Biology subjects

Chaugule, V. K.

Publications and source records attributed to Chaugule, V. K..

3 recordsLinked to original sources

bioRxiv2018

Specificity for deubiquitination of monoubiquitinated FANCD2 is driven by the N-terminus of USP1

The DNA damage response depends on ubiquitin signalling to orchestrate DNA repair. The Fanconi Anemia pathway for interstrand crosslink repair, and the translesion synthesis pathway for DNA damage tolerance, both require cycles of monoubiquitination and deubiquitination. The ubiquitin specific protease USP1 regulates both these pathways by deubiquitinating monoubiquitinated PCNA, FANCD2 and FANCI. Loss of USP1 activity gives rise to chromosomal instability. While many USPs hydrolyse ubiquitin-ubiquitin linkages, USP1 targets ubiquitin-substrate conjugates at specific sites. The molecular basis of USP1s specificity for multiple substrates is poorly understood. Here we show that the molecular determinants for substrate deubiquitination by USP1 reside within the highly conserved and extended N-terminus. We find that the N-terminus of USP1 harbours a FANCD2-specific binding sequence required for deubiquitination of K561 on FANCD2. In contrast, the N-terminus is not required for PCNA or FANCI deubiquitination. Furthermore, we show that the N-terminus of USP1 is sufficient to engineer specificity in a more promiscuous USP.

biochemistry↗

bioRxiv2018

Allosteric network in Ube2T drives specificity for RING E3 catalysed ubiquitin signals.

In eukaryotes, DNA damage repair is implemented by a host of proteins that are coordinated by defined molecular signals. One such signal that transpires during the Fanconi Anemia (FA) - interstrand crosslink (ICL) repair pathway is the site-specific monoubiquitination of FANCD2 and FANCI proteins by a large, multi-protein FA core complex. The mechanics for this exquisitely specific monoubiquitin signal has been elusive. Here we show FANCL, the RING E3 module of the FA core complex, allosterically activates its cognate E2 Ube2T for monoubiquitination by a mechanism distinct from the typical RING-based catalysis. FANCL triggers intricate re-wiring of Ube2Ts intra-residue network thus activating the E2 for precision targeting. This network is intrinsically regulated by conserved gates and loops which can be engineered to yield Ube2T variants that enhance FANCD2 ubiquitination by ~30-fold without compromising on target specificity. Finally, we also uncover allosteric networks in other ubiquitin E2s that can be leveraged by RING E3 ligases to drive specific ubiquitination.

biochemistry↗

bioRxiv2017

DHX15 regulates CMTR1-dependent gene expression and cell proliferation

CMTR1 contributes to mRNA cap formation by methylating the O-2 position of the 1st transcribed nucleotide ribose. mRNA cap O-2 methylation has roles in mRNA translation and self-RNA tolerance in innate immunity, however its role in cell physiology is unclear. We report that CMTR1 is recruited to Serine-5 phosphorylated RNA Pol II CTD, facilitating cotranscriptional methylation. We isolated CMTR1 in a complex with DHX15, an RNA helicase functioning in splicing and ribosome biogenesis, and characterised it as a regulator of CMTR1. When bound to DHX15, CMTR1 activity is repressed and prevented from binding to RNA pol II, thus constraining 1st nucleotide methylation to a co-transcriptional event. Conversely CMTR1 activates DHX15 helicase activity and influences its nuclear localisation, which is likely to impact on several nuclear functions. The impact of the CMTR1-DHX15 interaction is complex and will depend on the relative expression of these enzymes and their interactors, and the cellular dependency on different RNA processing pathways. In HCC1806 cells, the DHX15-CMTR1 interaction controls ribosome loading of a subset of mRNAs and impacts on cell proliferation.

cell biology↗