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B.S. Biology, 2003
Pontifical Catholic University of Puerto Rico
Ponce, Puerto Rico
Thesis Advisor: Sunita G. Kramer, PhD
Graduate Program in Cell & Developmental Biology
Life Science Building Auditorium
Tuesday, November 25, 2008
Heart morphogenesis requires the coordinated regulation of cell movements and cell-cell interactions between distinct populations of cardiac precursor cells. In this study, I analyzed the role of Slit, an extracellular matrix protein and its transmembrane receptors Roundabout (Robo) and Roundabout2 (Robo2) during morphogenesis of the Drosophila heart tube, a process analogous to early heart formation in vertebrates. During heart assembly, two types of progenitor cells align into rows and coordinately migrate to the dorsal midline of the embryo, where they meet to assemble a linear heart tube. Here I present that cardiac-specific expression of Slit is required to maintain adhesion between cells within each row during dorsal migration. Moreover, differential Robo expression determines the relative distance each row is positioned from the dorsal midline. The innermost cardioblasts (CB) express only Robo, whereas the flanking pericardial cells express both receptors. Removal of robo2 causes pericardial cells to shift toward the midline, whereas ectopic robo2 in CBs drives them laterally, preventing the heart tube from forming. I propose a model in which Slit has a dual role during assembly of the linear heart tube, functioning to regulate both cell positioning and adhesive interactions between migrating cardiac precursor cells.
After contralateral CBs meet at the dorsal midline, a process guided by Slit and Robo, they must form a lumen between the apical surfaces of the cells. Here, I present a second function for Slit and Robo during heart tube assembly. They are both required at CB apical domains for lumen formation. Mislocalization of Slit outside the apical domain causes ectopic lumen formation and the mislocalization of cell junction proteins, E-cadherin (E-Cad) and Enabled, without disrupting overall CB cell polarity. Ectopic lumen formation is suppressed in robo mutants, which indicates robo`s requirement for this process. I provide genetic evidence suggesting that Robo and Shotgun (Shg)/E-Cad function together in modulating CB adhesion. In robo loss-of-function or shg/E-Cad gain-of-function embryos, lumen formation is blocked because of inappropriate CB adhesion and an accumulation of E-Cad at the apical membrane. In contrast, shg/E-Cad loss-of-function or robo gain-of-function blocks lumen formation due to a loss of CB adhesion. This data shows that Slit and Robo pathways function in lumen formation as a repulsive signal to antagonize E-Cad-mediated cell adhesion. In this dissertation, I found that Slit and Robo receptors function in two distinct steps during Drosophila heart tube formation. First, keeping heart cells adhered to each other while migrating to the midline and the second, regulating apical cell adhesion during lumen formation.