Lei K J, Gluzman Y, Pan C J, Chou J Y
Human Genetics Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
Mol Endocrinol. 1992 May;6(5):703-12. doi: 10.1210/mend.6.5.1318503.
Human pregnancy-specific glycoproteins (PSGs) are a family of closely related placental proteins that, together with the carcinoembryonic antigen members, comprise a subfamily within the immunoglobulin superfamily. To facilitate study of the control of PSG expression, we immortalized human placental cell lines with adenovirus-origin-minus (ori-)-simian virus-40 (SV40) recombinant viruses containing either wild-type or temperature-sensitive (ts) A mutants of SV40. Cells transformed with the SV40 tsA chimera (HP-A1 and HP-A2), but not the SV40 wild-type chimera (HP-W1), were temperature sensitive for transformation. All three cell lines expressed trophoblast-specific genes, including PSG and the alpha- and beta-subunits of hCG. Human CG alpha expression was greatly stimulated by (Bu)2cAMP in all three cell lines; shifting HP-A1 and HP-A2 cells to the nonpermissive temperature (39.5 C) further increased hCG alpha expression. At both 33 C (permissive temperature) and 39.5 C, the transformed placental cells expressed PSG mRNAs of 2.2 and 1.7 kilobases; expression was greatly stimulated by sodium butyrate. In the absence of an inducer, the three placental lines synthesized a PSG of 64 kilodaltons (kDa). In the presence of butyrate, they synthesized PSGs of 72, 64, and 54 kDa, similar to the placental PSGs. However, in placenta the predominant species is the 72-kDa product. At 39.5 C, butyrate selectively increased synthesis of the 72-kDa PSG in HP-A1 and HP-A2 cells. To characterize PSG promoter activity, we constructed chloramphenicol acetyltransferase (CAT) fusion genes containing -809 to -44 basepairs up-stream of the translational start site of the PSG6 gene. Using transient expression assays, we demonstrated that the -809/-44 region of the PSG6 gene contained cis-acting sequences that can direct CAT expression in human placental cells. Sodium butyrate, which stimulates PSG expression, greatly increased CAT activity, indicating that butyrate-induced PSG expression is regulated primarily at the level of gene transcription.
人妊娠特异性糖蛋白（PSG）是一族密切相关的胎盘蛋白，它们与癌胚抗原成员一起，构成免疫球蛋白超家族中的一个亚家族。为便于研究PSG表达的调控，我们用含野生型或温度敏感（ts）A突变体猿猴病毒40（SV40）的腺病毒缺失起源（ori -）-SV40重组病毒使人胎盘细胞系永生化。用SV40 tsA嵌合体（HP - A1和HP - A2）而非SV40野生型嵌合体（HP - W1）转化的细胞对转化具有温度敏感性。所有这三种细胞系均表达滋养层特异性基因，包括PSG以及hCG的α和β亚基。在所有这三种细胞系中，（Bu）2cAMP极大地刺激了人CGα的表达；将HP - A1和HP - A2细胞转移至非允许温度（39.5℃）进一步增加了hCGα的表达。在33℃（允许温度）和39.5℃时，转化的胎盘细胞均表达2.2和1.7千碱基的PSG mRNA；丁酸钠极大地刺激了其表达。在没有诱导剂的情况下，这三种胎盘细胞系合成了一种64千道尔顿（kDa）的PSG。在丁酸钠存在的情况下，它们合成了72、64和54 kDa的PSG，类似于胎盘PSG。然而，在胎盘中主要的种类是72 kDa的产物。在39.5℃时，丁酸钠选择性地增加了HP - A1和HP - A2细胞中72 kDa PSG的合成。为了表征PSG启动子活性，我们构建了氯霉素乙酰转移酶（CAT）融合基因，其包含PSG6基因翻译起始位点上游-809至-44碱基对。使用瞬时表达分析，我们证明PSG6基因的-809 / -44区域包含可在人胎盘细胞中指导CAT表达的顺式作用序列。刺激PSG表达的丁酸钠极大地增加了CAT活性，表明丁酸钠诱导的PSG表达主要在基因转录水平受到调控。
