Tracking of C. Reinhardtii flagellar disassembly following femtosecond laser ablation

Abstract

Single-cell green algae (C. Reinhardtii) is a key model organism to study ciliogenesis. Cilia have important roles in sensory signaling pathways and in clearing the airways of mucus and dirt in multiple systems of the human body. As cilia are found on most eukaryotic cells, defects in ciliogenesis result in many symptoms and disorders. We are testing the hypothesis that when a flagellum is removed, the long flagellum shrinks because it is competing with the shorter regrowing flagellum. We used a 780-nm 200-fs laser to perform laser ablation to amputate one of two flagella on wildtype and mutant algae. Fla3 and Fla10 mutants were altered to inhibit the KAP kinesin motor that drives the intraflagellar transport (IFT) pathway. Impaired IFT pathways would demonstrate a lag in response to flagellar length equalization and a reduced disassembly rate. Quantified images following the long flagellum for 20 min post-ablation demonstrate a delayed disassembly rate in the Fla3 mutant compared to wildtype; Fla10 was inconclusive. Therefore, it was concluded that the proper function of KAP motor protein serves a significant role in length control of cilia. In the future, we will compare the assembly rates of flagellar regrowth for the wildtype and mutants.