Molecular Basis of Th1 Development

这个项目代表了一项确定Th1发育的分子基础的努力。在基础水平上对Th1发育进行分析的动机理由源于CD4T细胞，特别是Th1细胞，在包括1型糖尿病在内的几种类型的自身免疫性疾病中发挥的核心作用。这里所述的项目基于大量的初步数据，这些数据是根据上一个供资周期取得的进展而得出的。在对IL-12信号通路进行研究的同时，我们认识到Th1的发育首先进入STAT4依赖的阶段，然后进入STAT4非依赖的阶段。后一阶段代表了分化更充分的Th1细胞，在这种细胞中，仅在TCR激活后就可以产生高水平的IFN-γ，而不通过IL-12信号通路发挥作用。重要的是，这一结果直接表明，在Th1发育的这个阶段，STAT4的激活是一个非常弱的信号，可以诱导IFNGamma的产生，而它对TCR激活产生的IFNGamma的贡献很小。因此，我们得出假设，STAT4的作用是导致Th1特异性转录因子的表达，这些转录因子直接促进TCR衍生信号反应中IFNGamma的产生。因此，我们得出一个假设，即STAT4的作用是导致Th1特异性转录因子的表达，然后直接促进TCR衍生信号的产生。我们已经开始使用几种克隆方法来鉴定这种转录因子。我们的初步数据表明，我们已经确定了这样一个转录因子ERM，我们证明了它是由IL-12单独通过激活STAT4来诱导的。我们还开发了一种重要的方法，有助于我们分析Th1的发育和这些转录因子的作用。也就是说，我们现在已经开发出用逆转录病毒非常高效地感染原代T细胞并表达感兴趣的转录因子的技术。我们证明，在STAT4缺陷的T细胞中表达ERM可以显著增加其IFNGamma的产生。我们建议使用既定的方法，以非常直接的方式跟踪这些发现。我们将进行一项分析，以全面描述ERM在Th1发育中的作用，并将继续寻找更多的STAT4诱导因素。此外，我们将分析STAT4激活基因的特异性的基础，并定义T激活与其他统计数据之间的差异，特别是STAT1。最后，我们将重点放在小鼠和人类系统在激活STAT4方面的特定差异上，我们认为这是一个需要解决的重要问题，因为小鼠系统被大量用作人类疾病的模型。

This Project represents an effort to define the molecular basis of Th1 development. The motivating rationale for pursuing an analysis of Th1 development at a fundamental level stems from the central role that CD4 T cells, particularly Th1 cells, play in several types of autoimmune diseases, including the type 1 diabetes. The project described here is based on extensive preliminary data that have resulted fro, progress made in the previous funding cycle. While pursuing studies of the IL-12 signaling pathway, we recognized that Th1 development proceeds to first a Stat4-dependent, and then a Stat4- independent phase. This latter phase represents more fully differentiated Th1 cells in which high IFNgamma production can occur after TCR activation alone, without contribution through the IL-12 signaling pathway. Importantly, this result directly showed that in this phase of Th1 development, Stat4 activation is an extremely weak signal to induce IFNgamma production, and that it contributes very little to the IFNgamma produced by a TCR activation. Therefore, we arrived at the hypothesis that the role of Stat4 was to cause the expression of Th1- specific transcription factors which them directly promote the production of IFNgamma in response to TCR derived signals. Therefore, we arrived at the hypothesis that the role of Stat4 was to cause the expression of Th1- specific transcription factors when then directly promote the production of IFNgamma in response to TCR derived signals. We have set about to identify such transcription factors using several cloning approaches. Our preliminary data show that we have identified one such transcription factor, ERM, which we demonstrate is induced by IL-12, through Stat4 activation alone. We have also developed an important methodology that will facilitate our analysis of Th1 development and the role of these transcription factors. That is, we have now developed the technique to infect primary T cells by retrovirus at very high efficiency and express the transcription factors of interest. We demonstrate that expression of ERM in Stat4-deficient T cells can significantly augment their IFNgamma production. We propose to follow these findings in very direct ways using established methods. We will carry out an analysis to completely characterize the role of ERM in Th1 development, and will continue to search for additional Stat4-induced factors. Furthermore, we will analyze the basis for the specificity of gene activation by Stat4, and define the differences between T activation compared to other STATs, notably Stat1. Finally, we focus on a specific difference between the mouse and human systems in the activation of Stat4, which we argue is an important issue to resolve, since murine systems are used heavily as models for human diseases.