FEMALE REPRODUCTIVE TRACT - B
Wednesday, August 4, 2004
10:30 AM–12:30 PM
(820) TRANSCRIPTIONAL REGULATION OF CALBINDIN-D9K GENE AND PUTATIVE REPRESSOR AT THE PROMOTER REGION OF THIS GENE IN A SPECIES-SPECIFIC MANNER.
Lee, Geun-Shik 1, Choi, Kyung-Chul2, Jeung, Eui-Bae1, 3, 1 Laboratory of Veterinary Biochemistry and Molecular Biology, Cheongju, Republic of Korea2 Department of Obstetrics and Gynecology, University of British Columbia, Vancouver, BC, Canada3 Xenotransplantation Research Center, Seoul, Rebublic of Korea
ABSTRACT- Calbindin-D9k (CaBP-9k) expressed highly in duodenum, kidney and reproductive organs has high affinity for calcium which is an important factor related with metabolism and reproduction. Previous studies have shown that the expression of CaBP-9k gene is regulated by 1,25-dihydroxyvitamin D3, estrogen (E2) or progesterone (P4) in a tissue-specific manner. However, the molecular mechanisms governing its transcription level remain poorly understood due to the lack of an appropriate cell line. In the present study, we characterized the genomic structure of porcine and bovine CaBP-9k using BAC sequencing and Gene walking to identify the cis-acting elements functioned to activate or repress the CaBP-9k gene transcription. Promoter assays following transfection of the various cell lines (IEVT, JEG-3, MCF-7, T47D, and HUTU-80) with the short 5-flanking region of human (-1311 to +304), rat (-884 to +327), mice (-1186 to +150), porcine (-857 to +501) and bovine (-696 to +535) of CaBP-9k gene revealed that the promoter regions of human and mice have relatively high luciferase activity (40% vs. SV40 promoter). In addition, placental IEVT cell line has been shown to possess the highest promoter activity (2-folds vs. other cell lines) despites the lack of CaBP-9k expression. It is of interest that the AREB6 located nearby GATA1 is absent on the 5-flanking regions of rat, porcine and bovine CaBP-9k gene, whereas this region is present on the 5-flanking regions of human and mice, which show the high promoter activity. In an extended human CaBP-9k promoter (-4712 to +304), the promoter activity was completely lost, suggesting the existence a putative portent repressor between -4712 and -1312. The transcriptional activity of short promoter region was not altered by sex steroid hormone, estrogen (E2), progesterone (P4), or E2+P4, assumed as a key regulator on the uterine CaBP-9k. These findings indicate that the molecular mechanism of CaBP-9k is differentially implicated in a species-specific manner and a portent repressor may take part in the suppression of CaBP-9k gene transcription. [This work was supported by grant R01-2002-000-00015-0 (2002) from the Basic Research Program of the Korea Science & Engineering Foundation]
KEY WORDS: Uterus, Calbindin-D9k, Transcriptional regulation