ADAPTIVE RESISTANCE TO NITRIC OXIDE

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Nitric oxide (NO) is produced in mammalian cells by several NO synthases, including an inducible form with high output during inflammation. Since NO may contribute to carcinogenesis and age- related neurodegeneration, we wish to understand how mammalian cells counteract NO toxicity. We have identified a group of 12 proteins induced in normal human fibroblasts by exposure to subtoxic levels of NO, including the antioxidant enzyme heme oxygenase 1 (HO-1). We have shown that the induction of HO-1 mRNA occurs via NO-dependent changes in its stability. We will explore the mechanism of this regulation and try to define the signal transduction pathway for post-transcriptional control of HO-1. Other genes (e.g., MAP kinase phosphatase MKP-1/CL100) are controlled transcriptionally, and we will test specific transcription regulators for their roles in this expression. We have identified an adaptive resistance to NO in the rodent motor neurons: cells exposed to low-level NO that induces HO-1 become resistant to much higher (usually cytotoxic) levels of NO. We will characterize this response in various cell types by identifying NO-inducible genes and determining their roles in protection against NO-mediated toxicity. HO-1 knockout mice will allow us to define the defensive role of HO-1 in fibroblasts, neurons, and other tissues. This project involves parallel efforts to follow biochemical markers of nitric oxide damage (e.g., nitrotyrosine), to use genetics to establish the role of HO-1 in inducible resistance to NO, to elucidate regulatory and signal transduction pathways, and to identify and clone novel defense genes for NO resistance.