Mori K, Fujii R, Kida N, Takahashi H, Ohkubo S, Fujino M, Ohta M, Hayashi K
Department of Pharmaceutics, Gifu Pharmaceutical University.
J Biochem. 1990 Jan;107(1):73-6. doi: 10.1093/oxfordjournals.jbchem.a123014.
pS2 is a human gene whose transcription is directly triggered by estrogen in human breast cancer cells (MCF-7). We described here the complete primary structure of the pS2 gene product. The pS2 protein purified from conditioned medium of MCF-7 cells was S-pyridylethylated and digested with TPCK-trypsin. Five major fragments were obtained by reverse-phase HPLC. Amino acid sequence analysis of these tryptic peptides established that the pS2 protein comprises a 60-amino acid polypeptide. The sequence of the pS2 protein was completely identical to that deduced from the nucleotide sequence of the pS2 gene, if the signal polypeptide is excluded. Furthermore, two cDNA clones encoding an 84-amino acid precursor pS2 protein were isolated from a cDNA library which was constructed with RNA from MCF-7 cells cultured in the presence of estrogen. The nucleotide sequence of one clone (pS2B1) was identical to that of pS2 cDNA previously reported except for one nucleotide in the 3' untranslated region. The other clone (pS2B2) was longer by 73 nucleotides, at the 5' end, than pS2B1. The additional 73 nucleotides are located just upstream of the sequence of pS2B1 in the structure of the pS2 gene, indicating that the pS2 gene has two start sites for transcription. However, a mRNA molecule corresponding to pS2B1 but not to pS2B2 was detected in the cells on RNA blot hybridization analysis, indicating that one transcriptional start site is mainly used.
pS2是一种人类基因，其转录在人乳腺癌细胞（MCF-7）中由雌激素直接触发。我们在此描述了pS2基因产物的完整一级结构。从MCF-7细胞的条件培养基中纯化的pS2蛋白经S-吡啶基乙基化后用TPCK-胰蛋白酶消化。通过反相高效液相色谱法获得了五个主要片段。对这些胰蛋白酶肽段的氨基酸序列分析确定pS2蛋白由一个60个氨基酸的多肽组成。如果排除信号多肽，pS2蛋白的序列与从pS2基因的核苷酸序列推导的序列完全相同。此外，从一个用在雌激素存在下培养的MCF-7细胞的RNA构建的cDNA文库中分离出两个编码84个氨基酸的前体pS2蛋白的cDNA克隆。一个克隆（pS2B1）的核苷酸序列与先前报道的pS2 cDNA的序列相同，只是在3'非翻译区有一个核苷酸不同。另一个克隆（pS2B2）在5'端比pS2B1长73个核苷酸。在pS2基因结构中，额外的73个核苷酸位于pS2B1序列的上游，表明pS2基因有两个转录起始位点。然而，在RNA印迹杂交分析中，在细胞中检测到了与pS2B1对应的mRNA分子，而未检测到与pS2B2对应的mRNA分子，这表明主要使用一个转录起始位点。
