ACTIVITY OF UBIQUITIN-DEPENDENT PATHWAY IN RESPONSE TO OXIDATIVE STRESS - UBIQUITIN-ACTIVATING ENZYME IS TRANSIENTLY UP-REGULATED

Relations between the ubiquitin pathway and cellular stress have been noted, but data regarding responses of the ubiquitin pathway to oxidat ive stress are scanty. This paper documents the response of this pathw ay to oxidative stress in lens cells. A brief exposure of lens epithel ial cells to physiologically relevant levels of H2O2 induces a transie nt increase in activity of the ubiquitin-dependent pathway. Ubiquitin conjugation activity was maximal and increased 3.5-9.2-fold over the a ctivity noted in untreated cells by 4 h after removal of H2O2. By 24 h after removal of H2O2, ubiquitin conjugation activity returned to the level noted in untreated cells. In parallel to the changes in ubiquit in conjugation activity, the activity of ubiquitin-activating enzyme ( E1), as determined by thiol ester formation, increased 2-6.7-fold duri ng recovery from oxidation, Addition of exogenous E1 resulted in an in crease in ubiquitin conjugation activity and in the levels of ubiquiti n carrier protein (E2)-ubiquitin thiol esters in both the untreated ce lls and the H2O2-treated cells, These data suggest that E1 is the rate -limiting enzyme in the ubiquitin conjugation process and. that the in creases in ubiquitin conjugation activity which are induced upon recov ery from oxidation are primarily due to increased E1. activity. The ox idation-and recovery-induced up-regulation of E1 activity is primarily due to post-synthetic events, Substrate availability and up-regulatio n of E2 activities also appear to be related to the enhancement in ubi quitinylation upon recovery from oxidative stress, The oxidation-induc ed increases in ubiquitin conjugation activity were associated with an increase in intracellular proteolysis, suggesting that the transient increase in ubiquitinylation noted upon recovery from oxidative stress may play a role in removal of damaged proteins from the cells.