Beschreibung:
Abstract: Primary cilia are fundamentally important organelles involved in organ development, signaling pathways and cancer. The septin family of small GTP-binding proteins have been linked to cilia and ciliopathies but their function in ciliogenesis remains elusive. A detailed understanding of the process of cilia formation is critical for therapeutic approaches in the future.<br>This thesis demonstrated that cilia formation depends on the presence and dynamics of septins. Knockdown of septins and the inhibition of septin dynamics via forchlorfenuron (FCF) resulted in significantly decreased ciliogenesis. In addition, septins were visualized at the base of primary cilia or along the full length of the axoneme. <br>In the first part, it was shown that septins are regulated by the Rho GTPases Cdc42 and TC10 via their downstream effectors of the Borg family. Here it was shown that septin filaments associate with Borg proteins in the cytoskeleton as well as within cilia. Dominant active and negative Cdc42 inhibit cilia formation indicating that the free cycling of Cdc42 between its active and inactive state is essential for Borg and septin regulation during ciliogenesis. Even though Cdc42 and TC10 are close homologues, their functions during ciliogenesis are not redundant. The expression of dominant active TC10 inhibited cilia formation, whereas dominant negative TC10 had no influence on ciliogenesis. Knockdown experiments demonstrated that Cdc42 is essential for ciliogenesis but TC10 is not. Also the double knockdown of Cdc42 and TC10 showed no additive effect on ciliogenesis. However, an influence of TC10 on ciliary length was detected. Moreover, two distinct cellular distributions at cilia were observed: active Cdc42 was predominantly located as a spot at the base whereas a TC10 accumulation surrounded the base of cilia, suggesting different functions during ciliogenesis.<br>The second part demonstrated that septins are involved in exocytotic processes during ciliogenesis. Septins colocalize with the exocyst complex at the base of cilia. Rab10 vesicles were shown to accumulate at the base of cilia and to colocalize with septins. GST pull-down experiments revealed that septins interact with Rab10, independent of the nucleotide-binding state. Knockdown experiments revealed that Rab10 is important for cilia formation. Moreover, the inhibition of septin dynamics via FCF hindered the traffic of a cilia-targeted protein to the cilium. <br>This thesis investigated how septins are regulated via Borg proteins and their upstream Rho GTPases. Moreover, it presents a novel prospective for the role of septins in the process of ciliogenesis, away from the propagated diffusion barrier theory