This dissertation focuses on microtubule assembly mechanisms and functions in dividing and non-dividing cells. In dividing cells, centrosome is the major microtubule-organizing center (MTOC). Ninein (Nin) is a centrosomal protein whose gene is mutated in Seckel syndrome, an inherited recessive disease that results in primordial dwarfism, cognitive deficiencies, and increased sensitivity to genotoxic stress. Nin is evolutionarily conserved, yet its role in cell division and development has not been investigated in a model organism. Here, we characterize the single Nin ortholog in Drosophila. Drosophila Nin localizes to the periphery of the centrosome, but not at centriolar structures as in mammals. However, Nin shares the property of its mammalian ortholog of promoting microtubule assembly. In neural and germline stem cells, Nin localizes asymmetrically to the younger (daughter) centrosome, yet it is not required for the asymmetric division of dividing stem cells. Surprisingly, loss of nin expression from a nin mutant does not significantly impact embryonic and brain development, fertility, or locomotor performance of mutant flies, nor their survival upon exposure to DNA damaging agents. While not essential, Nin localizes to non-centrosomal MTOCs (ncMTOCs) in wing epithelia and muscle, two types of specialized and differentiated cell types, suggesting that Nin plays a supportive role in non-centrosomal microtubule organization. ncMTOCs have a variety of roles presumed to serve the diverse functions of the range of non-dividing cell types in which they are found. ncMTOCs are diverse in their composition, subcellular localization, and function. Here we report a novel perinuclear MTOC in another differentiated Drosophila cell type, fat body cells. This perinuclear ncMTOC in fat body cells is anchored by Msp300/Nesprin at the cytoplasmic surface of the nucleus. Msp300 recruits the MT minus-end protein Patronin/CAMSAP, which functions redundantly with Nin to assemble non-centrosomal MTs and does so independently of the widespread MT nucleation factor -tubulin. Patronin does not antagonize with known MT depolymerases in fat body ncMTOC, but acts cooperatively with Ninein to assemble circumferential MTs, and also recruit MT polymerase Msps to promote elongation of radial MTs. Functionally, the fat body ncMTOC is essential for retrograde dynein-dependent endosomal trafficking to restrict plasma membrane growth and for the secretion of basement membrane proteins. Together, we identify an ncMTOC with novel architecture and MT regulation properties that serves a vital secretory function.
