Mechanisms of extracellular vesicle biogenesis and uptake - Project Summary Small secreted extracellular vesicles (EVs) are released by most, if not all cells, to mediate intercellular transport of bioactive macromolecules. The protein, RNA, and metabolite cargoes carried by EVs regulate physiological processes and propagation of pathological conditions including neurodegenerative, cardiovascular, and kidney diseases. Consequently, it is important to understand fundamental mechanisms that impact EV cargo enrichment, biogenesis, targeting, uptake, and function. How does a cell release unique EV subpopulations to foster heterogeneous functionality? EV biogenesis occurs in response to signaling cues in native cellular environments, thus it is important to utilize an in vivo model to answer this question. One site of EV shedding is the primary cilium, a specialized organelle protruding from non-dividing cells that serves as a platform for organizing both signal reception and transmission. Shedding of bioactive EVs termed ectosomes from cilia of male tail sensory neurons can be observed in live C. elegans. Multiple different EV subpopulations are shed from these sensory cilia. The ectosomes that bud from the cilium distal tip and are released through a pore in the worm cuticle can be deposited on the vulva during mating and impact animal-animal communication. EVs shed from the ciliary base can be taken up by surrounding glia to influence cell-cell signaling in addition to being released into the environment. We have developed unique C. elegans strains expressing fluorescently-tagged EV cargoes and cutting-edge imaging methods, which enable us to quantitate: 1) EV shedding from specific, identified neuronal cilia, 2) EV cargo abundance in the releasing cilia, 3) glial uptake of EVs, and 4) cellular response to EV shedding and uptake. Our overarching goal is to define how specific conserved genes and cellular stress impact EV release, uptake, and response by using the strengths of the C. elegans model system combined with in vivo imaging, quantitative analyses, behavioral assays, and proteomic approaches. Our first thematic area of research focuses on answering fundamental questions in the EV biology field, using the power of genetics to characterize new genes that we discovered are important for biogenesis of individual EV subpopulations, EV shedding in response to physiological stimulus, and uptake of EVs into surrounding glia. In our second thematic area we will concentrate on how oxidative stress, which contributes to the pathogenesis of multiple diseases, impacts EV shedding and uptake as well as the potential function of redox-related cargoes in EVs. This project will have significant impact as defining factors that regulate EV production and functionality is essential to understand how transfer of cargoes can impact disease states and develop EV-based diagnostic tools and therapies.
