Channelrhodopsin-1: Cellular Localization and Role in Eyespot Assembly and Placement in Chlamydomonas reinhardtii

The eyespot of the single-celled alga Chlamydomonas aids the cell in detecting the direction of light in the environment. The complex assembly and asymmetric placement of the eyespot provides a model to ask questions about assembly and asymmetric placement of organelles. Understanding the mechanisms that underlie assembly and asymmetric placement of the eyespot can be applied more broadly to their functions in other eukaryotic organisms. This study sought to understand the role of a key protein in those processes, Channelrhodopsin-1 (ChR1). ChR1 was found to localize along the entire length of the D4 rootlet from the region around the daughter basal body to the eyespot. ChR1 was found to primarily localize to the plasma membrane side of the D4, suggesting that ChR1 was being pulled through the plasma membrane from the region around the basal bodies to the eyespot. Further, ChR1 was found to be able to localize to the eyespot even with the truncation of the large cytoplasmic C-terminal domain, suggesting that ChR1 is able to complex with another protein that is being trafficked to the eyespot. One such protein was thought to be ChR2, the other light-activated ion channel localized to the eyespot. Efforts to isolate a mutation in ChR2 were unsuccessful. Initial efforts were made in this dissertation to perform proteomic studies of ChR1 and identify its interacting partners. ChR1 is not the master regulator of either placement or assembly of the eyespot, but work in this study lays the groundwork to further investigate transport of ChR1 and interacting proteins to the eyespot and their role in assembly of the eyespot