Shirakata M, Nabeshima Y, Konishi K, Fujii-Kuriyama Y
Department of Biochemistry, Cancer Institute, Japanese Foundation for Cancer Research, Tokyo.
Mol Cell Biol. 1988 Jun;8(6):2581-8. doi: 10.1128/mcb.8.6.2581-2588.1988.
The expression of the fast type of myosin alkali light chain 1 is induced during the differentiation of muscle cells. To study the mechanism of its gene regulation, we joined the sequence of the 5'-flanking and upstream region of the chicken myosin alkali light-chain gene to the structural gene for chloramphenicol acetyltransferase (CAT). The fusion gene was introduced either into quail myoblasts transformed by a temperature-sensitive mutant of Rous sarcoma virus (tsNY68) or into chicken myoblasts, and the transiently expressed CAT activity was assayed after the differentiation of the myoblasts. From the experiments with the external and internal deletion mutants of the fusion gene, the cis-acting regulatory region responsible for the enhanced expression of the CAT activity in response to the cell differentiation was found to be localized at 2 kilobases upstream of the transcription initiation site. This region of 160 nucleotides contained two pairs of short sequences worthy of note, a direct repeat of 12 nucleotides, and an inverted repeat of 8 nucleotides. The nucleotide sequences of the 5'-flanking sequence up to nucleotide -3381 were determined and compared with those of the upstream activating elements of actin genes.
肌球蛋白碱性轻链1快速型的表达在肌肉细胞分化过程中被诱导。为了研究其基因调控机制，我们将鸡肌球蛋白碱性轻链基因的5'-侧翼及上游区域序列与氯霉素乙酰转移酶（CAT）的结构基因连接。融合基因被导入由劳氏肉瘤病毒温度敏感突变体（tsNY68）转化的鹌鹑成肌细胞或鸡成肌细胞中，成肌细胞分化后测定瞬时表达的CAT活性。通过对融合基因的外部和内部缺失突变体进行实验，发现负责响应细胞分化而增强CAT活性表达的顺式作用调控区域位于转录起始位点上游2千碱基处。这个160个核苷酸的区域包含两对值得注意的短序列，一个12个核苷酸的直接重复序列和一个8个核苷酸的反向重复序列。测定了直至核苷酸-3381的5'-侧翼序列的核苷酸序列，并与肌动蛋白基因的上游激活元件的序列进行了比较。
