Sunday, May 11, 2025
5/11/2025
Coordination of intracellular trafficking pathways by Ypt/Rab GTPases and their GEFs. - Abstract In the three major intracellular trafficking pathways–exocytosis, endocytosis and autophagy–proteins and membranes are secreted, internalized or shuttled for degradation, respectively. These pathways are regulated by the conserved Ypt/Rab GTPases that when activated by nucleotide exchangers (GEFs), recruit their effectors to membranes. These effectors are machinery components that mediate vesicular transport steps, from vesicle formation, through motility and targeting, to fusion. I have worked in the Ypt/Rab field since its inception and contributed to the formulation of principles that underlie their mode of action. These include ideas that they function in “GTPase modules”, which contain GEF/s, a GTPase, and effector/s, to organize pathway- or step-specific membrane microdomains. While mechanisms underlying Ypt/Rab function are currently known, questions regarding pathway and step coordination remain open. We propose that Ypt/Rab GTPases coordinate intracellular trafficking at three levels: Coordination of multiple pathways, integration of transport steps into whole pathways, and coordination of vesicular transport sub-steps of individual transport steps. The proposed research relies on our recent findings using yeast as a model system, and we will continue using yeast due to its smaller proteome that results in a much smaller interactome, which is important for exploring the following coordination issues: Multiple pathways coordination: I propose that Ypt/Rabs coordinate autophagy with secretion and endocytosis at two intersections. In the first, Ypt1/Rab1 is required for the beginning of secretion and autophagy in the context of two different GTPase modules. In the second intersection, merging of endocytosis and late autophagy is regulated by a shared Vps21/Rab5 GTPase module. Here, we will determine whether cells prioritize certain pathways under different environmental conditions and how such a priority is promoted. Integration of transport steps: Here, we will address two major questions: First, how do Ypts regulate the beginning of a pathway, especially when a single GTPase functions in the context of two different modules? Second, what are the specific mechanisms by which Ypts coordinate early and late steps in secretion and autophagy? Coordination of vesicular transport sub-steps: We will explore late steps of the secretory and autophagy pathways, for which members of the GTPase modules are known, and ask how effectors that function sequentially are recruited. We will use classical and molecular genetics combined with cell biology and biochemistry approaches to address these questions. An efficient and well-coordinated network of cellular trafficking pathways is important for all the systems of the human body, and even a minor defect can result in a severe disease. Ypt/Rabs in general were implicated in a spectrum of acquired and inherited diseases, and those we study were associated with cancer and neurodegeneration. Finally, we recently showed the value of yeast modeling in understanding how a conserved protein variant causes a neurodevelopmental disorder.
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