The directed rearrangement of epithelial cells is crucial during human development, including the events of gastrulation and neurulation. How basolateral protrusive activity contributes to this process is poorly understood. Dorsal intercalation in the epidermis of the C. elegans embryo is an outstanding model system for examining basolateral events during epithelial cell rearrangement at the level of single cells. We have shown that dorsal intercalation relies on a phylogenetically conserved Rac/RhoG cassette activated by the Rho family guanine nucleotide exchange factor (GEF), UNC-73/Trio, and is negatively regulated by the highly conserved actin capping protein regulator, CRML-1/CARMIL (capping protein-, Arp2/3- and myosin I-linker protein). This new proposal will investigate how this conserved Rac/RhoG cassette regulates directed cell rearrangement by performing experiments in four areas: (1) We will use embryological and structure-function approaches to determine how CRML- 1/CARMIL localizes to the rear of intercalating cells; (2) We will use biochemical and in vivo rescue experiments to determine whether CRML-1 negatively regulates the Trio/Rac/RhoG pathway via direct binding or indirectly through its effects on capping protein recruitment; (3) We will determine whether a Slit/Robo/srGAP cassette regulates Rac via localized repulsion or as a ?cell contact buffer?; and (4) We will determine how additional components regulate the Trio/Rac/RhoG pathway using candidate screens and forward genetic approaches. As a result of these studies, we will elucidate a novel pathway regulating cell intercalation via basolateral protrusive activity, a widespread process with implications for understanding major birth defects and normal human embryogenesis.
Understanding how cells directionally change position in sheets of tissue is important for understanding many common birth defects, including during the formation of the human central nervous system, and the pathways that regulate these movements are misregulated in cancer cells as they lose their connections to one another and invade the body. This proposal examines the machinery that controls how cells migrate by examining proteins that regulate the key proteins Rac and RhoG. These proteins regulate how cells extend appendages that allow them to crawl in a specific direction. By studying how these proteins work in living embryos, we will gain important information that serves as the backdrop for understanding human disease.!
