mass-spectrometry analysis of whole-cell extracts from 2D10 Jurkat T cells revealed that JIB-04 targets Serine Hydroxymethyltransferase 2 (SHMT2), a regulator of glycine biosynthesis and an adaptor for the BRCC36 K63Ub-specific deubiquitinase in the BRISC complex. Importantly, knockdown of SHMT1,2 or BRCC36, or exposure of cells to JIB-04, strongly increased Tat K63Ub-dependent destruction via autophagy. Moreover, point mutation of multiple lysines in Tat, or knockdown of BRCC36 or SHMT1,2, was sufficient to prevent destruction of Tat by JIB-04. We conclude that HIV-1 Tat levels are regulated through K63Ub-selective autophagy mediated through SHMT1,2 and the BRCC36 deubiquitinase.
AB - HIV-1 Tat is a key regulator of viral transcription, however little is known about the mechanisms that control its turnover in T cells. Here we use a novel proteomics technique, called DiffPOP, to identify the molecular target of JIB-04, a small molecule compound that potently and selectively blocks HIV-1 Tat expression, transactivation, and virus replication in T cell lines. Mass-spectrometry analysis of whole-cell extracts from 2D10 Jurkat T cells revealed that JIB-04 targets Serine Hydroxymethyltransferase 2 (SHMT2), a regulator of glycine biosynthesis and an adaptor for the BRCC36 K63Ub-specific deubiquitinase in the BRISC complex. Importantly, knockdown of SHMT1,2 or BRCC36, or exposure of cells to JIB-04, strongly increased Tat K63Ub-dependent destruction via autophagy. Moreover, point mutation of multiple lysines in Tat, or knockdown of BRCC36 or SHMT1,2, was sufficient to prevent destruction of Tat by JIB-04. We conclude that HIV-1 Tat levels are regulated through K63Ub-selective autophagy mediated through SHMT1,2 and the BRCC36 deubiquitinase.
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U2 - 10.1371/journal.ppat.1007071
DO - 10.1371/journal.ppat.1007071
M3 - Article
C2 - 29791506
AN - SCOPUS:85047982099
SN - 1553-7366
VL - 14
JO - PLoS pathogens
JF - PLoS pathogens
IS - 5
M1 - e1007071
ER -