RNF4-Dependent Oncogene Activation by Protein Stabilization

Abstract

SUMMARYUbiquitylation regulates signaling pathways critical for cancer development and, in many cases, targets proteins for degradation. Here, we report that ubiquitylation by RNF4 stabilizes otherwise short-lived oncogenic transcription factors, including β-catenin, Myc, c-Jun, and the Notch intracellular-domain (N-ICD) protein. RNF4 enhances the transcriptional activity of these factors, as well as Wnt- and Notch-dependent gene expression. While RNF4 is a SUMO-targeted ubiquitin ligase, protein stabilization requires the substrate’s phosphorylation, rather than SUMOylation, and binding to RNF4’s arginine-rich motif domain. Stabilization also involves generation of unusual polyubiquitin chains and docking of RNF4 to chromatin. Biologically, RNF4 enhances the tumor phenotype and is essential for cancer cell survival. High levels of RNF4 mRNA correlate with poor survival of a subgroup of breast cancer patients, and RNF4 protein levels are elevated in 30% of human colon adenocarcinomas. Thus, RNF4-dependent ubiquitylation translates transient phosphorylation signal(s) into long-term protein stabilization, resulting in enhanced oncoprotein activation.

Stabilization of nuclear oncoproteins by RNF4 and the ubiquitin system in cancer

“…In a recent study, 5 we identified that RNF4, a ubiquitin ligase, and the ubiquitin system is part of the machinery involved in stabilization, rather than degradation, of selected short-lived oncoproteins such as b-Catenin, c-Myc, c-Jun, and Notch intracellular domain protein (N-ICD): RNF4 potentiates the transcriptional activity of these oncoproteins, enhances the tumorigenic properties of cancer cells, and is essential for cancer cell survival. Importantly, high RNF4 levels correlate with poor outcome in a subset of cancers.…”

“…Importantly, high RNF4 levels correlate with poor outcome in a subset of cancers. 5 RNF4 belongs to a small group of RING (really interesting new gene) ubiquitin ligases termed SUMO-Targeted Ubiquitin ligases (STUbL). Conserved from yeast to man, these ligases bind to SUMO chains of SUMOylated proteins via multiple SIM domains at the N-termini of the protein and catalyze their ubiquitylation via the RING domain, thus physically connecting the ubiquitin and SUMO pathways.…”

“…These p-oncoproteins are recognized by RNF4 via a short arginine motif (ARM) located at the N-terminal part of RNF4 downstream of the SIM motifs. 5,7 Mutation of these phosphorylation sites or the ARM region prevented binding, ubiquitylation, stabilization, and transcriptional enhancement of oncoproteins by RNF4. Moreover, RNF4 stabilized and enhanced the transcriptional activity of otherwise stable oncoprotein mutants such as b-catenin S33A and c-Myc S62A .…”

“…RNF4 stabilizing activity also requires its association with nucleosomes. A point mutation, K179D, that blocks this association prevents ubiquitylation, stabilization, and transcriptional enhancement, 5,6 suggesting that RNF4 acts on, or in the vicinity of, chromatin. Nevertheless, how atypical ubiquitylation and association with nucleosomes results in oncoprotein potentiation is not clear.…”

“…Likewise, high RNF4 mRNA levels correlated with poor outcome in a cohort of breast cancer patients with a relatively less severe type of breast cancer (ER C , luminal type A). 5 In summary, RNF4 fits into a class of genes termed "nononcogene addiction" (NOA) genes encoding enzymes and proteins that are essential for the activity of oncogenes or negatively regulate tumor suppressor functions. 10 Generally, NOA genes are not oncogenic on their own but are crucial for tumor progression and maintaining the cancerous state.…”