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Characterization of the testis‐specific promoter region in the human pituitary adenylate cyclase‐activating polypeptide (PACAP) gene
Author(s) -
Tominaga Aiko,
Sugawara Hideki,
Futagawa Toshitaka,
Inoue Kazuhiko,
Sasaki Kazuki,
Minamino Naoto,
Hatakeyama Mamoru,
Handa Hiroshi,
Miyata Atsuro
Publication year - 2010
Publication title -
genes to cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.912
H-Index - 115
eISSN - 1365-2443
pISSN - 1356-9597
DOI - 10.1111/j.1365-2443.2010.01403.x
Subject(s) - biology , microbiology and biotechnology , adenylate kinase , gene , cyclase , frameshift mutation , exon , biochemistry , enzyme
Pituitary adenylate cyclase‐activating polypeptide (PACAP) is a pleiotropic neuropeptide localized in the testis at concentration comparable to that found in the brain, suggesting involvement in spermatogenesis. In this study, we identified the human PACAP testis‐specific exon (TSE) 10.9 kb upstream from the translational start site and found that the testis‐specific transcript of the human PACAP gene was found to be spliced from the TSE into a region of intron 2 without a frameshift. The resulting PACAP precursor has no signal peptide, suggesting that PACAP functions physiologically in an intracrine manner in the testis. The 5′‐flanking region of the TSE contains an 80‐bp fragment with potent promoter activity in testicular F9 cell. Electrophoresis mobility shift assays showed that proteins from the F9 nuclear extract interacted specifically with the 80‐bp fragment. DNA affinity chromatography allowed isolation of the specific proteins bound to the 80‐bp fragment, two of which were identified as Poly (ADP‐ribose) polymerase‐1 (PARP‐1) and TIA‐1‐related protein (TIAR) by mass spectrometry. By using their siRNAs, the depletion of their proteins in F9 cells affected the potent promoter activity of the 80‐bp fragment, suggesting that they might be involved in the testis‐specific gene expression of PACAP.