Abstract
Vision loss impacts >30 million people worldwide. Mutations in over 50 genes can predispose to the retinal-degenerative disease retinitis pigmentosa (RP). Despite this clear genetic component, disease progression in RP is potentially linked to environmental factors such as light, which can induce the formation of reactive oxygen species leading to oxidative stress and oxidative DNA damage and inducing photoreceptor cell death. Alkylating agents are used to produce animal models of RP as DNA alkylating agents induce photoreceptor cell death. This cytotoxicity is mediated via the DNA repair protein alkyladenine DNA glycosylase (Aag). Hence, we hypothesise that Aag-mediated processing of DNA base damage in photoreceptors affects their function and the light responses in the animal. In-vivo, light/dark cycle entrained wild-type and Aag null mice were treated with methyl methanesulfonate (MMS), under constant light or dark conditions, and harvested 6, 48, and 72-hours post-treatment for histopathological and gene expression analysis. In-vitro, wild-type and Aag null primary embryonic fibroblast cells (pMEFs) were treated with MMS at different circadian phases, and assessed for their circadian phase-response to MMS. Marginal light exacerbation of Aag-mediated retinal-degeneration upon alkylation-exposure is observed in wild-type retinas. Photoreceptor cell function is apparently affected basally and upon alkylation-exposure in wild-type and Aag null mice. Mice melanopsin expressing retinal ganglion cells are resistant to alkylation-damage with no reduction in melanopsin expression. Early gene expression analysis of wild-type and Aag null mice tissues in response to MMS-treatment showed a link between Aag status and upregulation of key cellular stress response mechanisms. MMS seemed to reset the circadian clock in pMEFs in a circadian-time dependent-manner without involving Aag. Aag-deficience can rescue alkylation sensitivity in specific cell types via reducing BER-intermediates accumulation, but Aag is required to play a role in the transcriptional activation of several key genes associated with cell survival, death and function upon alkylation-exposure.