Ypt-Rab GTPases are key regulators of the various steps of intracellular trafficking. Guanine nucleotide-exchange factors (GEFs) regulate the conversion of Ypt-Rabs to the GTP-bound state, in which they interact with effectors that mediate all the known aspects of vesicular transport. An interesting possibility is that Ypt-Rabs coordinate separate steps of the transport pathways. The conserved modular complex TRAPP is a GEF for the Golgi gatekeepers Ypt1 and Ypt31/32 (Refs 5-7). However, it is not known how Golgi entry and exit are coordinated. TRAPP comes in two configurations: the seven-subunit TRAPPI is required for endoplasmic reticulum-to-Golgi transport, whereas the ten-subunit TRAPPII functions in late Golgi. The two essential TRAPPII-specific subunits Trs120 and Trs130 have been identified as Ypt31/32 genetic interactors. Here, we show that they are required for switching the GEF specificity of TRAPP from Ypt1 to Ypt31. Moreover, a trs130ts mutation confers opposite effects on the intracellular localization of these GTPases. We suggest that the Trs120-Trs130 subcomplex joins TRAPP in the late Golgi to switch its GEF activity from Ypt1 to Ypt31/32. Such a 'switchable' GEF could ensure sequential activation of these Ypts, thereby coordinating Golgi entry and exit.

The key regulators of intracellular trafficking, Ypt/Rab GTPases, are stimulated by specific upstream activators and, when activated, recruit specific downstream effectors to mediate membrane-transport events. The yeast Ypt1 and its human functional homolog hRab1 regulate both endoplasmic reticulum (ER)-to-Golgi transport and autophagy. However, it is not clear whether the mechanism by which these GTPases regulate autophagy depends on their well-documented function in ER-to-Golgi transport. Here, we identify Atg11, the preautophagosomal structure (PAS) organizer, as a downstream effector of Ypt1 and show that the Ypt1-Atg11 interaction is required for PAS assembly under normal growth conditions. Moreover, we show that Ypt1 and Atg11 colocalize with Trs85, a Ypt1 activator subunit, and together they regulate selective autophagy. Finally, we show that Ypt1 and Trs85 interact on Atg9-containing membranes, which serve as a source for the membrane component of the PAS. Together our results define a Ypt/Rab module-comprising an activator, GTPase, and effector-that orchestrates the onset of selective autophagy, a process vital for cell homeostasis. Furthermore, because Atg11 does not play a role in ER-to-Golgi transport, we demonstrate here that Ypt/Rabs can regulate two independent membrane-transport processes by recruiting process-specific effectors.

TRAPP is a highly conserved modular multi-subunit protein complex. Originally identified as a “transport protein particle” with a role in endoplasmic reticulum-to-Golgi transport, its multiple subunits and their conservation from yeast to humans were characterized in the late 1990s. TRAPP attracted attention when it was shown to act as a Ypt/Rab GTPase nucleotide exchanger, GEF, in the 2000s. Currently, three TRAPP complexes are known in yeast, I, II, and III, and they regulate two different intracellular trafficking pathways: secretion and autophagy. Core TRAPP contains four small subunits that self assemble to a stable complex, which has a GEF activity on Ypt1. Another small subunit, Trs20/Sedlin, is an adaptor required for the association of core TRAPP with larger subunits to form TRAPP II and TRAPP III. Whereas the molecular structure of the core TRAPP complex is resolved, the architecture of the larger TRAPP complexes, including their existence as dimers and multimers, is less clear. In addition to its Ypt/Rab GEF activity, and thereby an indirect role in vesicle tethering through Ypt/Rabs, a direct role for TRAPP as a vesicle tether has been suggested. This idea is based on TRAPP interactions with vesicle coat components. While much of the basic information about TRAPP complexes comes from yeast, mutations in TRAPP subunits were connected to human disease. In this review we will summarize new information about TRAPP complexes, highlight new insights about their function and discuss current controversies and future perspectives.

Rab GTPases recruit myosin motors to endocytic compartments, which in turn are required for their motility. However, no Ypt/Rab GTPase has been shown to regulate the motility of exocytic compartments. In yeast, the Ypt31/32 functional pair is required for the formation of trans-Golgi vesicles. The myosin V motor Myo2 attaches to these vesicles through its globular-tail domain (GTD) and mediates their polarized delivery to sites of cell growth. Here, we identify Myo2 as an effector of Ypt31/32 and show that the Ypt31/32-Myo2 interaction is required for polarized secretion. Using the yeast-two hybrid system and coprecipitation of recombinant proteins, we show that Ypt31/32 in their guanosine triphosphate (GTP)-bound form interact directly with Myo2-GTD. The physiological relevance of this interaction is shown by colocalization of the proteins, genetic interactions between their genes, and rescue of the lethality caused by a mutation in the Ypt31/32-binding site of Myo2-GTD through fusion with Ypt32. Furthermore, microscopic analyses show a defective Myo2 intracellular localization in ypt31Delta/32ts and in Ypt31/32-interaction-deficient myo2 mutant cells, as well as accumulation of unpolarized secretory vesicles in the latter mutant cells. Together, these results indicate that Ypt31/32 play roles in both the formation of trans-Golgi vesicles and their subsequent Myo2-dependent motility.

The endoplasmic-reticulum quality-control (ERQC) system shuttles misfolded proteins for degradation by the proteasome through the well-defined ER-associated degradation (ERAD) pathway. In contrast, very little is known about the role of autophagy in ERQC. Macro-autophagy, a collection of pathways that deliver proteins through autophagosomes (APs) for degradation in the lysosome (vacuole in yeast), is mediated by autophagy-specific proteins, Atgs, and regulated by Ypt/Rab GTPases. Until recently, the term ER-phagy was used to describe degradation of ER membrane and proteins in the lysosome under stress: either ER stress induced by drugs or whole-cell stress induced by starvation. These two types of stresses induce micro-ER-phagy, which does not use autophagic organelles and machinery, and non-selective autophagy. Here, we characterize the macro-ER-phagy pathway and uncover its role in ERQC. This pathway delivers 20–50% of certain ER-resident membrane proteins to the vacuole and is further induced to >90% by overexpression of a single integral-membrane protein. Even though such overexpression in cells defective in macro-ER-phagy induces the unfolded-protein response (UPR), UPR is not needed for macro-ER-phagy. We show that macro-ER-phagy is dependent on Atgs and Ypt GTPases and its cargo passes through APs. Moreover, for the first time the role of Atg9, the only integral-membrane core Atg, is uncoupled from that of other core Atgs. Finally, three sequential steps of this pathway are delineated: Atg9-dependent exit from the ER en route to autophagy, Ypt1- and core Atgs-mediated pre-autophagsomal-structure organization, and Ypt51-mediated delivery of APs to the vacuole.