In sprouting angiogenesis, endothelial cells from a pre-existing tube are led by actively migrating tip cells to form new vessels. In the Drosophila respiratory organ (tracheal system), primary branching is also led by tip cells. Work from us and others showed that tip cells in both systems are selected through a competition-based mechanism. 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. Subsequent to initiating and guiding the outgrowth of a new branch, tip cells execute a second essential function: they must form a tube (lumenize) in order to make the new branch functional. Tip cells hollow out to form seamless tubes that lack junctional seams (adherens junctions and tight/septate junctions). In the Drosophila tracheal system, all tip cells reside in stereotyped positions and form seamless tubes; furthermore, tracheal terminal cells form new seamless tube side branches throughout the course of larval life. These qualities, combined with the power of genetic analysis in Drosophila, have made tracheal terminal cells an exceptionally useful model for dissecting seamless tube morphogenesis. Tip cell tube formation is critical, as it permits transport of blood or gas throug the cell; however, despite or recent progress (4 major publications in the last grant cycle), important questions remain: how does FGFR- signaling trigger distinct outputs at each stage? how does FGFR induce seamless tubes? how does FGFR regulate seamless tube branching? what are the effectors regulated downstream of FGFR? We propose to exploit or published and preliminary data to address these questions using innovative approaches. With the twin goals of understanding how signaling is coupled to changes in the mechanism of tubulogenesis, and how the seamless tubulogenesis machinery operates downstream of FGFR, we propose the following specific aims:
Aim 1 : Determine how FGFR activation induces seamless tubulogenesis. We will focus our studies of seamless tubulogenesis on the novel zinc finger transcription factor that we identify, in our preliminary data, as essential for seamless tube morphogenesis.
Aim 2 : Determine how FGFR activation regulates local branching of the seamless tube. We will focus on characterization of seamless tube branching using innovative optogenetic and live imaging tools. We will follow up on our preliminary data to establish the roles of three proteins - PALS2/MPP6/2 (Drosophila Varicose), Spectroplakin (Drosophila Short stop) and Moesin - in seamless tube branching in Drosophila and in the zebrafish vascular system. Completion of these specific aims will represent a dramatic advance in the field, connecting signal transduction to the cellular machinery of tube morphogenesis at an unprecedented level of resolution.

Public Health Relevance

In tubular networks, such as the human vascular system, fine connections between branches are often made by so-called 'seamless' endothelial cells, which make specialized, subcellular tubes that lack junctional seams. Formation of patent connections is required for blood flow, and misregulation of that process is implicated in human disease (cerebral cavernous malformations). We propose to use a Drosophila model of seamless tube formation (the terminal cells of the tracheal system) to better understand the genetic and molecular mechanisms at play.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM089782-07
Application #
9115620
Study Section
Cardiovascular Differentiation and Development Study Section (CDD)
Program Officer
Hoodbhoy, Tanya
Project Start
2009-12-01
Project End
2019-06-30
Budget Start
2016-07-01
Budget End
2017-06-30
Support Year
7
Fiscal Year
2016
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Francis, Deanne; Ghabrial, Amin S (2015) Compensatory branching morphogenesis of stalk cells in the Drosophila trachea. Development 142:2048-57
Schottenfeld-Roames, Jodi; Rosa, Jeffrey B; Ghabrial, Amin S (2014) Seamless tube shape is constrained by endocytosis-dependent regulation of active Moesin. Curr Biol 24:1756-64
Schottenfeld-Roames, Jodi; Ghabrial, Amin S (2013) Osmotic regulation of seamless tube growth. Nat Cell Biol 15:137-9
Song, Yanjun; Eng, Melissa; Ghabrial, Amin S (2013) Focal defects in single-celled tubes mutant for Cerebral cavernous malformation 3, GCKIII, or NSF2. Dev Cell 25:507-19
Schottenfeld-Roames, Jodi; Ghabrial, Amin S (2012) Whacked and Rab35 polarize dynein-motor-complex-dependent seamless tube growth. Nat Cell Biol 14:386-93
Ghabrial, Amin S (2012) A sweet spot in the FGFR signal transduction pathway. Sci Signal 5:pe1
Ghabrial, Amin S; Levi, Boaz P; Krasnow, Mark A (2011) A systematic screen for tube morphogenesis and branching genes in the Drosophila tracheal system. PLoS Genet 7:e1002087
Schottenfeld, Jodi; Song, Yanjun; Ghabrial, Amin S (2010) Tube continued: morphogenesis of the Drosophila tracheal system. Curr Opin Cell Biol 22:633-9