2000 — 2003 |
Ghabrial, Amin S |
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. |
Genome Wide Analysis of Epithelial Tube Fusion
The creation of tubular networks is essential to the production of such organs as the lungs, the kidneys, and the vascular system. The Drosophila tracheal system provides an excellent model system in which the formation of tubular networks can be studied using powerful genetic, molecular, and genomic tools. Previous work has shown that there is a high degree of conservation between flies and vertebrates in the molecular pathways used to mediated tube branching. This proposal focuses on another aspect of tracheal network formation, branch fusion. Growing branches must identify and move towards the appropriate fusion partner, then align and form an appropriate connection. These fusion events are essential to the formation of an interconnected network, and remain poorly understood at the cellular and molecular level. Fusion events are mediated by a single cell, called a fusion cell, found at the ends of growing branches. To identify the genes that act during branch fusion, fusion cells will be isolated from embryos by whole animal cell sorting (WACS), and cDNAs made from these cells will be used to probe DNA microarrays. Comparison of genes expressed in fusion cells to genes expressed in other tracheal cells allow identification of fusion cell- specific genes. The expression pattern of these candidate genes will be verified by in situ hybridization, and the loss of function phenotypes of these genes will be determined by RNA interference (RNAi) experiments. Genes will be categorized and ordered into pathways, laying the foundation for a detailed molecular understanding of each cellular step in the fusion process.
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0.954 |
2010 — 2021 |
Ghabrial, Amin S |
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. |
Competition and Morphogenesis in Tip Cell-Mediated Branching of Tubular Networks @ University of Pennsylvania
DESCRIPTION (provided by applicant): A critical mechanism for generating new branches in the human vascular system is sprouting. In sprouting angiogenesis, endothelial cells from a pre-existing tube are led by actively migrating tip cells to form new vessels. Similarly, in the Drosophila respiratory organ (tracheal system), primary branching occurs through a sprouting mechanism in which tip cells play the same essential role. In both models of branch sprouting, the tip cells are morphologically distinct, extending filopodia to sense the local environment and to lead migration up a concentration gradient of a branch-inducing signal (Vascular endothelial growth factor/VEGF and fibroblast growth factor/FGF, respectively). We have shown that tracheal cells compete with each other for tip cell positions in Drosophila, where competition is based on relative levels of FGF Receptor (FGFR) activity and is mediated in part by the lateral inhibitory signal Notch. If Notch signaling is abrogated, a vast excess of tip cells are selected and migrate to the leading end of the sprout while only one or two follower cells comprise the branch stalk. Competition for tip cell positions also occurs during sprouting angiogenesis, with Receptor Tyrosine Kinase (RTK) and Notch signaling playing the same key roles. Despite substantial progress in understanding tip cell selection, the direct targets of Notch signaling that antagonize tip cell behavior in stalk cells remain unknown. Likewise, while RTK signaling promotes tip cell migration, the direct targets of the RTKs in tracheal and endothelial tip cells are not known. We have identified a novel mutation that we have named too many leaders, that confer a tip cell bias to mutant cells, and mutations in the septate/tight junction gene polychaetoid/ZO-1, that confer a Notch-like extra tip cell defect. We propose that tracheal tip cells and endothelial tip cells are selected by a conserved competition-based mechanism, and will systematically dissect the signaling pathways and their downstream targets, with the goal of determining how signaling information is translated into changes in cell shape, cell position within an epithelium, and cell migration, such that a pre- existing tube reorganizes to sprout a side branch. The specific aims of this project are: 1. To define the RTK core components that mediate tip cell selection, and to determine whether a transcriptional response to RTK signaling is required. 2. To determine how Notch signaling is regulated and transduced during branch sprouting, and to examine how selective loss of Notch ligand expression affects tip cell competition. 3. To test the hypothesis that Polychaetoid and Too Many Leaders are downstream effectors of Notch signaling during tip cell competition.
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