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Minna Roh PhD Student Cell-Cell
Communication: Does Wnt act as a positional cue?  Establishing cell polarity is essential to generate
cellular diversity.
Cell polarity requires intricate cell-cell signalling and remodeling of
the underlying cytoskeleton - a phenomenon that remains under-explored
in developmental systems. To explore this intricate process, we are studying the four-cell stage C. elegans embryo. At this stage, signals from one cell result in polarization of it’s neighbouring responding cell to generate daughter cells with distinct developmental fates. The signalling cell produces two signals, MES-1 (a transmembrane protein) and a Wnt homolog, MOM-2, that polarize the responding cell, aligning the mitotic spindle as well as regulating gene expression. It is known that both pathways are required for polarized division, and cell manipulation experiments suggest that it is the position of the Wnt signal alone that determines spindle orientation (Goldstein et al., 2006). However, it is unknown whether this is a direct effect of Wnt, or an indirect downstream effect. To address this issue, we are using a novel approach in which beads coated with purified Wnt proteins are used to manipulate the position of the Wnt source on an isolated responding cell. We have also constructed a strain in which GFP-tagged tubulin is expressed in a Wnt background to examine the effects of a local Wnt signal on spindle dynamics in real-time. Our studies suggest that Wnt may act as an instructive cue at high levels, but as a permissive cue at low levels, and that there is another, perhaps weaker, instructive cue in Wnt mutant signalling cells. We are also genetically altering the levels of Wnt to further test whether this is true in vivo. Combining these cell manipulation experiments with imaging, we hope to extend this approach to investigate the effects of Wnt signalling on the localization of downstream cell polarity proteins by high resolution microscopy in C. elegans. Investigating the role of the Arp2/3 Complex during C. elegans gastrulation Gastrulation is a crucial
process that shapes the body plan of
organisms. Regulated cell movements are critical for gastrulation and
require intricate coordination of cytoskeletal components. The
potential to combine genetics, live imaging, and cell manipulations
makes C. elegans gastrulation an attractive model for studying
the cellular mechanisms of morphogenesis.
C. elegans gastrulation is initiated by the ingression of two endodermal precursor cells, Ea and Ep. The current model for this ingression event is through myosin activation and subsequent constriction of the apical region of the E cells, thereby pulling in the neighbouring cells such that they fill in the gap left by the ingressing E cells (Lee and Goldstein, 2003; Lee et al., 2006). Surprisingly, using live imaging techniques to visualize actin, we found that specific neighbouring cells extend short actin-rich structures near their apical borders with the E cells. These dynamic extensions occur where cell flattening has previously been reported by SEM (Nance and Priess, 2002). Cellular actin architecture is remodeled based on signalling events and upstream actin regulators in diverse cell types. One such regulator is the Arp2/3 complex. This complex acts to nucleate new actin filaments off existing actin filaments and, thus, affects overall actin organization. Severson et al., in 2002, discovered that depleting C. elegans Arp-related proteins, ARX-2 and ARX-1, results in gastrulation defects and the E cells divide on the surface of the embryo. We have found that in Arp2/3 depleted embryos, the E cells are polarized normally and myosin is activated in the apical region of these cells, but the F-actin-rich structures in neighbouring cells fail to form. Thus, these results suggest a new layer to our existing model of C. elegans gastrulation, in that in addition to apical constriction, E cell ingression may also involve Arp2/3-dependent, F-actin-rich extensions from specific neighbouring cells.
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