Reversible inactivation of deubiquitinases by reactive oxygen species in vitro and in cells

Abstract

In eukaryotes, deubiquitinases (DUBs) remove ubiquitin conjugates from diverse substrates, altering their stabilities, localizations or activities. Here we show that many DUBs of the USP and UCH subfamilies can be reversibly inactivated upon oxidation by reactive oxygen species in vitro and in cells. Oxidation occurs preferentially on the catalytic cysteine, abrogating the isopeptide-cleaving activity without affecting these enzymes’ affinity to ubiquitin. Sensitivity to oxidative inhibition is associated with DUB activation wherein the active site cysteine is converted to a deprotonated state prone to oxidation. We demonstrate that this redox regulation is essential for mono-ubiquitination of proliferating-cell nuclear antigen in response to oxidative DNA damage, which initiates a DNA damage-tolerance programme. These findings establish a novel mechanism of DUB regulation that may be integrated with other redox-dependent signalling circuits to govern cellular adaptation to oxidative stress, a process intimately linked to aging and cancer.

Mechanisms of regulation and diversification of deubiquitylating enzyme function

“…The DUBs USP32, MINDY1 and MINDY2 (also known as FAM63A and FAM63B , respectively) are also predicted to be modified with lipids, which might account for the association of USP32 with intracellular membranes (Abdul Rehman et al, 2016;Akhavantabasi et al, 2010). Finally, reversible oxidation of the catalytic cysteine of DUBs, which inhibits their activity, has also been reported (Cotto-Rios et al, 2012;Kulathu et al, 2013;Lee et al, 2013); this could provide an attractive model for how ubiquitin signals can be amplified when DUB activity is reversibly inhibited in a spatio-temporal manner in response to production of reactive oxygen species.…”

Deubiquitylases From Genes to Organism

“…Ubiquitylation of Ataxin-3 in the vicinity of the catalytic site enhances its activity (255), whilst sumoylation of USP25 inhibits activity (173). Interestingly, the catalytic cysteine of the cysteine protease DUBs is widely subject to reversible inactivation by modification with reactive oxygen species (ROS), similarly to protein tyrosine phosphatases (48,62,132,144,217).…”

The Expanding Landscape of the Thiol Redox Proteome

“…Most thiol redox modifications can be formed by the non-enzymatic reactions of reactive oxygen/nitrogen/sulfur species (ROS/RNS/RSS) with protein thiols, which has led to the widespread notion that these types of modifications are randomly distributed across the cysteine proteome. However, emerging evidence suggests that thiol redox modifications are well-controlled, sitespecific cellular events, which play important roles in regulation of diverse protein functions, including catalytic or ligand binding activities (5-7), protein-protein interactions (8,9), and protein stability (10,11).…”