RETINOIC ACID-MEDIATED DOWN-REGULATION OF OCT3 4 COINCIDES WITH THE LOSS OF PROMOTER OCCUPANCY IN-VIVO/

Oct3/4, a hallmark of the earliest stages of embryogenesis, is express ed in undifferentiated embryonal carcinoma (EC) and embryonic stem (ES ) cells. Oct3/4 gene expression is dependent on the promoter region, t he proximal enhancer and the newly identified distal enhancer. We have analysed in vivo occupancy of these elements. In undifferentiated EC and ES cells, strong footprints were detected at specific sites of all three regulatory elements. These were promptly lost upon RA treatment in ES cells and in P19 EC cells, in parallel with sharply reduced Oct 3/4 mRNA levels, Thus, the occupancy of regulatory elements is coupled with Oct3/4 expression, and RA treatment causes coordinated factor di splacement, leading to extinction of gene activity. In F9 EC cells, fo otprint was first abolished at the proximal enhancer. However, this lo ss of binding site occupancy did not result in a decrease in Oct3/4 mR NA levels. The partial factor displacement seen in F9 EC cells, combin ed with the observation that EC and ES cells utilize the proximal and distal enhancers in a differential manner, indicate the complex patter n of Oct3/4 gene regulation, which could reflect a cell type- and line age-specific expression of the gene in vivo.