Abstract
Purpose :
Incomplete separation of the lens vesicle from the surface ectoderm results in a persistent lens stalk between the lens and cornea, which clinically presents as adhesion between the cornea and the iris and/or lens in Peters anomaly (PA). This study aims to elucidate the Abl-Crk cellular signal network that regulates lens vesicle closure.
Methods :
Mouse lines carrying floxed genes of interest were crossed with the lens-specific Le-Cre (Tg/-) to generate conditional knockouts. Embryos carrying expected genes were harvested and sectioned for histochemical analysis. Immunoprecipitation and immunoblotting were carried out using primary mouse embryonic fibroblasts (MEFs) and lens epithelium cells.
Results :
Abelson murine leukemia viral oncogene homolog (Abl) and Abl related gene (Arg) mutants (Le-Cre;AblF/F;ArgF/F, AblCKO) presented incomplete separation of the lens vesicle from the surface ectoderm as early as embryonic day 10. In vitro, viral expression of Cre recombinase caused AblF/FArgF/F lens epithelium cells to scatter, losing their typical hexagon shape and adherens junctions. Heterozygous deletion of Abl substrates Crk adaptor proteins (Crk and its paralog CrkL) in the AblCKO mutants (Le-Cre;AblF/F;ArgF/F;CrkF/+;CrkLF/+) can rescue the lens stalk phenotype in an allele copy-dependent manner, and homozygous deletion of Shp2 was able to rescue the phenotype with partial penetrance. The lens epithelium in AblCKO embryos showed elevated phospho-ERK. But homozygous Abl, Arg, and Erk1/2 quadruple knockout (Le-Cre;AblF/F;ArgF/F;Erk1/2F/F) embryos still possessed lens stalks. Only homozygous deletion of Rac1 in the AblCKO mutants (Le-Cre;AblF/F;ArgF/F;Rac1F/F) was able to fully rescue the lens stalk phenotype.
Conclusions :
These genetic experimental results suggested that Crk and CrkL, as the direct substrates of Abl, serve as the linkage between Abl and the cell adhesion network. ERK1/2 and Rac1 abolition experiments demonstrated that Rac1, but not ERK, is the downstream effector of Abl in the lens-cornea separation.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.