CYTOKINE MODULATION OF COLLAGEN GENE EXPRESSION: SIGNAL

I型胶原蛋白的不受控制的积累，硬皮病的标志，导致皮肤和其他受影响器官的纤维化。该过程归因于硬皮病成纤维细胞中胶原基因转录的组成性激活。转化生长因子-β（TGF-β），一种用于胶原合成的有效刺激物，强烈地涉及病理性纤维形成，而干扰素γ（IFN-γ）拮抗TGF-β的作用并且对于预防瘢痕形成是重要的。最近，SMAD和STAT 1已被确定为TGF-β和IFN-γ的细胞内信号转导子。然而，这些细胞因子调节胶原基因转录的途径以及所涉及的转录机制仍然知之甚少。我们的实验室已经确定了SMAD 3在TGF-β刺激胶原基因调节I型胶原基因转录中的作用，并描绘了导致其在硬皮病中组成性上调的改变。为此，基于我们实验室最近关于成纤维细胞中TGF-β和IFN-γ信号传导以及p300/CBP在这些通路中的作用的见解，我建议检查这些共激活因子参与刺激和抑制胶原蛋白转录的假设，并将拮抗信号传导整合到Type 1胶原蛋白基因启动子中。本研究将从以下三个方面对这一假说进行验证：1）阐明p300/CBP参与TGF-β激活正常成纤维细胞中I型胶原基因转录的过程; 2）分析TGF-β和IFN-γ拮抗调节这些细胞中胶原基因转录的分子机制;和3）检测硬皮病成纤维细胞中SMAD-p300/CBP共激活因子的相互作用以及胶原基因转录的组成性上调。本申请中描述的试点研究是基于最近在理解TGF-β信号传导和共激活因子在转录调控中的作用方面的突破。通过增强我们对成纤维细胞中胶原基因转录如何调节的了解，这些研究最终可能导致设计新的治疗策略来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.