| The mechanisms underlying actin/microtubule (MT) coordination in axon targeting remain largely unknown. Here, I characterize Drosophila pod-1 (dpod1), a large protein with actin and MT binding domains. Purified Dpod1 crosslinks both actin and MTs. In cultured cells, endogenous lamellar Dpod1 is primarily actin-associated but often colocalizes with MTs. When actin is disrupted, Dpod1 delocalizes from the cell's edge and relocalizes to MTs. Overexpression of Dpod1 causes dose-dependent cytoskeletal remodeling: cells extend dynamic, neurite-like processes that are actin rich, actin-dependent, and MT-independent. They are invaded by MTs and often contain Enabled at their tips. In embryos, Dpod1 is strongly expressed in the developing nervous system, localizes to the tips of extending neurites, and often concentrates at axonal choice points. Dpod1 is required for proper axon guidance but not for cell polarity, cell fate determination, or continued neurite extension. In addition, Dpod1 is required in the PNS for proper cell migration. dpod1 genetically interacts with robo and enabled. Furthermore, postmitotic neuronal overexpression of Dpod1 disrupts axon targeting. Together, these data suggest that Dpod1 is an important cytoskeletal regulator whose levels must be maintained for accurate axon guidance and neuronal migration.; Interestingly, dpod1 is highly conserved: vertebrates possess a single pod-1 expressed in the developing nervous system.; In the developing Drosophila retina and optic lobes, Dpod1 is widely expressed at several developmental stages and is required for photoreceptor axon guidance. Also, in developing mechanosensory bristles, loss of Dpod1 causes defects in cytoskeletal organization, and overexpression dramatically changes cell shape in a dose dependent manner. Additionally, Dpod1 localizes to the cleavage furrow of dividing cells, but its function there remains mysterious.; Finally, I show that phosphatidylinositol-3-kinase (PI3K), a lipid kinase that produces phosphatidylinositol-3,4,5-trisphosphate (PIP3), may play a role in neuroblast asymmetric cell division. PIP3 forms a cell-cycle dependent apical crescent in neuroblasts. Pharmacological disruption of PI3K causes asymmetrically dividing neuroblasts to exhibit defects in basal (but not apical) polarity markers and mitotic spindle orientation. These preliminary results suggest that active PI3K is an important molecule in neuroblast polarity. |
