CYTOKINE MODULATION OF COLLAGEN GENE EXPRESSION: SIGNAL

硬皮病的特征是I型胶原的不受控制的积聚，导致皮肤和其他受影响器官的纤维化。这一过程归因于硬皮病成纤维细胞中胶原基因转录的结构性激活，转化生长因子-β(转化生长因子-β)是胶原合成的有力刺激因素，与病理纤维化密切相关，而干扰素-γ(干扰素-γ)可拮抗转化生长因子-β的作用，对预防瘢痕形成具有重要作用。最近，SMAD和STAT1分别被确定为转化生长因子-β和干扰素-γ的细胞内信号转导。然而，调节胶原基因转录以响应这些细胞因子的途径以及涉及的转录机制仍然知之甚少。我们实验室已经确定了Smad3在转化生长因子-β刺激胶原基因调控I型胶原基因转录中的作用，并描述了导致其在硬皮病中结构性上调的变化。为此，基于我们实验室关于成纤维细胞中转化生长因子-β和干扰素-γ信号转导的最新研究，以及p300/CBP在这些途径中的作用，我建议验证这样一种假设，即这些共激活因子参与了胶原转录的刺激和抑制，并将拮抗信号整合到Typ1胶原基因启动子中。这一假说将在以下三个特定目标中得到验证：1)阐明p300/CBP参与转化生长因子-β激活I型胶原基因转录的作用；2)剖析转化生长因子-β和干扰素-γ拮抗调节硬皮病成纤维细胞中胶原基因转录的分子机制；以及3)研究硬皮病成纤维细胞中Smad-p300/CBP共激活子的相互作用以及胶原基因转录的结构性上调。本申请中描述的先导性研究是基于最近在理解转化生长因子-β信号和共激活因子在转录调控中的作用方面的突破。通过加强我们对成纤维细胞中胶原基因转录调控的了解，这些研究最终可能导致设计新的治疗策略，选择性地调节硬皮病的这一过程。

Un controlled accumulation of Type I collagen, the hallmark of scleroderma, results in fibrosis of skin and other affected organs. This process is attributed to constitutive activation of collagen gene transcription in scleroderma fibroblasts, Transforming growth factor-beta (TGF-beta), a potent stimulus for collagen synthesis, is strongly implicated in pathological fibrogenesis , whereas interferon gamma (IFN- gamma) antagonize the effects of TGF-beta and as important for prevention of scarring. Recently, SMAD and STAT1 have been identified as intracellular signal transducers of TGF-beta and IFN-gamma, respectively. However, the pathways for modulating collagen gene transcription in response to these cytokines, and the transcriptional mechanisms involved, remain poorly understood. Our laboratory has established the role of SMAD3 in TGF-beta stimulation of collagen gene regulation of Type I collagen gene transcription, and to delineate alterations that result in its constitutive up-regulation in scleroderma. To this end, building on recent insights from our laboratory relating to TGF- beta and IFN-gamma signaling in fibroblasts and the role of p300/CBP in these pathways, I propose to examine the hypothesis that these co- activators are involved in stimulation as well as inhibition of collagen transcription, and integrate antagonistic signaling to the Typ1 collagen gene promoters. The hypothesis will be tested in the following three Specific Aims: 1) to elucidate the involvement of p300/CBP in activation of Type I collagen transcription by TGF-beta in normal fibroblasts; 2) to dissect the molecular mechanisms underlying antagonistic regulation of collagen gene transcription in these cells by TGF-beta and IFN-gamma; and 3) to examine SMAD-p300/CBP co-activator interactions in scleroderma fibroblasts with constitutive up-regulation of collagen gene transcription. The pilot studies described in this application are based on recent breakthroughs in understanding TGF-beta signaling and the role of co-activators in transcriptional regulation. By enhancing our knowledge of how transcription of collagen genes is regulated in fibroblasts, these studies could ultimately lead to the design of novel therapeutic strategies to selectively modulate this process in Scleroderma.