Ube2V2 Is a Rosetta Stone Bridging Redox and Ubiquitin Codes, Coordinating DNA Damage Responses

Posttranslational
modifications (PTMs) are the lingua franca of
cellular communication. Most PTMs are enzyme-orchestrated. However,
the reemergence of electrophilic drugs has ushered mining of unconventional/non-enzyme-catalyzed
electrophile-signaling pathways. Despite the latest impetus toward
harnessing kinetically and functionally privileged cysteines for electrophilic
drug design, identifying these sensors remains challenging. Herein,
we designed “G-REX”a technique that allows controlled
release of reactive electrophiles in vivo. Mitigating toxicity/off-target
effects associated with uncontrolled bolus exposure, G-REX tagged first-responding innate cysteines that bind electrophiles
under true kcat/Km conditions. G-REX identified two allosteric ubiquitin-conjugating
proteinsUbe2V1/Ube2V2sharing a novel privileged-sensor-cysteine.
This non-enzyme-catalyzed-PTM triggered responses specific
to each protein. Thus, G-REX is an unbiased method to identify
novel functional cysteines. Contrasting conventional active-site/off-active-site
cysteine-modifications that regulate target activity,
modification of Ube2V2 allosterically hyperactivated its enzymatically
active binding-partner Ube2N, promoting K63-linked client ubiquitination
and stimulating H2AX-dependent DNA damage response. This work establishes
Ube2V2 as a Rosetta-stone bridging redox and ubiquitin codes to guard
genome integrity.