Abstract
Saccharomyces cerevisiae is amenable to studying membrane traffic by live-cell fluorescence microscopy. We used this system to explore two aspects of cargo protein traffic through prevacuolar endosome (PVE) compartments to the vacuole. First, at what point during Golgi maturation does a biosynthetic vacuolar cargo depart from the maturing cisternae? To address this question, we modified a regulatable fluorescent secretory cargo by adding a vacuolar targeting signal. Traffic of the vacuolar cargo requires the GGA clathrin adaptors, which arrive during the early-to-late Golgi transition. Accordingly, the vacuolar cargo begins to exit the Golgi near the midpoint of maturation, significantly before exit of a secretory cargo. Second, how are cargoes delivered from PVE compartments to the vacuole? To address this question, we tracked biosynthetic and endocytic cargoes after they had accumulated in PVE compartments. The results suggest that stable PVE compartments repeatedly deliver material to the vacuole by a kiss-and-run mechanism.
Highlights
Budding yeast has been instrumental for defining mechanisms of membrane traffic
During the early stage of maturation, one set of resident Golgi membrane proteins recycles with the aid of the COPI vesicle coat, whereas during the late stage of maturation, another set of resident Golgi membrane proteins recycles with the aid of the AP-1 clathrin adaptor
We found previously that the regulatable secretory cargo persists in cisternae through the early-to-late transition of Golgi maturation, and that a fraction of the cargo molecules are recycled within the Golgi in an AP-1-dependent manner (Casler et al, 2019)
Summary
Budding yeast has been instrumental for defining mechanisms of membrane traffic. Genetic screens of Saccharomyces cerevisiae have identified many conserved components of the biosynthetic and endocytic machineries (Kaiser et al, 1997; Novick et al, 1980). Cell Biology known as the trans-Golgi network (TGN), plays an additional role as an early and recycling endosome (Day et al, 2018) According to this view, yeast cells have two types of endosomes: (a) the late Golgi/TGN, and (b) PVE compartments that are typically attached to the vacuole. The conventional view is that biosynthetic cargoes all travel together through the Golgi until reaching a terminal sorting stage (De Matteis and Luini, 2008; Griffiths and Simons, 1986), implying that vacuolar cargoes would remain in a cisterna throughout the maturation process This idea is called into question by studies of GGA dynamics at the Golgi. Evidence presented here suggests that stable PVE compartments deliver their contents to the vacuole by transient kiss-and-run fusion events
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