MEKK1-Dependent Activation of the CRL4 Complex Is Important for DNA Damage-Induced Degradation of p21 and DDB2 and Cell Survival

Abstract

Cullin-4 ubiquitin ligase (CRL4) complexes are differentially composed and highly dynamic protein assemblies that control many biological processes, including the global genome nucleotide excision repair (GG-NER) pathway. Here, we identified the ki-nase mitogen-activated protein kinase kinase kinase 1 (MEKK1) as a novel constitutive interactor of a cytosolic CRL4 complex that disassembles after DNA damage due to the caspase-mediated cleavage of MEKK1. The kinase activity of MEKK1 was important to trigger autoubiquitination of the CRL4 complex by K48-and K63-linked ubiquitin chains. MEKK1 knockdown prohibited DNA damage-induced degradation of the CRL4 component DNA-damage binding protein 2 (DDB2) and the CRL4 substrate p21 and also cell recovery and survival. A ubiquitin replacement strategy revealed a contribution of K63-branched ubiquitin chains for DNA damage-induced DDB2/p21 decay, cell cycle regulation, and cell survival. These data might also have implications for cancer, as frequently occurring mutations of MEKK1 might have an impact on genome stability and the therapeutic efficacy of CRL4-dependent immunomodulatory drugs such as thalidomide derivatives. C ullin-RING ligases (CRLs) are the largest family of ubiquitin E3 ligases and are composed of modular protein subunits (1). The human genome encodes seven cullin proteins which function as scaffolding proteins and associate with a RING finger protein (RING of cullins [ROC1/RBX1 or ROC2/RBX2]) to recruit the E2 ubiquitin-conjugating enzyme (2). The N-terminal part of cullins binds to a number of specific adapter proteins that, in turn, bind to substrate receptors. Cullin-4 (CUL4A and the highly related protein CUL4B) typically use the DNA-damage binding protein 1 (DDB1) adaptor to interact with a variety of DDB1-cullin-4-associated factors (DCAFs) that make contact to substrate proteins or that can be also ubiquitinated and degraded by themselves (3). Activation of the CRL4 complex needs the covalent but reversible ligation of the ubiquitin-like protein NEDD8 (neural precursor cell-expressed developmentally downregulated 8) to a conserved lysine in the C terminus of the cullin (4). The CRL4 complex is involved in the regulation of several processes, including transcription and embryonic development (5, 6). This complex also participates in the multistep global genome nucleotide excision repair (GG-NER) pathway, where a heterodimer of DDB1 and DDB2 recognizes photolesions in the DNA to promote several consecutive steps in the early phase of DNA repair (3, 7). While CRL4-mediated monoubiquitination of his-tone proteins leads to their eviction around the damaged site, DDB1/DDB2 allow the subsequent recruitment of the xeroderma pigmentosum complementation group C Citation Bacher S, Stekman H, Farah CM, Karger A, Kracht M, Schmitz ML. 2021. MEKK1-dependent activation of the CRL4 complex is important for DNA damage-induced degradation of p21 and DDB2 and cell survival. Mol Cell Biol 41:e00081-21. https://doi.