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R. Ayyagari, N. Dobri, B. Chang, V. R. Chavali, V. Vasireddy; Inner Retinal Morphology of Mice With a Null Mutation in the Mdm1 Gene and Interaction of This Protein With DNA. Invest. Ophthalmol. Vis. Sci. 2010;51(13):6140.
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© ARVO (1962-2015); The Authors (2016-present)
A null mutation in the mouse double minute-1 (Mdm1) gene that codes for a nuclear protein has been shown to cause autosomal recessive late-onset photoreceptor degeneration (arrd2) in mice. We studied the affect of Mdm1 mutation on the retina and evaluated the function of this protein.
Retinal changes of 4 to 11 months old arrd2 mice were studied by immunohistochemsitry using retinal cell marker antibodies. To identify the interacting partners of Mdm1, ChIP on Chip analysis was carried out using cells expressing MDM1 and Gene ChIP mouse tailing 2.0 R array. Evaluation of results obtained by ChIP on Chip was carried out by amplification of chromatin immunoprecipiate with specific primers, electrophoresis mobility shift assay, immuno precipitation and western blot analysis.
The Mdm1 protein is localized to the inner retinal layers. Analysis with medium wavelength (MW) cone opsin antibodies demonstrated significant loss of MW opsin expressing cones in the retina of arrd2 mice compared to controls. Significant increase in the immunoreactivity of GFAP and presence of GFAP positive muller cells were observed in the inner half of retina as early as age 4 months in arrd2 mice. In addition the thickness of inner nuclear layer of arrd2 mice retina was found to be less compared to controls. Intensity of hybridization signal with PSD95 antibodies was significantly less in arrd2 mice when compared to age-matched controls. Analysis with calretinin indicated significant loss of amacrine cells. ChIP on chip analysis revealed that the Mdm1 protein interacts with about 800 gene sequences. Amplification of the chromatin immunoprecipitate revealed that the Mdm1 does not interact directly with common retinal transcription factors CRX and NRL. Chromatin immunoprecipitation assay revealed that Mdm1 complexes with genes involved in cell cycle regulation and DNA damage repair.
Although the onset of photoreceptor degeneration in arrd2 mice is at age 9 months, significant changes were observed in the inner retina as early as 4 months. Majority of the genes that showed interaction with the Mdm1 are involved in cell cycle and DNA repair. Further studies are underway to understand the role of Mdm1 in cell cycle and DNA repair and the mechanism underlying late-onset degeneration due to the null mutation in the Mdm1 gene.
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