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Title: Role of a non-canonical surface of Rad6 in ubiquitin conjugating activity

Authors:
; ; ; ; ;  [1];  [2]
  1. JHU
  2. (
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
National Institutes of Health (NIH)
OSTI Identifier:
1224474
Resource Type:
Journal Article
Resource Relation:
Journal Name: Nucleic Acids Research; Journal Volume: 43; Journal Issue: (18) ; 10, 2015
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Kumar, Pankaj, Magala, Pearl, Geiger-Schuller, Kathryn R., Majumdar, Ananya, Tolman, Joel R., Wolberger, Cynthia, and JHU-MED). Role of a non-canonical surface of Rad6 in ubiquitin conjugating activity. United States: N. p., 2016. Web. doi:10.1093/nar/gkv845.
Kumar, Pankaj, Magala, Pearl, Geiger-Schuller, Kathryn R., Majumdar, Ananya, Tolman, Joel R., Wolberger, Cynthia, & JHU-MED). Role of a non-canonical surface of Rad6 in ubiquitin conjugating activity. United States. doi:10.1093/nar/gkv845.
Kumar, Pankaj, Magala, Pearl, Geiger-Schuller, Kathryn R., Majumdar, Ananya, Tolman, Joel R., Wolberger, Cynthia, and JHU-MED). 2016. "Role of a non-canonical surface of Rad6 in ubiquitin conjugating activity". United States. doi:10.1093/nar/gkv845.
@article{osti_1224474,
title = {Role of a non-canonical surface of Rad6 in ubiquitin conjugating activity},
author = {Kumar, Pankaj and Magala, Pearl and Geiger-Schuller, Kathryn R. and Majumdar, Ananya and Tolman, Joel R. and Wolberger, Cynthia and JHU-MED)},
abstractNote = {},
doi = {10.1093/nar/gkv845},
journal = {Nucleic Acids Research},
number = (18) ; 10, 2015,
volume = 43,
place = {United States},
year = 2016,
month = 6
}
  • No abstract prepared.
  • Mono- and multi-ubiquitination alters the functions and subcellular localization of many cellular and viral proteins. Viruses can co-opt or actively manipulate the ubiquitin network to support viral processes or suppress innate immunity. Using yeast (Saccharomyces cerevisiae) model host, we show that the yeast Rad6p (radiation sensitive 6) E2 ubiquitin-conjugating enzyme and its plant ortholog, AtUbc2, interact with two tombusviral replication proteins and these E2 ubiquitin-conjugating enzymes could be co-purified with the tombusvirus replicase. We demonstrate that TBSV RNA replication and the mono- and bi-ubiquitination level of p33 is decreased in rad6Δ yeast. However, plasmid-based expression of AtUbc2p could complement bothmore » defects in rad6Δ yeast. Knockdown of UBC2 expression in plants also decreases tombusvirus accumulation and reduces symptom severity, suggesting that Ubc2p is critical for virus replication in plants. We provide evidence that Rad6p is involved in promoting the subversion of Vps23p and Vps4p ESCRT proteins for viral replicase complex assembly. - Highlights: • Tombusvirus p33 replication protein interacts with cellular RAD6/Ubc2 E2 enzymes. • Deletion of RAD6 reduces tombusvirus replication in yeast. • Silencing of UBC2 in plants inhibits tombusvirus replication. • Mono- and bi-ubiquitination of p33 replication protein in yeast and in vitro. • Rad6p promotes the recruitment of cellular ESCRT proteins into the tombusvirus replicase.« less
  • DNA double strand break (DSB) responses depend on the sequential actions of the E3 ubiquitin ligases RNF8 and RNF168 plus E2 ubiquitin-conjugating enzyme Ubc13 to specifically generate histone Lys-63-linked ubiquitin chains in DSB signaling. In this paper, we defined the activated RNF8-Ubc13~ubiquitin complex by x-ray crystallography and its functional solution conformations by x-ray scattering, as tested by separation-of-function mutations imaged in cells by immunofluorescence. The collective results show that the RING E3 RNF8 targets E2 Ubc13 to DSB sites and plays a critical role in damage signaling by stimulating polyubiquitination through modulating conformations of ubiquitin covalently linked to the Ubc13more » active site. Structure-guided separation-of-function mutations show that the RNF8 E2 stimulating activity is essential for DSB signaling in mammalian cells and is necessary for downstream recruitment of 53BP1 and BRCA1. Chromatin-targeted RNF168 rescues 53BP1 recruitment involved in non-homologous end joining but not BRCA1 recruitment for homologous recombination. Finally, these findings suggest an allosteric approach to targeting the ubiquitin-docking cleft at the E2-E3 interface for possible interventions in cancer and chronic inflammation, and moreover, they establish an independent RNF8 role in BRCA1 recruitment.« less