Philip C. Hanawalt, Ph. D.
Stanford University School of Medicine

Repair of Oxidative DNA Damage in Human Neurons.

Accumulation of deleterious alterations in neuronal DNA has been invoked in models for neurological diseases and aging. While there has been a fair amount of speculation, there have been few definitive studies in this important field. We have been studying the excision repair of DNA damage induced by ultraviolet light in terminally differentiated human neurons, as compared to that in their precursor cells. A striking attenuation of overall genomic repair was observed in adult differentiated neurons. It could be rationalized that neurons do not repair the bulk of their genome because they will never again need to replicate this DNA, nor even transcribe most of it. However, they obviously do need to maintain the integrity of those genes that are still being expressed. Indeed, we have observed that transcribed genes are very efficiently repaired in differentiated neurons. Unlike the situation for so-called transcription-coupled DNA repair in growing cells, in which the transcribed strand is preferentially repaired, we have found that both DNA strands of active genes are repaired efficiently in these neurons. This mechanism may be required when global genomic repair has been shut off, because the non-transcribed strand is needed as a template to repair the transcribed strand. If neurons were to accumulate lesions in non-transcribed strands over an extended period, it would become increasingly likely that mutations would be introduced when using that damaged template for repair. A deficiency in this differentiation associated repair would be expected to result in age-related inactivation of genes, leading to progressive metabolic dysfunction and ultimately to premature neuron "aging" and cell death, causing early dementia.

Our analysis of gene expression profiles has thus far revealed that genes encoding several important repair enzymes are remarkably upregulated in adult neurons. One aim of our research is to examine the relevance of this gene induction to the unique repair phenotype of mature neurons. Another aim is to determine the potential role of the activated p53 tumor suppressor gene in the proficient repair of expressed genes in neurons.

We are extending our analysis to the case of endogenous oxidative base damage, that is a more likely threat than ultraviolet light for aging neurons. Recent work has shown that some oxidative DNA lesions, normally repaired by the base excision repair pathway, are also subject to transcription-coupled repair. These lesions are expected to cause arrest of translocating RNA polymerases which may generate a strong signal for a p53-dependent pathway of apoptosis. That could account for the excessive neuronal cell death in patients with Cockayne syndrome, in which transcription coupled DNA repair is defective, and it could also contribute to neuronal dysfunction in elderly humans.

Our experiments are designed to critically test the hypothesis that oxidative base damage may accumulate in the bulk DNA in differentiated neurons but that it continues to be efficiently repaired in expressed genes. We will examine human hereditary syndromes and clinical situations in which the maintenance of these essential genes may be impaired.

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