Ion Channel and Calcium Signaling in Ultraviolet Light Transduction in Human Skin

项目总结 皮肤，人体最大的器官，经常暴露在太阳紫外线辐射(UVR)下，这是一种 强烈的环境致癌物质。与其他哺乳动物的皮肤不同，人类皮肤通过以下方式对紫外线做出反应 表皮黑素细胞产生黑素所致的色素沉着增加及其转移到 角质形成细胞。黑色素通过直接吸收UVR保护黑素细胞和角质形成细胞的核DNA 和中和自由基，从而防止遗传损伤和癌症形成。 人体皮肤是如何检测和响应紫外线的？在地球表面的UVR由95%的长 波长紫外线(UVA)和5%短波紫外线(UVB)。尽管UVB引起的机制 DNA损伤和暴露一天多后黑色素合成延迟的结果是很好的特征， 人们对UVA的影响知之甚少。高剂量UVA会导致氧化损伤，但没有特定的途径 为了检测UVA的生理剂量，已经确定了其特征。我们的初步结果描述了一部小说 黑素细胞中UVA激活的信号通路是视网膜依赖和G蛋白偶联的。激活 这一途径通过生理剂量的UVA导致瞬时受体电位A1(TRPA1)的激活。 离子通道，细胞内钙离子的快速增加，以及黑色素含量的快速增加 (从UVA暴露后约一小时开始)。 这项提议的目标是描述UVA引起钙反应(AIM)的分子步骤 1)和细胞黑色素浓度的增加(目标2)。使用活细胞荧光成像技术， 电生理学和生化，我们将在Aim1中鉴定由UVA激活的G蛋白途径， 确定细胞信使的特征并确定导致快速和瞬时钙离子的机制 回应。在目标2中，我们将测试UVA介导的细胞去极化是一种 持续的钙离子反应和持续的PKC激活，导致早期黑色素产生。在《目标3》中我们 将测试UVA和UVB信号通路的假设，这两个信号通路同时被太阳激活 UVR，协同作用，调节黑色素的产生和着色。我们将使用鼠标和 人源化皮肤研究UVA和UVB激活途径之间的相互作用 描述调节这种相互作用的分子机制。 拟议的研究将确定和表征一种新的UVA光传导途径，并确定其 在皮肤中的功能，并将大大促进我们对黑素细胞功能和皮肤的了解 对太阳UVR的反应。

PROJECT SUMMARY Skin, the largest organ of the human body, is constantly exposed to solar ultraviolet radiation (UVR), a powerful environmental carcinogen. Unlike the skin of other mammals, human skin responds to UVR by increased pigmentation resulting from melanin production in epidermal melanocytes and its transfer to keratinocytes. Melanin protects the nuclear DNA of melanocytes and keratinocytes by directly absorbing UVR and neutralizing free radicals, thus preventing genetic damage and cancer formation. How does human skin detect and respond to UVR? UVR at the earth's surface consists of 95% long wavelength UVR (UVA) and 5% short wavelength UVR (UVB). Although the mechanism by which UVB causes DNA damage and results in delayed melanin synthesis more than a day after exposure is well characterized, the effects of UVA are poorly understood. High UVA doses cause oxidative damage, but no specific pathway for detecting physiological doses of UVA has been characterized. Our preliminary results describe a novel UVA-activated signaling pathway in melanocytes that is retinal-dependent and G-protein coupled. Activation of this pathway by physiological doses of UVA leads to the activation of transient receptor potential A1 (TRPA1) ion channels, a rapid increase in intracellular Ca2+, and increased melanin content on a rapid time scale (beginning about an hour after UVA exposure). The goal of this proposal is to characterize the molecular steps by which UVA causes a Ca2+ response (Aim 1) and an increase in cellular melanin concentration (Aim 2). Using live-cell fluorescence imaging, electrophysiology, and biochemistry, we will identify in Aim1 the G protein pathway activated by UVA, characterize the cellular messengers and determine the mechanisms that lead to a rapid and transient Ca2+ response. In Aim 2 we will test the hypothesis that UVA-mediated cellular depolarization is required for a sustained Ca2+ response and persistent PKC activation, which leads to early melanin production. In Aim 3 we will test the hypothesis that UVA and UVB signaling pathways, which are simultaneously activated by solar UVR, interact synergistically to regulate melanin production and pigmentation. We will use mice with "humanized" skin to investigate the interaction between the UVA and UVB activated pathways and characterize the molecular mechanisms mediating this interaction. The proposed studies will identify and characterize a novel UVA phototransduction pathway and determine its function in skin, and will significantly advance our understanding of melanocyte function and of the skin's response to solar UVR.