A Novel Conserved Isoform of the Ubiquitin Ligase UFD2a/UBE4B Is Expressed Exclusively in Mature Striated Muscle Cells

Yeast Ufd2p was the first identified E4 multiubiquitin chain assembly factor. Its vertebrate homologues later referred to as UFD2a, UBE4B or E4B were also shown to have E3 ubiquitin ligase activity. UFD2a function in the brain has been well established in vivo, and in vitro studies have shown that its activity is essential for proper condensation and segregation of chromosomes during mitosis. Here we show that 2 alternative splice forms of UFD2a, UFD2a-7 and -7/7a, are expressed sequentially during myoblast differentiation of C2C12 cell cultures and during cardiotoxin-induced regeneration of skeletal muscle in mice. UFD2a-7 contains an alternate exon 7, and UFD2a-7/7a, the larger of the 2 isoforms, contains an additional novel exon 7a. Analysis of protein or mRNA expression in mice and zebrafish revealed that a similar pattern of isoform switching occurs during developmental myogenesis of cardiac and skeletal muscle. In vertebrates (humans, rodents, zebrafish), UFD2a-7/7a is expressed only in mature striated muscle. This unique tissue specificity is further validated by the conserved presence of 2 muscle-specific splicing regulatory motifs located in the 3' introns of exons 7 and 7a. UFD2a interacts with VCP/p97, an AAA-type ATPase implicated in processes whose functions appear to be regulated, in part, through their interaction with one or more of 15 previously identified cofactors. UFD2a-7/7a did not interact with VCP/p97 in yeast 2-hybrid experiments, which may allow the ATPase to bind cofactors that facilitate its muscle-specific functions. We conclude that the regulated expression of these UFD2a isoforms most likely imparts divergent functions that are important for myogenisis.