| Budget Amount *help |
¥3,890,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥390,000)
Fiscal Year 2007: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2006: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
In addition to brain, pituitary adenylate cyclase-activating polypeptide (PACAP) is significantly localized also in testis and is expressed in a stage-specific manner, suggesting it's involvement in testicular function including spermatogenesis. Recently, it was reported that the testis-specific exon is involved in the expression of the testicular variant mRNA of rat PACAP gene (PB Daniel, et. al., 2000). However the detailed testis-specific regulatory mechanism of PACAP gene expression remains unknown. Therefore we attempted to characterize human testis specific promoter. We first surveyed the human genome data base, and found a human homologue of testis-specific exon of PACAP gene (hTE), which is localized 10.9 kb upstream from the transcriptional start site. RT-PCR analysis of the hTE with human testicular mRNA revealed the presence of testis-specific variant of PACAP mRNA. The 5'-flanking region of the contains potential binding sites for several transcription factors (SRY, OCT-1, GATA-1, etc.). It should be interesting that the luciferase reporter assay with the 1.2 kb of the 5'-flanking region of the demonstrated the potent promoter activity in the F9 (mouse testicular teratoma cells) and CHO cells, but neither in Swiss-3T3 cells, nor PC12 cells. We focused 80 by as a region containing the promoter activity, which contains the elements binding to AML1 and GATA. However, the data of electrophoretic mobility shift assay (EMSA) indicated the involvement of unknown transcription factors in spite of AML1 and GATA, suggesting the presence of novel testis-specific regulatory system of human PACAP gene expression.
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