RESUMO
The interplay between E2 and E3 enzymes regulates the polyubiquitination of substrates in eukaryotes. Among the several RING-domain E3 ligases in humans, many utilize two distinct E2s for polyubiquitination. For example, the cell cycle regulatory E3, human anaphase-promoting complex/cyclosome (APC/C), relies on UBE2C to prime substrates with ubiquitin (Ub) and on UBE2S to extend polyubiquitin chains. However, the potential coordination between these steps in ubiquitin chain formation remains undefined. While numerous studies have unveiled how RING E3s stimulate individual E2s for Ub transfer, here we change perspective to describe a case where the chain-elongating E2 UBE2S feeds back and directly stimulates the E3 APC/C to promote substrate priming and subsequent multiubiquitination by UBE2C. Our work reveals an unexpected model for the mechanisms of RING E3-dependent ubiquitination and for the diverse and complex interrelationship between components of the ubiquitination cascade.
Assuntos
Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ciclossomo-Complexo Promotor de Anáfase/química , Ciclossomo-Complexo Promotor de Anáfase/genética , Subunidade Apc4 do Ciclossomo-Complexo Promotor de Anáfase/química , Subunidade Apc4 do Ciclossomo-Complexo Promotor de Anáfase/genética , Subunidade Apc4 do Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Citidina Trifosfato/metabolismo , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Células HeLa , Humanos , Poliubiquitina/metabolismo , Ubiquitina/metabolismo , Enzimas de Conjugação de Ubiquitina/genética , Ubiquitina-Proteína Ligases/química , UbiquitinaçãoRESUMO
The Anaphase Promoting Complex/Cyclosome (APC/C) is a ubiquitin E3 ligase that functions as the gatekeeper to mitotic exit. APC/C activity is controlled by an interplay of multiple pathways during mitosis, including the spindle assembly checkpoint (SAC), that are not yet fully understood. Here, we show that sumoylation of the APC4 subunit of the APC/C peaks during mitosis and is critical for timely APC/C activation and anaphase onset. We have also identified a functionally important SUMO interacting motif in the cullin-homology domain of APC2 located near the APC4 sumoylation sites and APC/C catalytic core. Our findings provide evidence of an important regulatory role for SUMO modification and binding in affecting APC/C activation and mitotic exit.