Doa1 is a Cdc48 adapter that possesses a novel ubiquitin binding domain

Cdc48 (p97/VCP) is an AAA-ATPase molecular chaperone whose cellular functions are facilitated by its interaction with ubiquitin binding cofactors (e.g., Npl4-Ufd1 and Shp1). Several studies have shown that Saccharomyces cerevisiae Doal (Ufd3/Zzz4) and its mammalian homologue, PLAA, interact with Cdc48. However, the function of this interaction has not been determined, nor has a physiological link between these proteins been demonstrated. Herein, we demonstrate that Cdc48 interacts directly with the C-terminal PUL domain of Doal. We find that Dual possesses a novel ubiquitin binding domain (we propose the name PFU domain, for PLAA family ubiquitin binding domain), which appears to be necessary for Dual function. Our data suggest that the PUL and PFU domains of Doal promote the formation of a Doal-Cdc48-ubiquitin ternary complex, potentially allowing for the recruitment of ubiquitinated proteins to Cdc48. DOA1 and CDC48 mutations are epistatic, suggesting that their interaction is physiologically relevant. Lastly, we provide evidence of functional conservation within the PLAA family by showing that a human-yeast chimera binds to ubiquitin and complements doa1 Delta phenotypes in yeast. Combined, our data suggest that Doal plays a physiological role as a ubiquitin binding cofactor of Cdc48 and that human PLAA may play an analogous role via its interaction with p97/VCP.