| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1994: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1993: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Research Abstract |
(1) Sequence required for the basal transcriptional activity of two SPT gene promoters
A 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
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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 gene
It 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
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