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B. K. Chauhan, A. DiSanza, S.-Y. Choi, M. Lou, H. E. Beggs, G. Scita, Y. Zheng, R. A. Lang; Mammalian Lens Placode Invagination Requires Cdc42, RhoA and Rac1. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4774.
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The formation of the mammalian lens pit provides an excellent model to study the morphogenetic pathways involved in epithelial invagination. In this study, we attempt to address the importance of three chief small Rho GTPases, Cdc42, RhoA and Rac1, in this process.
Three conditional mouse mutants of Cdc42, RhoA and Rac1 were crossed with the le-cre mouse line to generate corresponding deleted alleles only in the induced lens ectoderm. Analysis, including immunofluorescent-staining and quantification, was conducted at specific stages of lens placode invagination.
We show that F-actin-rich filopodia link adjacent presumptive lens and retina. The filopodia, most of which originate in the presumptive lens, form at E9.5 when presumptive lens and retina first come into close contact, and have retracted by E11.5 when invagination is complete. Formation of filopodia is dependent on Cdc42 and its effector IRSp53 (Baiap2). Loss of filopodia results in reduced lens pit invagination. Pharmacological manipulation of the actin-myosin contraction pathway showed that the filopodia can respond rapidly in length to change the inter-epithelial distance.Next, we show that there are apical actomyosin complexes in the lens placode that respond to pharmalogical manipulation, thereby reducing invagination. We observe that activated myosin and F-actin is reduced in RhoA mutant lens pits, therefore producing shallower pits. In contrast, early invagination of placodes occurs in the Rac1 mutants compared to wild-types. Interestingly, the lens pits are thinner in the Rac1 mutants, but noticeably thicker for the RhoA mutants.
Collectively, our results suggest that basal lens filopodia provide a physical tether that coordinates invagination and correct positioning of the lens in the early eye. The role of RhoA is to regulate the apical actomyosin complexes that drive invagination, and Rac1 is responsible for migration of the lens placode towards the presumptive retina.
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