A molecular analysis of ventral enclosure in Caenorhabditis elegans

• Main Author: Raich, William B
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-1999
• Subjects: Cellular biology; Genetics; Molecular biology
• ISBN: 0599325380; 9780599325388

During the ventral enclosure of the Caenorhabditis elegans embryo, an epithelial sheet of cells called the hypodermis, or embryonic epidermis, spreads in a contralateral fashion from the dorsal surface of the embryo, ultimately wrapping the embryo in an epithelial monolayer. I have identified requirements for hmr-1 cadherin, hmp-2 β-catenin, hmp-1 α-catenin, and zen-4 kinesin during this process. HMP-1, HMP-2, and HMR-1 co-localize to adherens junctions, but mutant analysis indicates that these proteins are not essential for general cell adhesion. Instead, analysis of hmp-1, hmp-2, and hmr-1 suggests that these gene products anchor actin filament bundles at adherens junctions, thereby transmitting the force of bundle contraction into cell shape changes. During ventral enclosure, a translational fusion between HMP-l and Green Fluorescent Protein (GFP) localizes to filopodia extended by a subset of migrating epidermal cells called the leading cells. Cadherin mediated adhesive strengthening is specifically required to stabilize filopodial adhesion, since other ventral epidermal cells do not display long filopodia and successfully adhere following hmr-1 inactivation. These results indicate that epithelial cells that use filopodia for migration require α-catenin recruitment to sites of nascent junction formation to correctly regulate the assembly of mature junctions. Zygotic mutations in zen-4 prevent ventral enclosure. I cloned zen-4, and found that it encodes a kinesin-like protein belonging to the CHOl/MKLP1 subfamily. ZEN-4 antisera and a translational fusion between ZEN-4 and GFP reveal that ZEN-4 is strongly expressed in dividing cells, localizing to the midzone of the mitotic spindle. Embryos deprived of maternal and zygotic zen-4 gene product form multinucleate single-celled embryos, as they continue to cycle through mitosis but fail to complete cytokinesis. In these embryos, the midzone microtubules appear disorganized, indicating that ZEN-4 directly crosslinks the midzone microtubules. It is likely that zygotic mutations in zen-4 disrupt ventral enclosure indirectly, since homozygous zen-4 mutants contain multinucleate cells on their ventral surface immediately preceding enclosure. The work presented in this dissertation indicates that ventral enclosure in C. elegans will be a useful system for elucidating the intricate molecular mechanisms that underlie epithelial morphogenesis.