Reciprocation Sessions on Meat Science 726

We showed previously that vasoactive intestinal peptide (VIP) increases prolactin (PRL) gene expression and secretion in turkey primary pituitary cells. We have now used 5’flanking deletions and mutations of the turkey PRL promoter fused to the luciferase (Luc) reporter gene in transient transfection assays to further characterize sequences involved in stimulation of PRL gene expression by VIP. Promoter activities were determined by quantitative RT-PCR of Luc mRNA. The deletion analysis of turkey PRL promoter (tPRLP) indicated that the VIP-stimulated tPRLP activity was controlled by three major positive regulatory regions and two negative regions. From the -127 to -14 Luc construct, where the 7-8 fold increase of promoter activity by VIP occured, we did deletion assay with -92/-14 and -60/-14 Luc constructs for investigating the minimal VRE of the promoter. The 35-base pair (bp) segment (position -127 to -93) deletion induced complete suppression of VIPstimulated promoter activity, suggesting that the nucleotides between position -127 and -93 in the tPRL promoter are essential for the VIPstimulated promoter activity. A putative Pit-1 binding site was found in the middle of the 35-bp segment and the significance of this element (12 bp) was tested by Decoy assay, deletion, and mutation analysis. The result of the present study demonstrated that VRE (12bp) in the proximal prolactin promoter is an important cis-element for the VIP-stimulated PRL gene expression in turkey primary pituitary cells. USDA grant No. 00-02127