The endosome is a dynamic organelle which is responsible for collecting, packaging and sorting protein and lipid cargo to specified sub-cellular sites. These sorting activities are broadly regulated by a family of conserved GTPases belonging to the Rab5 family. In both yeast and higher eukaryotes, Rab5-family GTPases are activated by VPS9 domain-containing guanine nucleotide exchange factors (GEFs). Recently, our lab has demonstrated an important role for the yeast VPS9 GEFs in regulating the endosomal recruitment of the neurodegenerative disease-associated retromer sorting complex. In a mechanism involving local synthesis of PI3P, VPS9-domain GEFs physically bind to retromer and initiate a positive feedback loop of endosomal recruitment. Further characterization of the relationship between VPS9-domain GEFs and retromer may provide insights into regulatory strategies employed by the endosome that can be exploited therapeutically to treat neurodegenerative disease.
Localization of VPS13 to Membrane Contact Sites
Mutations in the four human VPS13 proteins (VPS13A-D) are causative for chorea-acanthocytosis, Cohen syndrome, early-onset Parkinson's disease and spastic ataxia respectively. There is evidence to suggest that the different isoforms localize to different membrane contact sites, regions where organelles are tethered in close proximity without fusing. The divergent localization and distinct diseases associated with each VPS13 protein suggest that these proteins have diverged. By determining which membranes these proteins localize to and the factors required for their recruitment we can begin to understand their function. Using yeast as a model due to its single conserved Vps13 protein, we have determined that Vps13 is recruited to membranes via competitive organelle-specific adaptors. These adaptors bind Vps13 at a region we named the Vps13 Adaptor Binding (VAB) domain through a related motif. Mutations in the VAB domain of VPS13B and VPS13D cause disease. Our research is aimed at further characterizing the adaptor-Vps13 interaction, determining the role of pathogenic VPS13 mutations and identifying VPS13 adaptors in human cells.
Dynamic Protein Depalmitoylation in Membrane Targeting
Palmitoylation – the only reversible lipid modification – is important for tethering signaling proteins to membranes. The dynamic removal of palmitate from NRas prevents its redistribution to intracellular sites and maintains signaling at the plasma membrane. This reversible cycle provides an important mechanism for controlling cell growth that can be exploited to treat cancers driven by oncogenic NRas. Palmitate is added to hundreds of proteins in the cell, but only a few undergo dynamic cycling. This is modulated by the opposing actions of enzymes that add and remove palmitate. We recently discovered that a new family of α/β hydrolase domain 17 (ABHD17) enzymes remove palmitate from NRas. Our studies will determine if these enzymes have other cellular targets, provide insights into their regulation and test if inhibiting the ABHD17 proteins blocks oncogenic NRas signaling. This work will help establish if the ABHD17 proteins are a good target for the development of drugs to treat cancers such as leukemia and melanoma.