Solid Immersion Lens Microscopy to Study Cilia Assembly

ended

Cilia and flagella are evolutionary conserved organelles indispensable for vital processes in eukaryotic organisms, such as environment sensing, cell motility, signaling and development. Broad spectrum of ciliary functions, together with omnipresence of the cilium throughout human body, explains the range of symptoms associated with congenital ciliary disorders called ciliopathies. On the other hand, cilia are essential for survival of parasites, such as trypanosomatids, in the host. Therefore, cilia are of great interest as a potential therapeutic target. The ciliary tip is an essential ciliary domain; it provides capping and mechanical stabilization of the ciliary cytoskeleton, it is a turning point of the intraflagellar transport trains, a sole place of cilium growth and a place of budding of signaling vesicles. Yet the tip is the most enigmatic of all ciliary domains, with structures constituting the ciliary tip largely unknown. This hampers our understanding of how are the tip-related processes brought about and orchestrated. To gain insight into the dynamic ultrastructure od the ciliary tip, I will develop a novel technique for cryogenic correlative light and electron microscopy based on solid immersion lens (SIL) optics. I will integrate the resulting imaging data with the tip proteome project project carried out by the host lab and provide mechanistic understanding of the resulting tip model by employing top-down synthetic biology and in vitro reconstitution approaches. Key achievements of this project will include: (i) development of the cryo-SIL technique and (ii) unraveling the functions of the the ciliary tip domain, which will broaden our knowledge of the principles of self-organization of biological systems.