An Arginine-Rich Motif of Ring Finger Protein 4 (RNF4) Oversees the Recruitment and Degradation of the Phosphorylated and SUMOylated KAP1/TRIM28 during Genotoxic Stress [Signal Transduction]

June 6th, 2014 by Kuo, C.-Y., Li, X., Kong, X.-Q., Luo, C., Chang, C.-C., Chung, Y., Shih, H.-M., Li, K. K., Ann, D. K.

KAP1 is a universal transcriptional co-repressor that undergoes multiple post-translational modifications (PTMs), including SUMOylation and Serine (S)824-phosphorylation. However, the functional interplay of KAP1 PTMs in regulating KAP1 turnover during DNA damage response (DDR) remains unclear. To decipher the role and crosstalk of multiple KAP1 PTMs, we show herein that DNA double-strand breaks (DSBs)-induced KAP1 S824-phosphorylation promoted the recruitment of SUMO-targeted Ub E3 ligase (STUbL), RNF4 and subsequent RNF4-mediated, SUMO-dependent degradation. Besides SUMO-interacting motif (SIM), a previously unrecognized, but evolutionarily conserved, Arginine-rich motif (ARM) in RNF4 acts as a novel recognition motif for a selective target recruitment. Results from combined mutagenesis and computational modeling studies suggested that RNF4 utilizes a concerted bi-modular recognition, namely SIM for K676-SUMOylation and ARM for pS824 of simultaneously phosphorylated and SUMOylated KAP1 (pS824-SUMO-KAP1). Furthermore, we proved that arginine (R)73 and 74 within the ARM of RNF4 are required for efficient recruitment to KAP1 or accelerated degradation of PML under stress. In parallel, results of bi-molecular fluorescence complementation assays validated the role of ARM in recognizing pS824 in living cells. Taken together, we establish that ARM is required for RNF4 to efficiently target pS824-SUMO-KAP1, conferring ubiquitin K48-mediated proteasomal degradation in the context of DSBs. The conservation of such motif may possibly explain the requirement for timely substrate selectivity determination among a myriad of SUMOylated proteins under stress conditions. Thus, ARM dynamically regulates the SIM-dependent recruitment of targets to RNF4, which could be critical to dynamically fine-tune the abundance of pS824-SUMO-KAP1, and potentially other SUMOylated protein during DDR.
  • Posted in Journal of Biological Chemistry, Publications
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