Growth differentiation control in primary keratinocytes

原代角质形成细胞的生长分化控制

• 批准号: 8096827
• 负责人: GIAN-PAOLO DOTTO
• 金额: $ 54.04万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-27 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8096827
• 关键词: Address; Biological; Biological Assay; Cells; Commit; Communication; Complex; Core-Binding Factor; Development; Differentiation and Growth; Equilibrium; Event; Future; Growth; Human; In Vitro; Interferons; Mus; Outcome; Pathway interactions; Phosphotransferases; Play; Population; ROCK1 gene; Regulation; Regulatory Pathway; Role; Signal Transduction; Skin; Stem cells; System; Testing; Transcriptional Activation; Work; cell type; in vivo; keratinocyte; keratinocyte differentiation; notch protein; reconstitution; research study; response; rho GTP-Binding Proteins; self-renewal; stem cell population; transcription factor; tumor; tumorigenesis

DESCRIPTION (provided by applicant): Notch signaling is an important form of inter-cellular communication with a key role in cell-fate determination and differentiation (Artavanis-Tsakonas et al., 1999). The biological outcome of activation or suppression of this pathway is highly context-dependent. In many mammalian systems, Notch signaling enhances stem cell potential and suppresses differentiation, while in others, notably keratinocytes, it exerts an opposite role. Our main working hypothesis is that this pro-differentiation function of Notch in keratinocytes depends on a cell-type specific integration with other key regulatory pathways. Signaling mechanisms involved in growth/differentiation control of human and mouse keratinocytes, including their Notch response, are only partially overlapping. Therefore, in our future work, we will focus preferentially on human cells, utilizing the mouse system whenever necessary and as a point of reference. Notch signaling proceeds through a "canonical" pathway involving transcriptional activation of the CBF-1/Maml_1 complex and a less defined "non-canonical" pathway independent of CBF-1/Maml_1 function. We will address the following aims : 1) We will test the hypothesis that the "canonical" Notch pathway promotes keratinocyte commitment to differentiation through a cross-talk with signaling by small Rho GTPases. In particular, we have found that the Notch/CBF1 pathway is involved in cell type specific negative regulation of three key effectors of small Rho GTPases : the ROCK1, 2 and MRCKa kinases. Accordingly, we will assess whether down-modulation of these molecules is a key event in the Notch response of keratinocytes, and whether these kinases in turn control Notch signaling in these cells 2) We will test the hypothesis that the "non-canonical" Notch pathway plays a parallel important function in keratinocyte growth/differentiation control through an interplay with Interferon Response Factors and in particular IRF6, a transcription factor with an essential specific role in keratinocytes. We will evaluate the involvement of Notch signaling, in connection with IKKa, another selective regulator of keratinocyte differentiation, in control of IRF6 expression. Concomitantly, we will assess the impact of IRF6 and other IRFs on the Notch-response of keratinocytes, with p63, a key transcription factor for these cells, as an important endpoint. 3) We will test the hypothesis that integration of the above mechanisms plays a key role in long term control of self-renewing stem cell populations, and tumorigenesis. By both in vitro and in vivo assays, we will assess the balance between self-renewing and reversibly versus irreversibly committed populations. Concomitantly, we will determine the role of the above pathways in control of keratinocyte tumor development, by skin reconstitution / grafting experiments with genetically modified keratinocytes.

描述（由申请人提供）：Notch 信号传导是细胞间通讯的重要形式，在细胞命运决定和分化中发挥关键作用（Artavanis-Tsakonas 等，1999）。该通路激活或抑制的生物学结果高度依赖于环境。在许多哺乳动物系统中，Notch 信号传导增强干细胞潜力并抑制分化，而在其他系统中，尤其是角质形成细胞中，它发挥相反的作用。我们的主要工作假设是，Notch 在角质形成细胞中的这种促分化功能取决于细胞类型与其他关键调控途径的特异性整合。人类和小鼠角质形成细胞生长/分化控制中涉及的信号机制（包括它们的 Notch 反应）仅部分重叠。因此，在我们未来的工作中，我们将优先关注人类细胞，并在必要时利用小鼠系统作为参考。 Notch 信号传导通过涉及 CBF-1/Maml_1 复合物转录激活的“规范”途径和独立于 CBF-1/Maml_1 功能的不太明确的“非规范”途径进行。我们将实现以下目标：1）我们将测试“规范”Notch 通路通过与小 Rho GTPases 信号传导的串扰促进角质形成细胞分化的假设。特别是，我们发现 Notch/CBF1 通路参与小 Rho GTPases 的三个关键效应子（ROCK1、2 和 MRCKa 激酶）的细胞类型特异性负调节。因此，我们将评估这些分子的下调是否是角质形成细胞 Notch 反应中的关键事件，以及这些激酶是否反过来控制这些细胞中的 Notch 信号传导 2) 我们将检验“非规范”Notch 的假设途径通过与干扰素反应因子，特别是 IRF6（一种在角质形成细胞中具有重要特定作用的转录因子）相互作用，在角质形成细胞生长/分化控制中发挥平行的重要功能。我们将评估 Notch 信号传导与 IKKa（角质形成细胞分化的另一种选择性调节因子）在 IRF6 表达控制中的关系。与此同时，我们将评估 IRF6 和其他 IRF 对角质形成细胞 Notch 反应的影响，其中 p63（这些细胞的关键转录因子）作为重要终点。 3）我们将检验以下假设：上述机制的整合在自我更新干细胞群的长期控制和肿瘤发生中起着关键作用。通过体外和体内测定，我们将评估自我更新和可逆与不可逆承诺群体之间的平衡。与此同时，我们将通过转基因角质形成细胞的皮肤重建/移植实验来确定上述途径在控制角质形成细胞肿瘤发展中的作用。