Abstract
Peptide linkers that tether WWP2 WW domains lock the HECT domain in an inactive conformation.
Major finding: Peptide linkers that tether WWP2 WW domains lock the HECT domain in an inactive conformation.
Mechanism: Phosphorylation at both ends of the WW2–WW3 linker promotes WWP2 activity and substrate ubiquitination.
Impact: WWP2 mutations found in cancer relieve linker-mediated autoinhibition to promote substrate ubiquitination.
HECT E3 ubiquitin ligases, including WWP2, catalyze ubiquitination of many transcription factors and signaling proteins. Thus, deregulated HECT activity can promote cancer and immune disorders. The WWP2 protein is comprised of four WW domains separated by linkers followed by a HECT domain formed by a large N-lobe that interacts with E2 proteins connected to a smaller C-lobe that catalyzes ubiquitin transfer. A conformational change allows the HECT domain to switch between catalytically active and inactive states; however, the molecular mechanisms underlying the transition are not well defined. Chen and colleagues investigated the regulation of WWP2, which ubiquitinates the tumor suppressor PTEN to promote its degradation. WWP2 catalytic activity was autoinhibited by the WW2–WW3 linker in the full-length protein. Determination of the X-ray crystal structure revealed that the WW2–WW3 linker promoted the catalytically inactive T-shape HECT conformation, with the linker blocking ubiquitin binding to the HECT exosite to prevent stimulation of WWP2 activity. Trp358 in the C-terminus of the WW2 domain mediates the interaction with the HECT domain, and the WWP2W358L mutation, which has been observed in cancer, increased the activity of WWP2. The hinge-loop residues Met752 and Gln753 also interact with the WW2–WW3 linker, and mutations in these residues, WWP2M752T and WWP2Q753L, have also been linked to cancer. Like WWP2W358L, these mutations increased WWP2 activity. Mechanistically, WWP2 was activated by phosphorylation of tyrosine 369 and 392 at each end of the WW2–WW3 linker. These phosphorylations loosened the interactions between the WW2–WW3 linker and the hinge and ubiquitin-binding exosite, allowing ubiquitin binding to the HECT exosite, E2-mediated WWP2 ubiquitination, and substrate ubiquitination. The finding that the peptide linker tethering the WW2–WW3 domains can promote autoinhibition of HECT E3 ligase catalytic activity reveals the mechanism by which HECT mutations found in tumors may promote HECT activity and cancer.
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