Structural Biology & Biochemistry (SBB) Theme Seminar - Assembly of Retromer–Rab7–Gyp6 Complex: A Novel Molecular Timer for Vacuolar Cargo Retrieval

Title: Assembly of Retromer–Rab7–Gyp6 Complex: A Novel Molecular Timer for Vacuolar Cargo Retrieval

Speaker: Dr Kevin Chen (IMB)

Abstract: The evolutionary conserved Retromer complex, composed of Vps35, Vps26 and Vps29, is a vital regulator of endosomal membrane trafficking, and mutations in its subunits and interactors lead to neurodegenerative disorders. Originally discovered in yeast, Retromer was identified as part of a larger complex with the membrane-associated SNX-BAR proteins Vps5 and Vps17, forming a heteropentameric assembly. Prior to this complex assembly, Retromer is known to be recruited to endosomal membranes in part by the active GTPase Rab7. Although the interaction of Retromer and Rab7 is implicated in membrane trafficking, its molecular details and functional mechanisms remain poorly understood. Here, we elucidate the structural basis of the yeast Retromer - Rab7 complex on membranes using cryo-electron tomography (cryoET). We found that Retromer and Rab7 co-assemble into a well-ordered tubular coat carrier in the absence of canonical membrane curvature drivers or sorting nexin proteins such as Vps5 and SNX3. In parallel, we applied proximity labelling and AlphaFold-based screening to identify Gyp6 as the relevant GTPase-activating protein (GAP) responsible for Rab7 inactivation in this context. Using X-ray crystallography, we found that Gyp6 binds to the Vps29 subunit of Retromer via a conserved PL motif. Disruption of the Retromer - Gyp6 interaction leads to the co-accumulation of Retromer and its cargo Vps10 in tubulovesicular structures. Together with our CryoET structure, these findings support a model in which Gyp6 mediated inactivation of Rab7 triggers a handover mechanism, facilitating the transition of Retromer from a Rab-GAP associated state to its pentameric complex assembly. 

Bio: I obtained my PhD at the Institute for Molecular Bioscience (IMB), University of Queensland (UQ), under the supervision of Professor Jenny Martin. During my time in her lab, I developed my expertise as a crystallographer. In 2016, I joined Professor Brett Collins’s lab, where I began investigating protein complexes involved in endosomal trafficking, particularly the Retromer complex. My current research employs a range of biochemical and structural techniques, including ITC, cryo-electron microscopy and X-ray crystallography, to understand how Retromer interacts with its cargo adaptors and regulatory proteins in a membrane environment.