Analysis of transcriptional control mechanism for the expression of type I collagen and related gene in fibrotic skin disorders

系统性硬化症（SSc）等纤维化性皮肤病的特点是皮肤和各种内脏器官中胶原蛋白的过度积累。胶原蛋白的积累被认为是纤维母细胞中胶原蛋白转录增强的结果。I型胶原蛋白是真皮中含量最多的蛋白，由两条α1(I)链和一条α2(I)链协调表达组成。Co1F1是一种特异性结合α2(I)胶原启动子片段的DNA结合蛋白，位于α2(I)胶原基因转录起始上游-400bp处，可激活α2(I)胶原基因的转录。我们对α2(T)胶原基因转录因子多肽的部分序列和cDNA克隆进行了研究。经序列特异性DNA亲和层析纯化的蛋白由42kDa和40.5 kDa的多肽组成。从42kDa多肽中提取的肽片段序列与Purα相同，Purα是一种与人类c-myc基因上游区域结合的核蛋白。40kDa多肽的部分片段与Purβ相同，已部分测序，与Purα具有较强的同源性。对Purβ cDNA进行全长测序。Purα和Purβ的演绎氨基酸序列同源性为61.4%。对于SSc患者，没有一种治疗方式能带来令人满意的改善。因此，我们想要展示红霉素类似物EM703对正常和硬皮病成纤维细胞胶原转录的影响。EM703以剂量依赖性的方式降低正常成纤维细胞的胶原生成（最大减少40%）和α1(I)胶原mRNA水平（最大减少30%）。EM703显著降低了所有9种SSc成纤维细胞。在荧光素酶实验中，COL1A1转录下调30%。EM703处理的成纤维细胞核提取物降低了ccaat结合因子（CBF）的结合活性，而SP1的结合活性不变。这些结果表明，EM703不仅在正常情况下，而且在SSc成纤维细胞中也能减少I型胶原的转录，并且CBF参与了这种减少的表达。少

Fibrotic skin diseases such as systemic sclerosis(SSc) are characterized by excessive accumulation of collagen in the skin and a variety of internal organs. The accumulation of collagen is thought to result from enhanced transcription of collagen in fibroblasts. Collagen type I, a most abundant protein in the dermis, consists of two α1(I) chain and one α2(I) chain which are coordinately expressed. Co1F1, a DNA binding protein which specifically binds to a segment of the α 2(I)collagen promoter at -400bp upstream of the start of transcription, activates transcription of the α2(I)collagen gene in vitro. We investigated on the partial sequences of polypeptide and the cDNA cloning of this transcriptional factor of the α2(T)collagen gene. The protein purified by sequence-specific DNA affinity chromatography were found to consist of 42kDa and 40.5 kDa polypeptides. Sequences of peptide fragments from 42kDa polypeptide were identical to Purα, which is a nuclear protein which has been reported … More to binds to a upstream region of human c-myc gene. Some of those from 40kDa polypeptide were identical to Purβ, has been partially sequenced and has regions of strong homology to Purα. Full length of Purβ cDNA were sequenced. The deductive amino acid sequences of Purα and Purβ showed 61.4% homology. None of treatment modalities for patients with SSc has brought satisfactory improvement. We, therefore, would like to present effects of the Erythromycin analog EM703 on collagen transcription in normal and scleroderma fibroblasts. EM703 reduced collagen production(to 40% in maximum) and mRNA levels of α1(I) collagen(to 30% in maximum) in a dose dependent manner in normal fibroblasts. EM703 markedly reduced those in all 9 SSc fibroblasts. COL1A1 transcription was down-regulated by 30% in the Luciferase assay. Nuclear extracts from fibroblasts treated with EM703 reduced the CCAAT-binding factor(CBF) binding activity, whereas SP1 binding activity was unchanged. These results suggest that EM703 reduce the type I collagen transcription not only in normal but also in SSc fibroblasts and that the CBF is involved in this reduced expression. Less