Prenylation-dependent membrane localization of a deubiquitinating enzyme and its role in regulating G protein-mediated signaling in yeast.

Miy1 is a highly conserved deubiquitinating enzyme in yeast with MINDY1 as its human homolog. Miy1 is known to act on K48-linked polyubiquitin chain, but its biological function is unknown. Miy1 has a putative prenylation site, suggesting it as a membrane-associated protein that may contribute to the regulation of cell signaling. Here, we demonstrate that Miy1 is localized in the plasma membrane and nuclear periphery. Mutating the putative prenylation site in Miy1 or disrupting the farnesyltransferase activity impairs its localization. Consistent with a role of Miy1 in regulating the ubiquitination status of membrane proteins, the miy1Δ mutants exhibit a higher level of ubiquitinated conjugates at the plasma membrane. To examine a role of Miy1 in regulating cell signaling across plasma membrane, we focused on the pheromone response, as both Ste2, the receptor for mating pheromone, and Gpa1, the cognate Gα protein of Ste2, are well known to be regulated by ubiquitination. We find that Miy1 interacts with Gpa1, regulates its level of ubiquitination and abundance. Pheromone-induced MAP kinase Fus3 activation is also altered in the MIY1-disrupted mutants. The findings demonstrate that Miy1 is a membrane-associated deubiquitinating enzyme and a regulator of G protein-mediated signaling.