Activation of autophagy in the outer retina exhibits a bimodal pattern that correlates with shifts in transduction proteins within the photoreceptor and by circadian ingestion of outer segments in the RPE. These dynamic shifts suggest a critical role for this pathway in maintaining homeostasis, with further study needed to define the mechanisms underlying the regulation of this phenomenon.

Senescence is a permanent growth arrest that restricts the lifespan of primary cells in culture, and represents an in vitro model for aging. Senescence functions as a tumor suppressor mechanism that can be induced independent of replicative crisis by diverse stress stimuli. RNase-L mediates antiproliferative activities and functions as a tumor suppressor in prostate cancer, therefore, we examined a role for RNase-L in cellular senescence and aging. Ectopic expression of RNase-L induced a senescent morphology, a decrease in DNA synthesis, an increase in senescence-associated b-galactosidase activity, and accelerated replicative senescence. In contrast, senescence was retarded in RNase-L-null fibroblasts compared with wildtype fibroblasts. Activation of endogenous RNase-L by 2-5A transfection induced distinct senescent and apoptotic responses in parental and Simian virus 40-transformed WI38 fibroblasts, respectively, demonstrating cell type specific differences in the antiproliferative response to RNase-L activation. Replicative senescence is a model for in vivo aging; therefore, genetic disruption of senescence effectors may impact lifespan. RNase-L À/À mice survived 31.7% (Po0.0001) longer than strain-matched RNase-L +/+ mice providing evidence for a physiological role for RNase-L in aging. These findings identify a novel role for RNase-L in senescence that may contribute to its tumor suppressive function and to the enhanced longevity of RNase-L À/À mice.

Photoreceptor (PR) cells receive oxygen and nutritional support from the underlying retinal pigment epithelium (RPE). Retinal detachment results in PR hypoxia and their time-dependent death. Detachment also activates autophagy within the PR, which serves to reduce the rate of PR apoptosis. In this study, we test the hypothesis that autophagy activation in the PR results, at least in part, from the detachment-induced activation of hypoxia-inducible factors (HIF). Retina-RPE separation was created in Brown-Norway rats and C57BL/6J mice by injection of 1% hyaluronic acid into the subretinal space. Retinas were harvested and assayed for HIF protein levels. Cultured 661W photoreceptor cells were subjected to hypoxic conditions and assayed for induction of HIF and autophagy. The requirement of HIF-1α and HIF-2α in regulating photoreceptor autophagy was tested using siRNA in vitro and in vivo. We observed increased levels of HIF-1α and HIF-2α within 1 day post-detachment, as well as increased levels of BNIP3, a downstream target of HIF-1α that contributes to autophagy activation. Exposing 661W cells to hypoxia resulted in increased HIF-1α and HIF-2α levels and increase in conversion of LC3-I to LC3-II. Silencing of HIF-1α, but not HIF-2α, reduced the hypoxia-induced increase in LC3-II formation and increased cell death in 661W cells. Silencing of HIF-1α in rat retinas prevented the detachment-induced increase in BNIP3 and LC3-II, resulting in increased PR cell death. Our data support the hypothesis that HIF-1α, but not HIF-2α, serves as an early response signal to induce autophagy and reduce photoreceptor cell death.