THE CHINOOK SALMON GONADOTROPIN-II-BETA SUBUNIT GENE CONTAINS A STRONG MINIMAL PROMOTER WITH A PROXIMAL NEGATIVE ELEMENT

The salmon pituitary expresses two distinct gonadotropins, gonadotropin I (GTHI) and gonadotropin II (GTHII). These two hormones are synthesized in distinct pituitary cells and secreted at different stages during the reproductive cycle. To study the transcriptional regulation of the hormone-specific beta-subunit of GTHII (sGTHII beta) gene, approximately 3.5 kilobases of the 5'-flanking region was characterized and sequenced. The pituitary specificity of sGTHII beta was examined by analyzing sGTHII beta promoter activity in homologous primary pituitary cells derived from spawning rainbow trout and in a collection of heterologous cell lines. Various lengths of the 5'-flanking region of the sGTHII beta gene were ligated into a vector encoding the bacterial chloramphenicol acetyltransferase (CAT) gene, and the resulting sGTHII beta/CAT chimeric constructs were analyzed using transient expression systems. Several constructs (-3500CAT, -1260CAT, -563CAT, and -39CAT) displayed readily detectable CAT activity in the pituitary cells derived from spawning male and female rainbow trout. In contrast, three of the constructs (-3500CAT, -1260CAT, and -563CAT) were expressed only at background levels in a variety of heterologous cell lines, suggesting that the 5'-flanking sequence of sGTHII beta contains information dictating its pituitary specificity. A silencer sequence (-95 to -35, pSil) was identified, which might function to repress sGTHII beta gene expression in the nongonadotropes or in the gonadotropes at developmental stages that precede final maturation and spawning.