L人晶体膜内蛋白MP19基因(LIM2)启动子上一个晶体特异顺式元件的鉴定(英文)

Identification of a Lens-specific cis-acting Element within the Basal Promoter of the Human Lens Intrinsic Membrane Protein MP19 Gene(LIM2)

使用重叠和变异的寡核苷酸作为探针 ,凝胶迁移分析和竞争实验分析了LIM2转录起始位点上游 - 4 7至- 32的区域 ,与其高度亲和结合的一个蛋白复合体看来仅仅结合到这个DNA双链区域的“敏感”位点。这个位点的序列由 4个G核苷 ,接着 7个其他核苷酸 (AACCTAA)及连着另外 4个G核苷组成 ,即GGGGAACCTAAGGGG ;我们称其为Hsu元件。使用含有这个元件或相应的变异元件所构建的LIM2基因启动子 CAT质粒的活性分析表明Hsu元件是位于LIM2基因启动子之内 ,它是LIM2基因表达所必须的。结合到Hsu元件的反式因子存于晶体发育期间 ,看来是晶体特异性的。由于LIM2基因启动子并不包含一个经典的TATA盒 ,这个Hsu元件可能充当RNA复制酶复合体结合的位点。

Using overlapping and mutant oligonucleotides as probes, gel mobility assays and competition experiments identified a sequence from -47 to -32 bp upstream of the LIM2 CAP site, which a lens protein complex bound with high affinity which appeared to bind only to the “sense” strand of the double stranded DNA molecule. This sequence consisted of a string of four guanine residues followed by seven other nucleotides (AACCTAA) and followed by another four guanines, i.e. GGGGAACCTAAGGGG, called the Hsu element. Promoter CAT constructs containing this sequence or mutations of the sequence indicated that the Hsu element is located within the basal promoter, and is essential for expression of the LIM2 gene. The trans factors binding to the Hsu element are present throughout development, and appear to be lens specific. Since the LIM2 gene promoter does not contain a classic TATA box, the Hsu element may serve as the site for binding the RNA polymerase complex.