4 research outputs found
Cellular Mechanisms Regulating Single Lumen Formation in the Zebrafish Gut
<p>The formation of a single lumen during tubulogenesis is crucial for the development and function of many organs. Although 3D cell culture models have identified molecular mechanisms controlling lumen formation in vitro, their function during vertebrate organogenesis is poorly understood. In this work we used the zebrafish gut as a model to investigate single lumen formation during tubulogenesis. Previous work has shown that multiple small lumens enlarge through fluid accumulation and coalesce into a single lumen. However, since lumen formation occurs in the absence of apoptosis, other cellular processes are necessary to facilitate single lumen formation. </p><p>Using light sheet microscopy and genetic approaches we identified a distinct intermediate stage in lumen formation, characterized by two adjacent un-fused lumens. These lumens are separated by cell contacts that contain basolateral adhesion proteins. We observed that lumens arise independently from each other along the length of the gut and do not share a continuous apical surface. Resolution of this intermediate phenotype into a single, continuous lumen requires the remodeling of basolateral contacts between adjacent lumens and subsequent lumen fusion. </p><p>Furthermore, we provide insight into the genetic mechanisms regulating lumen formation through the analysis of the Hedgehog pathway. We show that lumen resolution, but not lumen opening, is impaired in <italic>smoothened (smo)</italic> mutants, indicating that fluid-driven lumen enlargement and resolution are two distinct processes. We also show that <italic>smo</italic> mutants exhibit perturbations in the Rab11 trafficking pathway, which led us to demonstrate that Rab11-mediated recycling, but not degradation, is necessary for single lumen formation. Taken together, this work demonstrates that lumen resolution is a distinct genetically-controlled process, requiring cellular rearrangement and lumen fusion events, to create a single, continuous lumen in the zebrafish gut.</p>Dissertatio
Stabilization of β-catenin in XY gonads causes male-to-female sex-reversal
During mammalian sex determination, expression of the Y-linked gene Sry shifts the bipotential gonad toward a testicular fate by upregulating a feed-forward loop between FGF9 and SOX9 to establish SOX9 expression in somatic cells. We previously proposed that these signals are mutually antagonistic with counteracting signals in XX gonads and that a shift in the balance of these factors leads to either male or female development. Evidence in mice and humans suggests that the male pathway is opposed by the expression of two signals, WNT4 and R-SPONDIN-1 (RSPO1), that promote the ovarian fate and block testis development. Both of these ligands can activate the canonical Wnt signaling pathway. Duplication of the distal portion of chromosome 1p, which includes both WNT4 and RSPO1, overrides the male program and causes male-to-female sex reversal in XY patients. To determine whether activation of β-catenin is sufficient to block the testis pathway, we have ectopically expressed a stabilized form of β-catenin in the somatic cells of XY gonads. Our results show that activation of β-catenin in otherwise normal XY mice effectively disrupts the male program and results in male-to-female sex-reversal. The identification of β-catenin as a key pro-ovarian and anti-testis signaling molecule will further our understanding of the mechanisms controlling sex determination and the molecular mechanisms that lead to sex-reversal