EXTRINSIC CUES, INTRINSIC CUES AND MICROFILAMENTS REGULATE ASYMMETRICPROTEIN LOCALIZATION IN DROSOPHILA NEUROBLASTS

Background: The Drosophila central nervous system develops from stem c ell like precursors called neuroblasts, which divide unequally to bud off a series of smaller daughter cells called ganglion mother cells. N euroblasts show cell-cycle-specific asymmetric localization of both RN A and proteins: at late interphase, prospero RNA and Inscuteable, Pros pero and Staufen proteins are all apically localized; at mitosis, Insc uteable remains apical whereas prospero RNA, Prospero protein and Stau fen protein form basal cortical crescents. Here we use in vitro cultur e of neuroblasts to investigate the role of intrinsic and extrinsic cu es and the cytoskeleton in asymmetric localization of Inscuteable, Pro spero and Staufen proteins. Results: Neuroblast cytokinesis is normal in vitro, producing a larger neuroblast and a smaller ganglion mother cell. Apical localization of Inscuteable, Prospero and Staufen in inte rphase neuroblasts is reduced or eliminated in vitro, but all three pr oteins are localized normally during mitosis (apical Inscuteable, basa l Prospero and Staufen). Microfilament inhibitors result in delocaliza tion of all three proteins. Inscuteable becomes uniform at the cortex, whereas Prospero and Staufen become cytoplasmic; inhibitor washout le ads to recovery of microfilaments and asymmetric localization of all t hree proteins. Microtubule disruption has no effect on protein localiz ation, but disruption of both microtubules and microfilaments results in cytoplasmic localization of Inscuteable. Conclusions: Both extrinsi c and intrinsic cues regulate protein localization in neuroblasts. Mic rofilaments, but not microtubules, are essential for asymmetric protei n anchoring (and possibly localization) in mitotic neuroblasts. Our re sults highlight the similarity between Drosophila, Caenorhabditis eleg ans, vertebrates, plants and yeast: in all organisms, asymmetric prote in or RNA localization and/or anchoring requires microfilaments.