p63 Signaling in Epithelial Cell Growth and Cancer

描述(申请人提供)：该研究的目标是探讨P53家族成员p63在上皮细胞信号和功能中的作用，p63表达于几种复层上皮组织的基底层，包括表皮、口腔粘膜、食道和膀胱，以及几种腺体结构，如支气管、乳腺和前列腺，p63基因缺陷的小鼠和具有p63胚系突变的人类表现出严重的上皮相关缺陷。P63基因编码六个剪接变异体，具有几个预测的生化活性。在成人中，p63亚型deltaNp63α在表达它的组织中的上皮细胞基底层表达最高，在鳞状细胞癌中经常过表达。 基于我们最近发现的DeltaNp63α：(I)可作为转录抑制因子，(Ii)受生长刺激和抑制信号调节，以及(Iii)丢失可导致上皮细胞衰老，我们提出了以下相关假设。DeltaNp63α通过生长刺激和抑制信号的差异磷酸化，并通过选择靶基因的转录调控，在维持表皮储备细胞群的增殖状态中发挥作用，将更多分化的细胞从基质中分离出来。我们将通过分析deltaNp63pha的磷酸化，识别在体内结合和调节deltaNp63pha的新的靶基因，以及确定选定的p63靶基因在上皮细胞自我更新中的作用来检验这些假设。在我们的研究中，我们将主要使用原代、正常的人类表皮角质形成细胞和乳腺肌上皮细胞。将利用新的技术和试剂通过以下具体目标来检验这些假说：(I)确定deltaNp63pha在体内结合和调节的靶基因；(Ii)确定选定的deltaNp63pha靶基因在决定细胞在增殖、衰老、分化和应激条件下的结果中的作用；以及(Iii)分析deltaNp63pha的磷酸化及其在调节deltaNp63pha蛋白水平和活性中的作用。几种人类肿瘤类型中deltaNp63α过表达的频率，p63胚系突变导致的人类发育障碍的发生，以及调控人类癌细胞中的deltaNp63α信号可能导致生长停滞或凋亡的希望，突显了了解deltaNp63α调控和功能的重要性。

DESCRIPTION (provided by applicant): The goal of the proposed studies is to address the role of one p53 family member, p63, in epithelial cell signaling and function, p63 is expressed in the basal layer of several stratified epithelial tissues including the epidermis, oral mucosa, esophagus, and bladder, as well as in several glandular structures such as bronchi, breast and prostate, p63-deficient mice and humans with p63 germline mutations display severe epithelium-related defects. The p63 gene encodes six splice variants with several predicted biochemical activities. In adults, expression of one p63 isoform, deltaNp63alpha is highest in the basal subpopulation of epithelial cells in tissues in which it is expressed and is frequently overexpressed in squamous cell carcinomas. Based on our recent discoveries that deltaNp63alpha: (i) can function as a transcriptional repressor, (ii) is regulated by growth stimulatory and inhibitory signaling, and (iii) loss can lead to epithelial cell senescence, we put forth the following interrelated hypotheses. DeltaNp63alpha is differentially phosphorylated by growth stimulatory and inhibitory signaling, and through transcriptional regulation of select target genes plays a role in maintenance of the proliferative state of epidermal reserve cell populations that separate the more differentiated cells from the stroma. We will test these hypotheses by analyzing deltaNp63alpha phosphorylation, identifying novel target genes that deltaNp63alpha binds and regulates in vivo, and determining the role of select p63 target genes in epithelial cell self-renewal. For our studies, we will predominantly use primary, normal, human epidermal keratinocytes and breast myoepithelial cells. New technologies and reagents will be exploited to test the hypotheses through the following Specific Aims: (i) To identify target genes that deltaNp63alpha binds and regulates in vivo; (ii) To determine the role of select deltaNp63alpha target genes in dictating cellular outcome under conditions of proliferation, senescence, differentiation, and stress; and (iii) To analyze deltaNp63alpha phosphorylation and the role it plays in regulating deltaNp63alpha protein levels and activity. The importance of understanding deltaNp63alpha regulation and function is underscored by the frequency of deltaNp63alpha overexpression in several human tumor types, the occurrence of human developmental disorders with germline mutations in p63, and the hope that modulating deltaNp63alpha signaling in human cancer cells may induce growth arrest or apoptosis.