Convergent extension is the directed intercalation of cells in an embryonic tissue along a preferred axis, resulting in dramatic elongation of the tissue. Convergent extension is a common yet poorly understood process. In particular, virtually nothing is known about the cellular mechanisms or molecular control of convergent extension of epithelial tissues. This proposal seeks to investigate the cellular and molecular mechanisms of convergent extension in an epithelium, using the hypodermis, or embryonic epidermis, of the C. elegans embryo as a model system. The Hardin laboratory has performed extensive analysis of cell intercalation in the dorsal hypodermis of wild-type embryos, and has shown that intercalation involves the extension of basolateral protrusions in advance of rearrangement of junctional complexes. Intercalation of dorsal hypodermal cells in C. elegans is highly ordered and reproducible, and constitutes the simplest known system for studying convergent extension in vivo. In addition, studying this process in C. elegans allows the use of the powerful genetic and molecular techniques available in this system. Using "4D microscopy", laser scanning confocal and multiphoton excitation microscopy in conjunction with translational fusions of the green fluorescent protein (GFP) and adherens-junction associated proteins, as well as the membrane dye FM4-64, this proposal seeks to answer the following questions:
(1) What role does adherens junctional adhesion play during dorsal intercalation? Mutations have been characterized in three components of cadherin-mediated cell adhesion in C. elegans: a-catenin (encoded by the hmp-1 locus), b-catenin (encoded by the hmp-2 locus) and a cadherin homologue (encoded by the hmr-1 locus). Dorsal intercalation will be examined in hmr-1 and hmp-1 homozygotes as well as embryos from germline mosaic hermaphrodites which lack both maternal and zygotic hmr-1 and hmp-1 mRNA. (2) What role does the novel gene die-1 play during dorsal intercalation? A novel mutation has been isolated in a gene that results in specific defects in both dorsal intercalation and elongation (the Die phenotype). The die-1 gene has been cloned; it encodes a zinc-finger protein of the Kruppel class of putative transcriptional regulators. The phenotypic defects will be characterized in die-1 homozygotes, the molecular lesion in die-1(w34) mutants will be determined, and the tissue-specific expression pattern of die-1 in embryos will be determined. (3) What role does regional patterning play in regulating convergent extension? Mutations in several genes result in severe morphogenetic defects in the posterior of C. elegans embryos, including dorsal hypodermal cells derived from the C founder cell. These include the gene pal-1 (a caudal homologue), nob-1 (a homeodomain protein) and vab-7 (an even-skipped homologue). Possible dorsal intercalation defects will be examined in these patterning mutants, as well as alterations in the expression of die-1.
As a result of these studies, fundamental insights will be gained into the mechanisms of convergent extension, a process that is fundamental to a diverse set of morphogenetic events, including gastrulation, neurulation, wound healing, and many other morphogenetic movements.
