Dissecting the functional interactions between primary cilia, extracellular vesicles, and endo-lysosomal pathways

Recent studies have revealed that primary cilia release extracellular vesicles (EVs) as a means to regulate ciliary membrane homeostasis and signalling function, but the underlying mechanisms and disease relevance are still being investigated. DC4 will use cell-based and automatic imaging approaches to investigate, on the one hand, how mutations in specific ciliopathy disease genes (e.g. BBS1 and NPHP1) affect the subcellular/ciliary distribution and dynamics of relevant EV biogenesis/endo-lysosomal regulators recently identified by the lab and, conversely, how depletion of these regulators affects ciliary membrane protein composition as well as EV release frequency and composition. Focus will be on kidney epithelial cells and both small and large EV populations will be studied. Apart from advanced imaging approaches, the project involves generation of relevant mutant and fluorescent reporter cell lines by CRISPR/Cas9-mediated gene knockout and lentiviral transduction; biochemical isolation of EVs; nanoparticle tracking and mass spectrometry analysis of EVs. Cell-based signalling assays may also be carried out. The results of this project will provide new insight into the mechanisms underlying EV release by primary cilia, and how perturbation of these mechanisms may contribute to disease pathology of ciliopathies.

Fellow profile: Master degree in Cell Biology, Molecular Biology, Developmental Biology, or related field. This project is very suitable for students with a knowledge of mammalian cell culture, molecular biology and biochemistry techniques, fluorescence microscopy.