Abstract
Cullin-RING E3 ligases (CRLs) are essential ubiquitylation enzymes that combine a catalytic core built around cullin scaffolds with ∼300 exchangeable substrate adaptors. To ensure robust signal transduction, cells must constantly form new CRLs by pairing substrate-bound adaptors with their cullins, but how this occurs at the right time and place is still poorly understood. Here, we show that formation of individual CRL complexes is a tightly regulated process. Using CUL3KLHL12 as a model, we found that its co-adaptor PEF1-ALG2 initiates CRL3 formation by releasing KLHL12 from an assembly inhibitor at the endoplasmic reticulum, before co-adaptor monoubiquitylation stabilizes the enzyme for substrate modification. As the co-adaptor also helps recruit substrates, its role in CRL assembly couples target recognition to ubiquitylation. We propose that regulators dedicated to specific CRLs, such as assembly inhibitors or co-adaptors, cooperate with target-agnostic adaptor exchange mechanisms to establish E3 ligase complexes that control metazoan development.
Highlights
With the average human protein engaging five partners (Goodsell and Olson, 2000; Huttlin et al, 2017), multisubunit complexes play critical roles in cellular information transfer
Co-adaptor dependent assembly of CUL3KLHL12 Several CRL3 enzymes rely on co-adaptors to bind their targets at specific locations in the cell, but how substrate recognition and ubiquitylation are coordinated is not known
We focused on PEF1-ALG2, a heterodimeric coadaptor that allows CUL3 and its adaptor KLHL12 to monoubiquitylate SEC31A at budding COPII vesicles (McGourty et al, 2016)
Summary
With the average human protein engaging five partners (Goodsell and Olson, 2000; Huttlin et al, 2017), multisubunit complexes play critical roles in cellular information transfer. Illustrating the importance and prevalence of such modular signaling, human cells contain $40 PP2A phosphatases, $30 p97/VCP AAA-ATPases, and $300 Cullin-RING E3 ligases (CRLs) (Bennett et al, 2010; Buchberger et al, 2015; Goguet-Rubio et al, 2020; Reitsma et al, 2017). How these pivotal enzymes achieve their proper composition and select their targets at the right time and place remains a central question in biology. Whereas the adaptor exchange factor CAND1 is known to dismantle used CRLs (Liu et al, 2018; Pierce et al, 2013; Reichermeier et al, 2020; Reitsma et al, 2017), how cells choose the adaptor to build a new E3 ligase is still poorly understood
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