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priyanka arya, Mark Rainey, Sohinee Bhattacharyya, Bhopal Mohapatra, Murali Kuracha, Vimla Band, venkatesh govindarajan, Hamid Band; The Endocytic Recycling Regulatory Protein EHD1 Is Required for Ocular Lens Development. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4008.
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© ARVO (1962-2015); The Authors (2016-present)
The C-terminal Eps15 homology domain-containing (EHD) proteins play a key role in endocytic recycling, a fundamental cellular process that ensures the return of endocytosed membrane components and receptors back to the cell surface. Towards understanding the biological roles of mammalian EHD family proteins, we and others have begun to use mouse gene deletion models.<br /> Here, we provide evidence for a critical requirement of EHD1 for development of ocular lens and cornea. Our studies show that EHD1-/- mice display pleiotropic ocular phenotypes, including anophthalmia, microphthalmia and congenital cataracts.
EHD1 CKO were generated by crossing EHD1 floxed mice with Le-Cre transgenic mice. For timed-pregnancy experiments, the noon of vaginal plug detection was considered E0.5. Pregnant dams were euthanized, embryos collected, fixed in 10% formalin, transferred to 70% ethanol prior to paraffin embedding. For antibody staining, rehydrated tissues were boiled in antigen unmasking solution, blocked, incubated with primary antibodies, followed by dectection with flurosecent secondary antibodies.
Histological analyses showed pleiotropic ocular defects in EHD1-/- mice that included a smaller or absent lens, persistence of lens stalk and the hyaloid vasculature, and deformed optic cup. To investigate whether these profound ocular defects resulted from a role in lens versus optic vesicle, we deleted EHD1 selectively in the presumptive lens ectoderm using Le-Cre. EHD1 deletion in the lens resulted in developmental defects that included thinner epithelial layer with reduced cell count, smaller lenses and absence of corneal endothelium. Lens epithelium-specific EHD1 mutants exhibited reduced proliferation and cell survival, and downregulation of junctional proteins such as E-cadherin and ZO-1.
The ocular phenotypes caused by the loss of a single regulator of endocytic recycling, EHD1, provide a novel model system to elucidate mechanistic links between surface receptor recycling and the cellular processes that control orderly development of the compartments of mammalian eye. Taken together, these data reveal a unique cell-autonomous role for EHD1 in early lens development and suggest a previously unknown link between endocytic recycling pathway and regulation of key developmental processes events including proliferation, differentiation and morphogenesis.
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