E3 ubiquitin ligase Mule ubiquitinates Miz1 and is required for TNFα-induced JNK activation

The zinc finger transcription factor Miz1 is a negative regulator of TNFα-induced JNK activation and cell death through inhibition of TRAF2 K63-polyubiquitination in a transcription-independent manner. Upon TNFα stimulation, Miz1 undergoes K48-linked polyubiquitination and proteasomal degradation, thereby relieving its inhibition. However, the underling regulatory mechanism is not known. Here, we report that HECT-domain-containing Mule is the E3 ligase that catalyzes TNFα-induced Miz1 polyubiquitination. Mule is a Miz1-associated protein and catalyzes its K48-linked polyubiquitination. TNFα-induced polyubiquitination and degradation of Miz1 were inhibited by silencing of Mule and were promoted by ectopic expression of Mule. The interaction between Mule and Miz1 was promoted by TNFα independently of the pox virus and zinc finger domain of Miz1. Silencing of Mule stabilized Miz1, thereby suppressing TNFα-induced JNK activation and cell death. Thus, our study reveals a molecular mechanism by which Mule regulates TNFα-induced JNK activation and apoptosis by catalyzing the polyubiquitination of Miz1.