A Novel Regulatory Role of Tomosyn in HOPS-dependent Endo-lysosomal Function in C. elegans Coelomocytes

Intracellular membrane fusion events between endosomes, lysosomes, and autophagosomes are critical to maintaining cellular health. Dysfunction of these steps can lead to severe functional defects such as cognitive impairments and premature death. We previously identified VPS-39 as a binding partner of the WD40-repeat domains of C. elegans Tomosyn in a yeast two-hybrid assay. Tomosyn is a known inhibitor of exocytic membrane fusion and prevents excessive neurotransmitter release. VPS-39 is a member of the HOPS complex which is essential for mediating endomembrane fusion and embryonic survival. The interaction of these proteins revealed a potential novel role for Tomosyn in regulating endomembrane fusion. In this thesis, we characterize the novel relationship of VPS-39 and Tomosyn in regulating endomembrane fusion events in C. elegans coelomocytes. We hypothesized that Tomosyn and VPS-39 function together at the junction of late endosomes and lysosomes to regulate the fusion of these organelles. Our investigation of this hypothesis using fluorescence markers and ultrastructural data provides evidence to support our proposed mechanism. Additionally, this body of work provides the first reported analysis of entire coelomocytes using ultrathin serial sections, the first evidence that Tomosyn functions in regulating endomembrane fusion events, and that VPS-39 displays in vivo RAB-7 GEF activity.