Chromatin condensation as a DNA protective response to UV radiation in human melanocytes and aging C. elegans

紫外线诱导的DNA损伤是与年龄相关的皮肤病和光老化的主要原因。虽然身体 可以随着时间的推移发展对紫外线应激的防御（即，黑色素浓度和层增厚 角质层），在理解细胞是否可以立即有效地 对紫外线的保护反应。我们最近发现了一种直接和强大的紫外线诱导的DNA保护 这一机制涉及由钙离子内流触发的整体染色质致密化。染色质 在人HeLa细胞中证实了致密化和DNA保护反应， 在NIH 2/4小鼠胚胎成纤维细胞和蛔虫C.优雅我们的长期 我们的目标是了解这种机制的分子基础，并研究它是否可以被操纵， 增加我们对紫外线伤害的天然保护。我们将探讨这种机制是否会随着年龄的增长而衰退 就像其他抗压机制一样。本申请的目标是开发C。优雅作为一个 模型系统，以探索这种应激抗性机制的年龄依赖性，并使用人表皮 黑素细胞与C.第一次看到这种紫外线的分子途径， 诱导DNA保护性染色质致密化。中心假设是染色质致密化是 由感光细胞的保守分子机制触发，通过Gαq/11偶联的 光转导途径激活钙内流。我们假设这是一个进化上保守的 在老年生物体中效率较低的反应。选择C的理由。它是一种很好的 建立了研究衰老的模型系统，使用了极好的遗传和发育工具。人原代 选择表皮黑素细胞是因为它们是人类皮肤细胞，其中涉及 光感受器和Gαq/11被证明控制UV诱导的钙内流。具体目标是 研究内容包括：1）检测紫外线诱导的染色质致密化是否能保护人类DNA免受进一步的损伤 表皮黑素细胞（HEMs）和青年和老年C.优雅我们用紫外光照射HEMs和C。elegans Southwestern印迹法检测光产物的去除率（HEMs和C. elegans）和 vivo（C. elegans）。2)确定UVR诱导的染色质致密化是否涉及Gαq/11偶联的 人表皮黑素细胞和C.优雅这一目标将 通过RNAi/siRNA敲唐斯. elegans和人原代表皮黑素细胞。的 这项研究是创新的，因为它探索了一种以前从未描述过的DNA防御机制， 紫外线辐射和它设置为目标建立C。作为一个模型系统，将这种现象与 衰老增加我们的创新能力是一个C。elegans实验室和a 哺乳动物染色质实验室结果将是重要的，因为控制DNA的新的潜在目标- 保护途径将被发现，这可以导致更好的药物策略，以保护免受紫外线。

UV-induced DNA damage is a major cause of age-related skin diseases and photoaging. Although the body can develop defenses over time to UV stress (i.e., melanin concentration and thickening of the stratum corneum), there is a fundamental gap in understanding whether cells can mount an effective immediate protective response to UV. We recently discovered an immediate and robust UV-induced DNA protection mechanism that involves a global chromatin compaction triggered by calcium influx. The chromatin compaction and DNA protection responses were demonstrated in human HeLa cells, and the compaction was also demonstrated in NIH2/4 mouse embryonic fibroblasts and in the roundworm C. elegans. Our long-term goal is to understand the molecular basis of this mechanism, and to investigate if it can be manipulated to increase our natural protection from UV damage. We will explore whether this mechanism declines with age like other stress resistance mechanisms. The objectives of this application are to develop the C. elegans as a model system to probe the age-dependence of this stress resistance mechanism and to use human epidermal melanocytes in combination with C. elegans to gain the first glimpse into the molecular pathway of this UV- induced DNA protective chromatin compaction. The central hypothesis is that chromatin compaction is triggered by a conserved molecular machinery of a photoreceptor acting through the Gαq/11-coupled phototransduction pathway to activate a calcium influx. We hypothesize that this is an evolutionarily conserved response that is less efficient in aged organisms. The rationale for choosing C. elegans is that it is a well- established model system for studying aging with superb genetic and developmental tools. Human primary epidermal melanocytes were chosen because they are human skin cells in which a specific pathway involving photoreceptors and Gαq/11 was shown to control a UV-induced calcium influx. The specific aims for this research are: 1) Test whether UV-induced chromatin compaction protects DNA from further damage in human epidermal melanocytes (HEMs) and in young and old C. elegans. We will UV irradiate HEMs and C. elegans and detect the rate of removal of photoproducts by Southwestern blotting (for HEMs and C. elegans) and in vivo (for C. elegans). 2) Determine whether UVR-induced chromatin compaction involves the Gαq/11-coupled phototransduction pathway and calcium influx in human epidermal melanocytes and C. elegans. This aim will be achieved by RNAi/siRNA knock downs in C. elegans and human primary epidermal melanocytes. The proposed research is innovative because it explores a previously undescribed DNA defense mechanism from UV radiation and it sets as a goal establishing C. elegans as a model system for relating this phenomenon to aging. Adding to our innovative capacity is the ongoing cross-pollination between a C. elegans lab and a mammalian chromatin lab. The results will be significant because new potential targets that control the DNA- protective pathway will be discovered which can lead to better pharmaceutical strategies to protect from UV.