Identification of a novel hypoxia-inducible factor 1-responsive gene, RTP801, involved in apoptosis.

Hypoxia is an important factor that elicits numerous physiological and pathological responses. One of the major gene expression programs triggered by hypoxia is mediated through hypoxia-responsive transcription factor hypoxia-inducible factor 1 (HIF-1). Here, we report the identification and cloning of a novel HIF-1-responsive gene, designated RTP801. Its strong up-regulation by hypoxia was detected both in vitro and in vivo in an animal model of ischemic stroke. When induced from a tetracycline-repressible promoter, RTP801 protected MCF7 and PC12 cells from hypoxia in glucose-free medium and from H(2)O(2)-triggered apoptosis via a dramatic reduction in the generation of reactive oxygen species. However, expression of RTP801 appeared toxic for nondividing neuron-like PC12 cells and increased their sensitivity to ischemic injury and oxidative stress. Liposomal delivery of RTP801 cDNA to mouse lungs also resulted in massive cell death. Thus, the biological effect of RTP801 overexpression depends on the cell context and may be either protecting or detrimental for cells under conditions of oxidative or ischemic stresses. Altogether, the data suggest a complex type of involvement of RTP801 in the pathogenesis of ischemic diseases.

Inhibition of oxygen-induced retinopathy in RTP801-deficient mice.

PURPOSE: Ischemic proliferative retinopathy, which occurs as a complication of diabetes mellitus, prematurity, or retinal vein occlusion, is a major cause of blindness worldwide. In addition to retinal neovascularization, it involves retinal degeneration, of which apoptosis is the main cause. A prior report has described the cloning of a novel HIF-1-responsive gene, RTP801, which displays strong hypoxia-dependent upregulation in ischemic cells of neuronal origin, both in vitro and in vivo. Moreover, inducible overexpression of RTP801 promotes the apoptotic death of differentiated neuron-like PC12 cells and increases their sensitivity to ischemic injury and oxidative stress. The purpose of the study was to examine the potential role of RTP801 in the pathogenesis of retinopathy, using RTP801-deficient mice. METHODS: Wild-type and RTP801-knockout mice were used in a model of retinopathy of prematurity (ROP). Their retinas were collected at postnatal day (P)14 and P17. They were examined by fluorescein angiography and by analysis of VEGF expression, neovascularization, and apoptosis. RESULTS: The expression of RTP801 was induced in the wild-type retina after hypoxia treatment. The retinal expression of VEGF after transfer to normoxic conditions was similarly upregulated in both wild-type and knockout mice. Nevertheless, the retinas of the RTP801-knockout mice in an ROP model showed a significant reduction in retinal neovascularization (P < 0.0001) and in the number of apoptotic cells in the inner nuclear layer (P < 0.0001). CONCLUSIONS: In the absence of RTP801 expression, development of retinopathy in the mouse model of ROP was significantly attenuated, thus implying an important role of RTP801 in the pathogenesis of ROP.

Rtp801, a suppressor of mTOR signaling, is an essential mediator of cigarette smoke-induced pulmonary injury and emphysema.

Rtp801 (also known as Redd1, and encoded by Ddit4), a stress-related protein triggered by adverse environmental conditions, inhibits mammalian target of rapamycin (mTOR) by stabilizing the TSC1-TSC2 inhibitory complex and enhances oxidative stress-dependent cell death. We postulated that Rtp801 acts as a potential amplifying switch in the development of cigarette smoke-induced lung injury, leading to emphysema. Rtp801 mRNA and protein were overexpressed in human emphysematous lungs and in lungs of mice exposed to cigarette smoke. The regulation of Rtp801 expression by cigarette smoke may rely on oxidative stress-dependent activation of the CCAAT response element in its promoter. We also found that Rtp801 was necessary and sufficient for nuclear factor-kappaB (NF-kappaB) activation in cultured cells and, when forcefully expressed in mouse lungs, it promoted NF-kappaB activation, alveolar inflammation, oxidative stress and apoptosis of alveolar septal cells. In contrast, Rtp801 knockout mice were markedly protected against acute cigarette smoke-induced lung injury, partly via increased mTOR signaling, and, when exposed chronically to cigarette smoke, against emphysema. Our data support the notion that Rtp801 may represent a major molecular sensor and mediator of cigarette smoke-induced lung injury.