Branching Out: Genetic analysis of branch outgrowth in terminal cells. Tiffani A. Jones, Mark M. Metzstein. Human Gen, Univ Utah, Salt Lake City, UT.

   Cellular morphology is critical for cell function. However, little is known about how individual cells generate their specific shapes. Larval terminal cells, a component of the respiratory system, are an excellent model for investigating questions of cell shape due to their elaborate branched morphology. Terminal cells initiate branching from a central branch, containing the cell body and nucleus. Subsequent side branches bifurcate from this central branch, with a general reduction in the diameter of successive branches. In previous work, we showed that PAR-polarity proteins (Par-6/Baz/aPKC/Cdc42) are required for terminal cell branching, but not outgrowth, and are downstream of the branchless/breathless FGF signaling pathway required for terminal cell outgrowth and branching. However, how branch outgrowth occurs mechanistically and how the PAR complex may regulate this process is unknown. Our recent work has turned to testing vesicle trafficking pathways to elucidating the molecular mechanisms required for terminal cell outgrowth. In particular, we are focusing on a conserved complex, the exocyst. The exocyst complex is composed of the proteins Sec3, Sec5, Sec6, Sec8, Sec10, Sec15, Exo70, and Exo84 and is best known for its roles in facilitating polarized addition of post-Golgi derived vesicles to the plasma membrane. We find all 8 members of the exocyst complex, as well as the small GTPases Rab11 and Rab8, which are known to function in exocyst complex assembly, are required for terminal cell branching and outgrowth. The PAR protein Cdc42 has been shown to be required for exocyst protein localization in yeast and mammalian cell culture. We have found Cdc42 is required for exocyst protein localization in terminal cells and that PAR proteins and the exocyst act in a single genetic pathway to control terminal cell development. We suggest a model in which terminal cell development occurs through a process of branch specification via PAR complex activity, which directs exocyst complex mediated polarized exocytosis to facilitate terminal branch outgrowth.