Presence of a spermatogenic-specific promoter in the rat growth hormone-releasing hormone gene.

A GH-releasing hormone (GHRH) messenger RNA (mRNA) has been identified in hypothalamus, placenta, and testicular germ cells. The GHRH mRNA produced by spermatogenic cells is approximately 1700 nucleotides in length, whereas GHRH transcripts in hypothalamus and placenta are 750 nucleotides. To correlate the structure of testicular GHRH mRNA with cell type-specific expression, we determined its sequence. A GHRH clone isolated from a rat testicular complementary DNA library was found to be identical in the coding sequence to hypothalamic GHRH. Rapid amplification of complementary DNA ends analysis of the 5'-end of germ cell GHRH mRNA and comparison with the genomic sequence revealed that GHRH transcription in testis initiates approximately 700 basepairs 5' to transcription initiation in placenta and 10.7 kilobasepairs 5' to that in hypothalamus. Reverse transcription-polymerase chain reaction analysis of germ cell RNA using primers from testicular exons 1 and 4 demonstrated that part of the placental exon 1 sequence is contained in some testicular GHRH transcripts, as an extra exon, between testicular exon 1 and the common exon 2. This was confirmed by a Northern blot of testicular mRNA using a testicular exon 1 probe. The 5'-flanking region of the testicular GHRH gene was analyzed and found to contain a TATA-like motif and sequences homologous to spermatogenic-specific cis-acting elements. Southern blot analysis of rat liver DNA suggested that just one GHRH gene is present in rat. These results indicate that both alternative transcription initiation and splicing of the GHRH gene exist in rat testicular germ cells.