A TSG101/MDM2 regulatory loop modulates MDM2 degradation and MDM2/p53 feedback control

The p53 tumor suppressor protein and the MDM2 oncoprotein form a feedback-c ontrol loop that up-regulates cellular MDM2 production, blocks p53 activity , and promotes p53 decay, tsg101 was discovered as a gene whose deficiency results in neoplastic transformation of NIH 3T3 cells and the ability to ge nerate metastatic tumors in nude mice. Its protein product contains a domai n, Ubc, characteristic of the catalytic domain of ubiquitin conjugase (E2) enzymes but lacking an active-site cysteine crucial for ubiquitin conjugase activity. Here we report that TSG101 participates with MDM2 in an autoregu latory loop that modulates the cellular levels of both proteins, and also o f p53, by affecting protein decay, We show that the Ubc domain of TSG101 in terferes with ubiquitination of MDM2, that TSG101 inhibits MDM2 decay and e levates its steady-state level, and that these events are associated with d own-regulation of p53 protein. Conversely, pulse-chase and Western blot exp eriments in wild-type and mutant fibroblasts indicate that elevation of MDM 2 by overexpression of wild-type p53, by amplification of the endogenous MD M2 gene, or by transfection of MDM2-expressing constructs promotes TSG101 l oss, which we show occurs by 26S proteasome-dependent decay. Our results id entify TSG101 as both a regulator of, and target of, MDM2/p53 circuitry.