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

项目总结 黑素细胞是皮肤中的关键细胞系，是皮肤色素沉着所必需的。黑素细胞缺陷 发育和功能可导致色素沉着障碍，引起色素减少或过度色素沉着 最致命的皮肤癌之一，黑色素瘤。许多转录和信号事件会导致 这些疾病已经被探索过，然而，关于代谢的作用的研究很少。 黑素细胞发育过程中的环境和细胞内代谢状态。我们实验室最近的工作 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 脂肪酸到黑色素瘤细胞，促进侵袭。这提出了一个问题，即脂肪酸摄取是否起到了作用 在正常的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。 通过脂肪酸运输蛋白(FATPs)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，以确定哪种机制起作用 在黑素细胞发育和图案形成中最重要的作用。我们还将在一个特定的阶段中剖析 当黑素细胞在使用遗传扰动的分化过程中需要这些途径时，这些途径是如何进行的。 我们假设，黑素细胞中脂肪酸的重要性反映了某些谱系的特定需求， 包括高水平的脂肪酸，以支持磷脂的合成和树枝的形成。在《目标2》中我们将 检查作为磷脂合成基础的脂肪酸的重要性相对于 通过b-氧化分解脂肪酸的要求。有证据表明...的重要性 其他类型树突状细胞的磷脂合成。因为黑素细胞也会形成广泛的树突 对于适当的色素沉着至关重要，我们将剖析主要的磷脂合成途径对 黑素细胞以一种特定阶段的方式。相反，黑素细胞可能会分解脂肪酸，将其作为 支持细胞增殖和迁移的ATP。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靶向这一途径中的关键蛋白来进行发育。我们将评估这些措施的影响 这些对能量产生和黑素细胞发育的遗传扰动。为了进行这些研究，我们 将主要依靠斑马鱼作为体内黑素细胞发育的模型。我们将使用特定于阶段的 神经脊、黑素母细胞和黑素细胞的促进剂，以产生稳定的生殖系转基因。使用 结合遗传方法和图像分析，我们将研究脂肪酸和脂类代谢的作用 在黑素细胞发育过程中。在适当的时候，我们还将应用黑素细胞发育的体外模型， 基于人类多能干细胞对黑素细胞的分化，这些联合实验将 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 对色素沉着性疾病的认识和治疗。

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.