Function of albinism gene oca2 in non-melanocyte cell development

项目摘要 眼皮肤白化病2（oca2）基因的功能丧失突变不仅导致 黑色素（黑色素）合成的缺陷对肤色很重要，但也会增加 发展"副作用"或"多效性"的可能性。多效性效应的一些例子 包括感觉或神经系统发育缺陷以及胚胎死亡 这取决于突变基因的身份。虽然多效性效应通常 在患有色素减退症如Waardenburg综合征的人中观察到， 白化病的潜在机制是知之甚少，主要是由于低样本量。 因为色素细胞很容易通过透明的皮肤看到，卵子可以收集在 我们建议使用斑马鱼的银细胞来更好地了解 多效性效应出现。当oca2发生突变时，斑马鱼幼虫继续进行正确的 黑色素细胞、黑色素细胞数量减少，但黑色素合成严重减少。 相反，银色素细胞不表达OCA 2，但在不同的阶段数量增加， 符合神经嵴的规格使用oca2突变体虹膜作为模型，我们将 解决以下R03具体目标：1）确定虹膜发育阶段 受OCA 2功能的影响。通过aim 1实验，我们将重新测试oca2在虹膜细胞中的作用。 在多个附加阶段，包括 增殖、分化和存活-可能影响细胞数量的所有细胞事件; 2） 确定oca2是否直接或间接调节虹膜细胞数量。使用我们独特的收藏 黑素细胞突变体，我们将确定OCA 2是否直接（细胞自主）或间接（非- 细胞自主）调节虹膜细胞的发育。在其他实验中，我们将 进行中试[批量和单细胞RNA-Seq]分析，以确定这些方法是否可以 提供了深入了解oca2的自主与非自主的作用， 发展具体来说，这些RNA-seq数据将被分析，以确定是否改变 在表达的内在或外在基因的重要虹膜发展发生与 OCA2功能丧失。一旦这些实验完成，我们将有一个特点， 用于测试oca 2在多效性效应中的功能的模型和大量的初步研究 用于制定R01水平研究假设的数据，旨在阐明 黑色素细胞/黑色素与听觉/视觉/皮肤系统之间的机械联系 人类的发展。

Project summary Loss of function mutations in the oculocutaneous albinism 2 (oca2) gene not only lead to defects in black pigment (melanin) synthesis important for skin color, but also increase the chances of developing “side” or “pleiotropic” effects. Some examples of pleiotropic effects include deficiencies in sensory or neural system development, as well as embryonic death depending on the identity of the mutated gene. Although pleiotropic effects are commonly observed in humans with hypopigmentation disorders like Waardenburg Syndrome and albinism, the underlying mechanisms are poorly understood primarily due to low sample size. Because pigment cells are easily viewed through transparent skin and eggs can be collected in high numbers year round, we propose to use zebrafish silver cells to better understand how pleiotropic effects arise. When oca2 is mutated, zebrafish larvae continue to make the correct number of black pigment cells, melanocytes, but melanin synthesis is severely decreased. Conversely, silver pigment cells do not express oca2, but are increased in number at stages following specification from neural crest. Using oca2 mutant iridophores as a model, we will address the following R03 specific aims: 1) Determine the iridophore developmental stages impacted by oca2 function. With aim 1 experiments, we will retest the role of oca2 in iridophore specification and examine iridophore development at multiple, additional stages including proliferation, differentiation and survival – all cellular events that could impact cell number; 2) Determine if oca2 directly or indirectly regulates iridophore number. Using our unique collection of melanocyte mutants, we will determine if oca2 directly (cell-autonomously) or indirectly (non- cell autonomously) regulates iridophore development. In additional experiments, we will conduct pilot [bulk and single cell RNA-Seq] analysis to determine if these methods can provide insight into oca2’s autonomous versus non-autonomous role during iridophore development. Specifically, these RNA-seq data will be analyzed to determine whether changes in the expression of intrinsic or extrinsic genes important for iridophore development occur with oca2 loss of function. Once these experiments are complete, we will have a characterized model for testing oca2 function in pleiotropic effects and a substantial amount of preliminary data for formulating hypotheses appropriate for R01 level research aimed at elucidating mechanistic connections between melanocytes/melanin and hearing/visual/skin system development in humans.