Ozz-E3, A Muscle-Specific Ubiquitin Ligase, Regulates β-Catenin Degradation during Myogenesis

Abstract

The identities of the ubiquitin-ligases active during myogenesis are largely unknown. Here we describe a RING-type E3 ligase complex specified by the adaptor protein, Ozz, a novel SOCS protein that is developmentally regulated and expressed exclusively in striated muscle. In mice, the absence of Ozz results in overt maturation defects of the sarcomeric apparatus. We identified beta-catenin as one of the target substrates of the Ozz-E3 in vivo. In the differentiating myofibers, Ozz-E3 regulates the levels of sarcolemma-associated beta-catenin by mediating its degradation via the proteasome. Expression of beta-catenin mutants that reduce the binding of Ozz to endogenous beta-catenin leads to Mb-beta-catenin accumulation and myofibrillogenesis defects similar to those observed in Ozz null myocytes. These findings reveal a novel mechanism of regulation of Mb-beta-catenin and the role of this pool of the protein in myofibrillogenesis, and implicate the Ozz-E3 ligase in the process of myofiber differentiation.

Alix Protein Is Substrate of Ozz-E3 Ligase and Modulates Actin Remodeling in Skeletal Muscle

“…Antibodies and Reagents-Rabbit anti-Ozz antibody was prepared as described (29). The antibody was diluted 1:500 for immunoblotting and 1:10 for immunofluorescence.…”

“…of Genetics, St. Jude Children's Research Hospital) for putative Ozz-binding partners was performed as described previously (29,30). Three cDNA clones encoding Alix were isolated, the smallest of which encoded a fragment from 424 to 868 amino acids.…”

“…We now report that in muscle cells Alix is a substrate of the RING-type ubiquitin ligase, Ozz-E3. Ozz (also known as Neurl2 or Neutralized-like protein 2) is a member of the SOCS (suppressor of cytokine signaling) family of proteins and is the substrate-recognition module of Ozz-E3 (29). This ligase complex includes, beside Ozz, Elongin B/C (Elo B/C), Rbx1, and Cullin 5 (Cul5) (29).…”

“…Ozz (also known as Neurl2 or Neutralized-like protein 2) is a member of the SOCS (suppressor of cytokine signaling) family of proteins and is the substrate-recognition module of Ozz-E3 (29). This ligase complex includes, beside Ozz, Elongin B/C (Elo B/C), Rbx1, and Cullin 5 (Cul5) (29). We have shown previously that Ozz-E3 plays an active role in myofiber differentiation and maturation, during embryogenesis and muscle regeneration, by targeting and ubiquitinating two proteins within macromolecular complexes as follows: the sarcolemma-associated pool of ␤-catenin, and the sarcomeric embryonic myosin heavy chain (MyHC emb ) (29,30).…”

“…This ligase complex includes, beside Ozz, Elongin B/C (Elo B/C), Rbx1, and Cullin 5 (Cul5) (29). We have shown previously that Ozz-E3 plays an active role in myofiber differentiation and maturation, during embryogenesis and muscle regeneration, by targeting and ubiquitinating two proteins within macromolecular complexes as follows: the sarcolemma-associated pool of ␤-catenin, and the sarcomeric embryonic myosin heavy chain (MyHC emb ) (29,30). Ozz Ϫ/Ϫ mice develop overt sarcomeric defects, which we attribute in part to impaired turnover of cadherin-associated ␤-catenin and to incomplete exchange of embryonic MyHC isoforms with their neonatal and adult counterparts within the sarcomere, during myofiber differentiation (29,30).…”