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R. K. Tittle, R. Sze, A. Ng, R. J. Nuckels, J. M. Gross; Uhrf1 and Dnmt1 Function is Required for Vertebrate Lens Development. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2352.
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© ARVO (1962-2015); The Authors (2016-present)
DNA methylation results in epigenetic transcriptional repression, and this is a mechanism whereby cell-type specific gene expression patterns are set during terminal differentiation. The specific role of DNA methylation in terminal differentiation and organogenesis has so far been studied in very few cell types, due in part to the early embryonic lethality of knockout mice lacking genes required for DNA methylation. Unlike mouse models, zebrafish with mutations in two key epigenetic regulators, DNA Methyltransferase 1 (dnmt1) and Ubiquitin-like, Containing PHD and RING Finger Domains 1 (uhrf1), survive to late embryonic stages, at which time many complex organs (including the eye) have formed. We have utilized these mutant zebrafish lines to study the role of DNA methylation in lens development.
Lens formation has been assayed using histological and immunohistochemical techniques. In situ hybridizations have been performed to identify relevant mRNA distribution in the eye. Genomic DNA methylation has been assayed using a SouthWestern slot blot, and by comparison of methylation-sensitive and insensitive restriction enzyme cleavage. To study lens- and cell-autonomy of the phenotype, whole lens transplants and shield-stage transplants (used to generate embryos with mosaic lenses) have been performed.
Loss of Dnmt1 and/or Uhrf1 function leads to morphologically abnormal lenses which contain disorganized and apoptotic lens fibers. Genomic DNA methylation in dnmt1 and uhrf1 mutants is reduced by 75%, however many genes required for lens fiber cell differentiation have reduced expression in mutant lenses compared to siblings, suggesting that decreased DNA methylation does not lead to universal gene upregulation. The results of lens transplant experiments demonstrate that Uhrf1 and Dnmt1 functions are required lens-autonomously, but perhaps not cell-autonomously, during lens development in zebrafish.
This study provides the first evidence demonstrating that Uhrf1 and Dnmt1 function is required for vertebrate lens development. In the absence of either Uhrf1 or Dnmt1 function, differentiating lens fiber cells are not properly maintained, leading to apoptosis, severe defects in lens morphology, and cataracts.
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