1998 — 2000 |
Lee, Laura A [⬀] Lee, Laura A [⬀] |
F32Activity Code Description: To provide postdoctoral research training to individuals to broaden their scientific background and extend their potential for research in specified health-related areas. |
G Protein Pathway in Drosophila @ University of California Berkeley
Heterotrimeric guanine nucleotide binding proteins (G proteins) transduce a variety of signals (e.g. hormones, neurotransmitters, and light) across the plasma membrane by coupling transmembrane receptors with intracellular effectors. G proteins are composed of three distinct subunits: alpha (which defines the heterotrimer, beta, and gamma. Alpha subunits can be divided into four classes based on sequence similarities. Members of the G12 class are potential oncogenes because activating mutants have highly potent transforming activities. Signal transduction pathways mediated by members of the G12 class are poorly understood. I propose to study the G12 pathway in Drosophila melanogaster, a powerful genetic system that is ideal for dissection of novel signaling pathways. The Drosophila gene concertina encodes a G protein alpha subunit of the G12 class (CTA) that is required for coordinated cell shape changes during gastrulation. Another Drosophila gene required during gastrulation, fog, encodes a secreted protein (FOG) that potentially activates the CTA pathway. To elucidate the G12 signaling pathway, I plan to examine the effects of ectopic activation of the CTA pathway in Drosophila, identify new components of the pathway by performing a dominant modifier screen and a two-hybrid assay, and demonstrate that FOG is a ligand for a CTA-coupled receptor. Numerous examples of conservation of signaling networks between invertebrates and vertebrates have been described, so identification of new components of the G12 pathway in Drosophila will likely have relevance for our understanding of this important pathway in humans.
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0.937 |
2005 — 2013 |
Lee, Laura Anne [⬀] Lee, Laura Anne [⬀] |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Developmental Regulation of the Cell Cycle in Drosophila
DESCRIPTION (provided by applicant): The high degree of functional conservation of genes involved in the cell cycle combined with the superb genetics and cytology of Drosophila melanogaster make it an ideal model organism for studying cell-cycle regulation in a developmental context. Spermatogenesis utilizes mitotic and meiotic cell cycles coordinated with growth and differentiation programs to generate functional sperm. By mutational analysis, we have identified asunder (asun), which encodes an evolutionarily conserved protein, as an essential regulator of Drosophila spermatogenesis. asun spermatocytes arrest during prophase of meiosis I. Strikingly, arrested spermatocytes contain free centrosomes that fail to stably associate with the nucleus. Spermatocytes that overcome arrest exhibit severe defects in meiotic spindle assembly, chromosome segregation, and cytokinesis. Furthermore, the centriole-derived basal body is detached from the nucleus in asun postmeiotic spermatids, resulting in abnormalities later in spermatogenesis. We find that asun spermatocytes and spermatids exhibit drastic reduction of perinuclear dynein. Dynein is a minus end-directed microtubule motor complex that is required for diverse biological processes, from transport of intracellular cargo to cell migration. Dynein is controlled at multiple levels, including regulation of its subcellular localization; the mechanisms underlying the targeting of dynein to various sites within cells, however, are not well understood. Our current model is that asun coordinates spermatogenesis by promoting dynein recruitment to the nuclear surface, a critical step that is required for nucleus-centrosome coupling at M-phase entry and fidelity of meiotic divisions. ASUN exhibits a dynamic localization pattern during Drosophila male meiosis, and the timing of its release from the nucleus to the cytoplasm correlates with the appearance of dynein on the nuclear surface in G2 spermatocytes. We will assess whether this regulated movement of ASUN within spermatocytes is critical for controlling the activity of dynein. We propose experiments that will allow us to gain a more detailed understanding of the mechanism by which dynein is recruited to the nuclear surface with a focus on elucidating the role of ASUN in this process. Our preliminary data suggest that dynein anchored on the nuclear surface of spermatocytes may preferentially bind to microtubules that are post-translationally modified by acetylation. We will test our hypothesis that this pool of acetylated microtubules mediates key events of Drosophila male meiosis, including nucleus-centrosome coupling. The proposed experiments have the potential to illuminate the mechanism of action of ASUN, to identify additional factors required for recruitment of the dynein motor complex to the nuclear surface, and to define the role of microtubule acetylation during spermatogenesis.
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