The Cullin-RING E3 Ubiquitin Ligase CRL4−DCAF1 Complex Dimerizes via a Short Helical Region in DCAF1

MuRF1 is an E3 ubiquitin ligase central to muscle catabolism. It belongs to the TRIM protein family characterized by a tripartite fold of RING, B-box and coiled-coil (CC) motifs, followed by variable C-terminal domains. The CC motif is hypothesized to be responsible for domain organization in the fold as well as for high-order assembly into functional entities. But data on CC from this family that can clarify the structural significance of this motif are scarce. We have characterized the helical region from MuRF1 and show that, contrary to expectations, its CC domain assembles unproductively, being the B2- and COS-boxes in the fold (respectively flanking the CC) that promote a native quaternary structure. In particular, the C-terminal COS-box seemingly forms an α-hairpin that packs against the CC, influencing its dimerization. This shows that a C-terminal variable domain can be tightly integrated within the conserved TRIM fold to modulate its structure and function. Furthermore, data from transfected muscle show that in MuRF1 the COS-box mediates the in vivo targeting of sarcoskeletal structures and points to the pharmacological relevance of the COS domain for treating MuRF1-mediated muscle atrophy.

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Identification of a PGXPP degron motif in dishevelled and structural basis for its binding to the E3 ligase KLHL12

Open Biology ◽

10.1098/rsob.200041 ◽

2020 ◽

Vol 10 (6) ◽

pp. 200041 ◽

Cited By ~ 1

Author(s):

Zhuoyao Chen ◽

Gregory A. Wasney ◽

Sarah Picaud ◽

Panagis Filippakopoulos ◽

Masoud Vedadi ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Molecular Mechanisms ◽

Hydrophobic Interactions ◽

E3 Ubiquitin Ligase ◽

Structural Basis ◽

E3 Ligases ◽

Kelch Domain ◽

Ring E3 Ligases ◽

Ring E3 ◽

New Protein

Wnt signalling is dependent on dishevelled proteins (DVL1-3), which assemble an intracellular Wnt signalosome at the plasma membrane. The levels of DVL1-3 are regulated by multiple Cullin-RING E3 ligases that mediate their ubiquitination and degradation. The BTB-Kelch protein KLHL12 was the first E3 ubiquitin ligase to be identified for DVL1-3, but the molecular mechanisms determining its substrate interactions have remained unknown. Here, we mapped the interaction of DVL1-3 to a ‘PGXPP' motif that is conserved in other known partners and substrates of KLHL12, including PLEKHA4, PEF1, SEC31 and DRD4. To determine the binding mechanism, we solved a 2.4 Å crystal structure of the Kelch domain of KLHL12 in complex with a DVL1 peptide that bound with low micromolar affinity. The DVL1 substrate adopted a U-shaped turn conformation that enabled hydrophobic interactions with all six blades of the Kelch domain β-propeller. In cells, the mutation or deletion of this motif reduced the binding and ubiquitination of DVL1 and increased its stability confirming this sequence as a degron motif for KLHL12 recruitment. These results define the molecular mechanisms determining DVL regulation by KLHL12 and establish the KLHL12 Kelch domain as a new protein interaction module for a novel proline-rich motif.

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