To build a functional vascular system, a network of endothelial tubes must be generated through a combination of vasculogenesis and angiogenesis. During sprouting angiogenesis, endothelial tip cells lead the outgrowth of new branches. Tip cells anastomose with other tip cells or with pre-existing tubes. At the same time, tip cells must also lumenize so that the tubes become patent. Studies in zebrafish indicate tip cells lumenize by a process that an intracellular tube that, in cross section, lacks junctional seams (seamless tube). Even tip cells that later remodel to contribute to multicellular tubes, pass through a seamless tube intermediate stage. Likewise, during primary branching of the Drosophila respiratory system, tracheal tip cells lead the outgrowth of new branches. To form a network, some tracheal tip cells anastomose (fusion cells) while others (terminal cells) branch extensively to produce dozens of blind ended tubes that ramify on target tissues and act as the sites of gas exchange. Like endothelial tip cells, tracheal tip cells form seamless tubes. Cell biological studies have led to the current model of seamless tube formation by inverse membrane blebbing; however, very little is known about the genetic and molecular pathways that are required. We have been exploiting the powerful forward genetic approaches possible in Drosophila to meet our long-term objective of pioneering an understanding of the genetic and molecular framework required to make, shape and maintain seamless tubes. The rationale of our approach is that the fundamental rules and genetic pathways operative in Drosophila are likely to be conserved throughout the animal kingdom. As we build our understanding by identifying novel genes required for seamless tube morphogenesis (AIM 1: Determine the molecular identity and function of the cystic lumens gene.), we also go deeper ? using molecular genetic, cellular, and proteomic approaches ? to identify additional components and mechanisms of action for genetic pathways we have previously identified, and extending our findings to endothelial seamless tubes (AIM 2: Elucidate the cellular and molecular mechanisms of TBC1D10 and Rab35 action in seamless tube growth). Additionally, one seamless tubulogenesis pathway (the Cerebral Cavernous Malformations 3-Germinal Center Kinase III pathway) we identified, is known to be critical in endothelial cells, as mutations in orthologous human genes lead to familial vascular disease. Using Drosophila genetic and proteomic approaches, we have identified factors required both upstream and downstream in the CCM3-GCKIII pathway, and now propose to more clearly define the biological consequences of perturbing the pathway and to identify the downstream targets of the signaling pathway. Given the conserved role of CCM3 in tubulogenesis, we also seek to extend our results to the vertebrate endothelial system, making use of zebrafish, whose unique properties will allow both genetic manipulation and live imaging in vivo of vascular lesion formation (AIM 3: To characterize the role of the NDR kinase, Tricornered, in regulating seamless tube shape.).

Public Health Relevance

This project is relevant to human health on two levels. First, we pioneer a basic science understanding of the formation and maintenance of tubular networks that is relevant for all organs. Second, we focus on a poorly understood pathway that is directly affected in human vascular disease (cavernous angiomas).

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM089782-10
Application #
9818887
Study Section
Cardiovascular Differentiation and Development Study Section (CDD)
Program Officer
Hoodbhoy, Tanya
Project Start
2010-07-01
Project End
2023-07-31
Budget Start
2019-08-08
Budget End
2020-07-31
Support Year
10
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Columbia University (N.Y.)
Department
Pathology
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
Poon, Carole L C; Liu, Weijie; Song, Yanjun et al. (2018) A Hippo-like Signaling Pathway Controls Tracheal Morphogenesis in Drosophila melanogaster. Dev Cell 47:564-575.e5
Rosa, Jeffrey B; Metzstein, Mark M; Ghabrial, Amin S (2018) An Ichor-dependent apical extracellular matrix regulates seamless tube shape and integrity. PLoS Genet 14:e1007146
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
Ghabrial, Amin S (2012) A sweet spot in the FGFR signal transduction pathway. Sci Signal 5:pe1
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; 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