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Dennis S. Rice, Matthew M. Newhouse, David Potter, Mike J. Crist, Nianhua Xu, Gwenn M. Hansen, Alejandro Abuin, Brian P. Zambrowicz, Peter J. Vogel; Limk2 Controls Epithelial Sheet Migration During Eyelid Development. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4914.
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Genetic disruption of several genes involved in actin cytoskeleton remodeling induces an eyes open at birth (EOB) phenotype. We have used retroviral gene trapping to disrupt the expression of all known isoforms of LIM motif-containing protein kinase 2 (Limk2). Limk2 regulates the biochemical function of actin binding proteins such as cofilin. Mice deficient in Limk2 (Limk2-/-) exhibit an EOB phenotype. The purpose of this study is to investigate the molecular mechanisms associated with the EOB phenotype.
Histology was performed on embryos and postnatal mice at relevant stages of eyelid development. In situ hybridization, immunohistochemistry and immunoblotting techniques were used to investigate Limk2 expression and function during eyelid development.
The EOB phenotype is fully penetrant in Limk2-/- mice, whereas eyelid development is comparable between wild type (Limk2+/+) and heterozygous (Limk2 +/-) mice. Limk2 is expressed in palpebral epidermis and conjunctiva during eyelid formation. Periderm cells also express high levels of Limk2. The EOB phenotype becomes apparent at E15.5 in Limk2-/- mice, when the epithelial sheet fails to migrate over the corneal surface. Levels of phospho-cofilin, a biochemical substrate of Limk2, and of F-actin are decreased in the Limk2-/- eyelids.
Inactivation of Limk2 results in an EOB phenotype. This phenotype arises through failure in epithelial sheet migration associated with decreased levels of F-actin. Decreased levels of F-actin presumably arise as a result of the increased actin-severing activity of non-phosphorylated cofilin in the knockout eyelid. These results demonstrate that proper eyelid development requires F-actin nucleation in migrating epithelial cells.
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