The regulation of melanocyte stem cells by Wnt signaling

描述（由申请人提供）：黑素细胞干细胞（McSC）在整个生命周期中维持黑素细胞的健康群体。了解它们的行为和调节与黑素细胞损失引起的疾病有关，如炎症后色素减退、白癜风和头发变白，以及黑色素瘤的生物学。目前，对支配MCSC的细胞和分子机制知之甚少。这项资助的目的是了解Wnt信号传导的功能及其在毛囊中的McSCs中的调节。在毛囊的周期性再生过程中，McSC与毛囊上皮干细胞（EpSC）协同激活。它们为毛球提供分化的黑素细胞，用于产生毛发色素。我们发现，在生长期开始时，McSC在邻近EpSC分泌Wnt配体后激活Wnt信号传导。通过转基因删除或激活2-catenin，在McSCs中的Wnt信号的关键介质，我们已经表明，Wnt信号是至关重要的McSC分化。我们假设Wnt信号传导是McSC分化所必需的，并且在毛囊周期期间，EpSC协调调节McSC和EpSC自身中的Wnt信号传导。为了解决这一假设，我们将追求以下具体目标。在Aim 1中，我们将通过在黑素细胞中表达Wnt信号传导的分泌抑制剂Dkk 1或Wnt信号传导的核介质Lef 1的显性阴性形式来抑制McSC中的Wnt信号传导。我们将研究这是如何影响他们的能力，产生功能性黑色素细胞在头发周期。在Aim 2中，为了确定EpSC分泌的Wnt配体是否可以激活邻近McSC中的Wnt信号传导并促进其分化，我们将使用遗传小鼠模型在滤泡EpSC中删除Wnt配体分泌所必需的Wntless基因或迫使EpSC表达Wnt配体。在Aim 3中，我们将测试Wnt抑制剂是否可以促进衰老过程中的McSC维持，这是一个已知导致McSC衰竭和异常分化后损失的过程。具体来说，我们将利用转基因小鼠模型在McSCs中过表达Dkk 1或显性阴性形式的Lef 1，以检查这是否促进了未分化的McSCs在衰老过程中的维持。我们还将测试Wnt抑制是否可以逆转McSC分化，这一过程预示着它们随后的损失。我们的研究将为我们提供一个新的理解的分子机制调节MCSC和黑素细胞的行为是如何与头发再生协调，提供一个科学的基础，了解疾病，其中黑素细胞是不适当的损失，如头发变灰和白癜风或经历不受控制的细胞生长，如黑色素瘤。 公共卫生相关性：Wnt信号在黑素细胞干细胞中的功能和调节目前尚不清楚。我们建议调查的假设，Wnt信号调节黑素细胞干细胞的维持和分化，滤泡上皮干细胞是黑素细胞Wnt信号的关键介质。这项研究将促进我们对调节黑素细胞干细胞的分子信号的理解，并最终有助于开发新的策略来治疗黑素细胞不适当丢失的疾病，如头发变白和白癜风，或经历不受控制的细胞生长，如黑色素瘤。

DESCRIPTION (provided by applicant): Melanocyte stem cells (McSCs) sustain a healthy population of melanocytes throughout life. Understanding their behavior and regulation is relevant to disorders resulting from melanocyte loss such as post inflammatory hypopigmentation, vitiligo, and hair graying, as well as to the biology of melanoma. Currently, little is known about the cellular and molecular mechanisms that govern McSCs. The goal of this grant is to understand the function of Wnt signaling and its regulation in McSCs residing in the hair follicle. McSCs are coordinately activated with follicular epithelial stem cells (EpSCs) during the cyclical regeneration of the hair follicle. They provide differentiated melanocytes to the hair bulb for the production of hair pigment. We discovered that McSCs activate Wnt signaling following Wnt ligand secretion from adjacent EpSCs at the onset of anagen. By transgenically deleting or activating 2-catenin, a key mediator of Wnt signaling in McSCs, we have shown that Wnt signaling is crucial for McSC differentiation. We hypothesize that Wnt signaling is required for McSC differentiation and that EpSCs coordinately regulate Wnt signaling in McSCs and EpSCs themselves during the hair follicle cycle. To address this hypothesis, we will pursue the following specific aims. In Aim1, we will inhibit Wnt signaling in McSCs by expressing either a secreted inhibitor of Wnt signaling, Dkk1, or a dominant- negative form of a nuclear mediator of Wnt signaling, Lef1, in melanocytes. We will examine how this influences their ability to give rise to functional melanocytes during the hair cycle. In Aim2, to determine if Wnt ligand(s) secreted by EpSCs can activate Wnt signaling in neighboring McSCs and promote their differentiation, we will delete the Wntless gene, which is necessary for Wnt ligand secretion, in follicular EpSCs or force expression of a Wnt ligand by EpSCs, using genetic mouse models. In Aim3, we will test if Wnt inhibitors can promote McSC maintenance during aging, a process known to result in McSC exhaustion and loss following their aberrant differentiation. Specifically, we will utilize transgenic mouse models to overexpress either Dkk1 or a dominant-negative form of Lef1 in McSCs to examine if this promotes the maintenance of undifferentiated McSCs during aging. We will also test if Wnt inhibition can reverse McSC differentiation, a process that presages their ensuing loss. Our study will provide us with a novel understanding of the molecular mechanisms regulating McSCs and how melanocyte behavior is coordinated with hair regeneration, providing a scientific foundation for understanding diseases in which melanocytes are either inappropriately lost such as hair graying and vitiligo or undergo uncontrolled cell growth as in melanoma. PUBLIC HEALTH RELEVANCE: The function and regulation of Wnt signaling in melanocyte stem cells is currently unknown. We propose to investigate the hypothesis that Wnt signaling regulates the maintenance and differentiation of melanocyte stem cells and that follicular epithelial stem cells are critical mediators of melanocyte Wnt signaling. This study will advance our understanding of the molecular signals that regulate melanocyte stem cells and will ultimately help development of novel strategies to treat diseases in which melanocytes are either inappropriately lost such as hair graying and vitiligo or undergo uncontrolled cell growth as in melanoma.