The major objectives of this proposal are to study the genetic, biosynthetic, and macromolecular assembly processes involved in the morphogenesis of the eukaryotic flagellum. The proposed studies will be carried out in the biflagel-late alga, Chlamydomonas reinhardtii, because of its proven advantages as a model system for molecular analysis of flagellar structure and assembly. Given the structural and biochemical complexity of the flagellum, molecular dissection of its biogenesis requires that highly specific and sensitive probes of the system area available. This research project is directed toward generating and utilizing such immunological and genetical probes. A library of hybridomas secreting monoclonal antibodies have been generated already in our laboratory, and additional ones will be isolated. These monoclonal antibodies will be specifically used to follow different flagellar proteins from their time of synthesis to assembly into structure. These studies will utilize the rapid and synchronous regeneration of flagella which occurs in Chlamydomonas following experimental deflagellation. Concurrent with these studies, we will pursue a molecular and genetic characterization of a series of non-conditional and thermosensitive mutants of Chlamydomonas with specific defects for flagellar biogenesis. Through our analysis of these mutants, already at hand, we anticipate that structural and regulatory genes which play critical functions in flagellar morphogenesis will be identified.
