ANALYSIS OF TWO PROMOTERS AND 5'FLANKING REGION OF RAT SERINE : PYRUVATE AMINOTRANSFERASE GENE

(1)两个SPT基因启动子的基础转录活性所需的序列单个SPT基因具有两个启动子，上游启动子（转录自+1），其具有TATA盒并且其活性通过cAMP施用和下游启动子增强（从+66转录），其不含TATA盒并且其活性不受cAMP影响。我构建了许多5 '端缺失的重组质粒，其3'端对于上游启动子为+36，对于下游启动子为+106。在含有SPT基因-192 ~+36序列的质粒中，从上游起始位点的转录达到最大水平，这表明除了位于-103 ~+36之间的CCAAT盒和TATA盒外，含有-192 ~-103之间的某些元件是最大活性所必需的。另一方面，检测下游启动子的转录活性， ...更多信息 在含有+36至+106序列的质粒中达到最大水平，在含有-51至+106序列的质粒中达到最大水平。这表明，即使HIP 1（管家起始蛋白1）作为下游启动子起作用，其也是下游启动子的最大转录活性所必需的，HIP 1被认为是无TATA启动子之一，并且位于+11和+106之间。(2)SPT基因的转录调控和细胞特异性表达研究表明，位于-684和-621之间的CRE（cAMP responsive element）与cAMP增强SPT基因的作用无关，而位于-103下游的区域对cAMP增强SPT基因的作用有贡献。仅在HepG 2细胞中观察到这种增强，而在HeLa、CHO-K1、CV-1和COS-1细胞中未观察到这种增强。由于cAMP对SPT基因转录的增强作用被蛋白质合成抑制剂放线菌酮所抑制，因此，CREB（CRE binding protein）可能是先增强了某些转录因子的肝脏特异性表达，然后翻译因子作用于SPT基因的近端区域，导致SPT基因转录增强。少

(1) Sequence required for the basal transcriptional activity of two SPT gene promotersA single SPT gene has two promoters, the upstream promoter (transcribed from+1) which has TATA box and whose activity is enhanced by cAMP administration and the downstream promoter (transcribed from +66) which contains no TATA box and whose activity is not affected by cAMP.To identify the regions necessary for the basal transcription by these two promoters, I constructed many 5'-deleted recombinant plasmids whose 3'-ends are +36 for the upstream promoter or +106 for the downstream promoter. The transcription from the upstream initiation site was reached to a maximum level in a plasmid containing a sequence from -192 to +36 of SPT gane, suggesting that, in addition to CCAAT box and TATA box which are located between -103 and +36 ; some element (s) contained between -192 and -103 is necessary for the maximum activity. The transcriptional activity of the downstream promoter, on the other hand, was detect … More ed in a plasmid having a sequence from +36 to +106 and was reached to a maximum level in a plasmid containing a sequence from -51 to +106. This suggests that some element contained between -51 and +11 are necessary for the maximum transcriptional activity of the downstream promoter even if HIP1 (Housekeeping initiation protein 1), which is proposed as one of TATA-less promoters and is located between +11 and +106, functions as the downstream promoter.(2) Transcriptional regulation and cell-specific expression of SPT geneIt has been indicated that the region downstream from -103 contributes to the enhancement of SPT gene by cAMP rather than CRE (cAMP responsive element) located between -684 and -621. This enhancement was observed only in HepG2 cells, not in HeLa, CHO-K1, CV-1 and COS-1 cells. Because the transcriptional enhancement of SPT gene by cAMP is repressed by cycloheximide, a inhibitor of protein synthesis, it seems that CREB (CRE binding protein) first increases the liver-specific expression of some transcriptional factor and then the translation factor acts on the proximal region of SPT gene to cause the enhancement of SPT gene transcription. Less