Investigating the role of fatty acid metabolism in melanocyte differentiation: Insights into the metabolic requirements of development

项目概要 黑素细胞是皮肤中的关键细胞谱系，是皮肤色素沉着所必需的。黑素细胞缺陷 发育和功能可能导致色素沉着障碍，导致色素沉着不足或过度，以及一种 最致命的皮肤癌——黑色素瘤。许多转录和信号转导事件导致 人们已经对这些疾病进行了探索，但是，对于代谢性疾病的作用却知之甚少。 黑素细胞发育过程中的环境和细胞内代谢状态。我们实验室的最新工作 已经证明，在黑色素瘤进展过程中，肿瘤微环境中的脂肪细胞可以转移 脂肪酸对黑色素细胞有促进侵袭作用。这就提出了脂肪酸摄取是否发挥作用的问题。 在正常黑素细胞中发挥作用。我们的初步数据表明，黑素细胞需要外源性摄取脂肪 通过脂肪酸转运蛋白 (FATP) 进行适当的分化。在目标 1 中，我们将调查 黑素细胞中脂肪酸摄取和脂肪酸从头合成的相对重要性。我们会 比较脂肪酸摄取和脂肪酸合成的多种机制，以确定哪种机制发挥作用 在黑素细胞发育和模式形成中发挥最重要的作用。我们还将在具体阶段进行剖析 当黑素细胞在利用遗传扰动分化期间需要这些途径时，以这种方式进行。 我们假设脂肪酸在黑素细胞中的重要性反映了某些谱系的特定需求， 包括高含量的脂肪酸以支持磷脂合成和树突形成。在目标 2 中，我们将 检查脂肪酸作为磷脂合成的基本成分相对于 通过 b-氧化分解脂肪酸的需要。有证据表明其重要性 其他树突状细胞类型中的磷脂合成。因为黑素细胞也形成广泛的树突， 对于适当的色素沉着至关重要，我们将剖析主要磷脂合成途径对色素沉着的重要性 黑素细胞以阶段特定的方式。相反，黑素细胞可能会分解脂肪酸作为 ATP 支持细胞增殖和迁移。我们还将确定 b-氧化在黑素细胞中的作用 通过以阶段特定的方式针对该途径中的关键蛋白质来进行开发。我们将评估以下措施的影响 这些基因扰动对能量产生和黑素细胞发育的影响。为了进行这些研究，我们 将主要依靠斑马鱼作为黑素细胞发育的体内模型。我们将使用阶段特定的 神经嵴、成黑素细胞和黑素细胞的启动子，以产生稳定的种系转基因。使用 结合遗传学方法和图像分析，我们将研究脂肪酸和脂质代谢的作用 在黑色素细胞发育中。在适当的时候，我们还将应用黑素细胞发育的体外模型， 基于黑素细胞与人类多能干细胞的分化，这些组合实验将 导致关于代谢在黑素细胞发育中的作用的新发现，这可能会增加我们的 了解和治疗色素沉着疾病。

PROJECT SUMMARY Melanocytes are a key cell lineage in the skin and are required for skin pigmentation. Defects in melanocyte development and function can lead to pigmentation disorders causing hypo or hyperpigmentation as well as one of the most deadly forms of skin cancer, melanoma. Many of the transcriptional and signaling events which cause these diseases have been explored, however, there has been little investigation into the role of the metabolic environment and intracellular metabolic state of the melanocyte during development. Recent work from our lab has demonstrated that during melanoma progression, adipocytes in the tumor microenvironment can transfer fatty acids to the melanoma cells, promoting invasion. This raises the question of whether fatty acid uptake plays a role in normal melanocytes. Our preliminary data suggests that melanocytes require extrinsic uptake of fatty acids through Fatty Acid Transport Proteins (FATPs) for proper differentiation. In Aim 1, we will investigate the relative importance of fatty acid uptake and de novo fatty acid synthesis in the melanocyte. We will compare multiple mechanisms of fatty acid uptake as well as fatty acid synthesis to determine which play the most important role in melanocyte development and pattern formation. We will also dissect in a stage specific manner when these pathways are required by the melanocyte during differentiation using genetic perturbations. We hypothesize that the importance of fatty acids in the melanocyte reflects certain lineage specific needs, including high levels of fatty acids to support phospholipid synthesis and dendrite formation. In Aim 2 we will examine the importance of fatty acids as building blocks for phospholipid synthesis relative to a requirement for fatty acid breakdown through b-oxidation. There is evidence for the importance of phospholipid synthesis in other dendritic cell types. Because melanocytes also form extensive dendrites which are critical for proper pigmentation we will dissect the importance major phospholipid synthesis pathways to the melanocyte in a stage specific manner. Conversely, melanocytes might break down fatty acids as a source of ATP to support cell proliferation and migration. We will also determine the role b-oxidation in melanocyte development by targeting key proteins in this pathway in a stage specific manner. We will assess the effects of these genetic perturbations on energy production and melanocyte development. To perform these studies we will primarily rely on zebrafish as an in vivo model of melanocyte development. We will use stage specific promoters for the neural crest, melanoblast, and melanocyte to generate stable germline transgenics. Using a combination of genetic methods and image analysis we will investigate the role of fatty acid and lipid metabolism in melanocyte development. When appropriate, we will also apply an in vitro model of melanocyte development, based on the differentiation of melanocytes from human pluripotent stem cells These combined experiments will lead to novel discoveries regarding the role of metabolism in melanocyte development, which could increase our understanding and treatment of pigmentation diseases.