Pituitary adenylate cyclase-activating polypeptide gene expression regulated by a testis-specific promoter in germ cells during spermatogenesis.
Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the glucagon-related family of hormones that is widely expressed in various tissues. The PACAP messenger RNA (mRNA) and protein is expressed at high levels in the germ cells of the testis, where it locally activates cAMP-coupled receptors located in the somatic Sertoli cells. The PACAP mRNA expressed specifically in the testis is shorter than the mRNA expressed in hypothalamus and includes 127 nucleotides of novel sequence at the 5'-end, suggesting a different start site of transcription in the testes and the utilization of a tissue-specific promoter. Here we present evidence that a single PACAP gene uses a testis-specific promoter to express a mRNA containing a unique exon located 13.5 kb upstream from the first coding exon. As determined by RT-PCR analysis of testis mRNA, the expression of the first testis-specific exon is relatively specific for the testis, as no PACAP mRNA containing the testis-specific first exon was detected in hypothalamic mRNAs. The promoter for the testis-specific PACAP gene was cloned, and a start site for transcription was mapped by primer extension. The testis-specific promoter sequence directs germ cell-specific expression upon transfection of promoter-transcriptional reporter plasmids to populations of testicular cells in vitro and upon expression of a promoter-reporter transgene in mice. Analyses of PACAP gene expression during the spermatogenic cycle, accomplished by RT-PCR of segments of isolated seminiferous tubules, identified intense expression in the postmeiotic round spermatids during developmental stages I-VIII. These observations establish the existence of a specialized PACAP gene promoter whose activity is highly regulated during the spermatogenic cycle.
Daniel
P B
PB
Laboratory of Molecular Endocrinology, Massachusetts General Hospital, Howard Hughes Medical Institute, and Harvard Medical School, Boston 02114, USA.
Habener
J F
JF
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D015139:Blotting, Southern; D004247:DNA / Q000096:biosynthesis / Q000235:genetics; D005091:Exons / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology*; D005854:Germ Cells / Q000378:metabolism / Q000502:physiology*; D008297:Male; D008540:Meiosis / Q000502:physiology; D051379:Mice; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000096:biosynthesis*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D012333:RNA, Messenger / Q000096:biosynthesis / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012671:Seminiferous Tubules / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013091:Spermatogenesis / Q000502:physiology*; D013737:Testis / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D014162:Transfection
C495402:Adcyap1 protein, mouse; C495403:Adcyap1 protein, rat; D009479:Neuropeptides; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D012333:RNA, Messenger; D004247:DNA