Occasionally in humans neural tube formation fails to work normally and an infant is born with a neural tube defect. Experimentally, to model this congenital problem, mice have been shown to exhibit neural tube defects, in some cases with mutations in the same genes that affect humans. This proposal will look at adhesion and cytoskeletal contributions to neural tube closure in a highly quantitative way as a measure of this morphogenetic process. Six major adhesive components to be measured include N-CAM, N-Cadherin, E-Cadherin, cell-laminin, cell-fibronectin, and cell-versican. A highly quantitative adhesion assay was miniaturized to assess regional adhesive forces using very small numbers of cells.
The specific aims i nclude: (1) To test the hypothesis that a wave of cell-cell adhesion changes progresses from the site of first fusion of the neural tube to the caudal end. The investigators will separately measure contributions of six adhesion molecules in each progressive region of the neural tube. The data will reveal the changing role of adhesion in neural tube closure. An assay that distinguishes between cell-ligand affinities and cytoskeletal contribution to the adhesion will be used so both parameters will be followed temporally and spatially as the neural tube forms. (2) To test the hypothesis that homozygous mutants for patterning molecules, with known neural tube closure defects, will have significant deficits in adhesion as a consequence. This hypothesis has often been suggested but has not been tested. Results from Specific Aim 1 will establish the norm. It is predicted that one or more of the six adhesion systems will vary significantly from the norm in the mutants. An overproduction of an adhesion molecule might make the cells too adhesive for morphogenesis, the system might fail to utilize the cytoskeleton adequately, or it might have insufficient adhesion. Each of these parameters will be tested. (3) To test the hypothesis that heterozygotes will have minor adhesion deficits and double heterozygotes will have more significant adhesion deficits. The ability to quantify adhesion and cytoskeletal contributions will allow to determine whether heterozygotes with a low or negligible penetrance of neural tube defects will nevertheless have measurable deficits in adhesion.
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