Non–SCF-type F-box protein Roy1/Ymr258c interacts with a Rab5-like GTPase Ypt52 and inhibits Ypt52 function

摘要

Skp1/Cul1/F-box (SCF)–type F-box proteins are a component of the Cullin-RING SCF ubiquitin E3 ligase, which is involved in numerous cellular processes. However, the function of non–SCF-type F-box proteins remains largely unknown. The Rab5-like small guanosine 5′-triphosphatase Vps21/Ypt51 is a key regulator of intracellular transportation; however, deletion of its isoforms, Ypt52 and Ypt53, results in only a modest inhibition of intracellular trafficking. The function of these proteins therefore remains largely elusive. Here we analyze the role of a previously uncharacterized non–SCF-type F-box protein, Roy1/Ymr258c, in cell growth and intracellular transport in Saccharomyces cerevisiae. Roy1 binds to Ypt52 under physiological conditions, and Skp1 is indispensable for the association of Roy1 with Ypt52. The vps21Δ yeast cells exhibit severe deficiencies in cell growth and intracellular trafficking, whereas simultaneous deletion of roy1 alleviates the defects caused by deletion of vps21. However, additional disruption of ypt52 in roy1Δvps21Δ cells largely suppresses the cell growth and trafficking observed in roy1Δvps21Δ cells. We demonstrate that Roy1 interacts with guanosine 5′-diphosphate–bound and nucleotide-free Ypt52 and thereby inhibits the formation of guanosine 5′-triphosphate–bound, active Ypt52. These results thus indicate that Roy1 negatively modulates cell viability and intracellular transport by suppressing Ypt52.