Organization of Mouse DNA Polymerase β Gene Silencer Elements and Identification of the Silencer‐binding Factor(s)

Different portions of the 5‐upstream region of the mouse DNA polymerase β gene were combined with bacterial chloramphenicol acetyltransferase (CAT) gene of the CAT vector. Transfection of these recombinant plasmids into mouse NIH/3T3 cells has revealed that each of the previously identified two negatively acting regions (silencers I and II) of this gene consists of multiple sub‐domains. The distal silencer (silencer I) at around ‐1.5 kb consists of four sub‐domains (‐1852 to ‐1667, ‐1663 to ‐1616, ‐1564 to ‐1525 and ‐1355 to ‐1257). The promoter‐proximal silencer (silencer II) at around – 0.5 kb consists of two functional domains (‐ 681 to – 523 and – 490 to – 447) separated by a neutral region of 33 base pairs. Silencer II functioned efficiently when silencer I was deleted. Conversely, the distal silencer I functioned efficiently when silencer II was deleted. Thus, these silencers functioned redundantly to each other in NIH/3T3 cells. Nucleotide sequence analysis revealed no extensive sequence similarity between these two silencers. Significant sequence similarity is present between a distal portion of silencer II and the c‐myc gene silencer, and also between a proximal portion of silencer II and the mouse F9 cell‐specific silencer. A protein factor(s) that specifically bound to the silencer elements was detected in nuclear extracts of NIH/3T3 cells and mouse liver in which DNA polymerase β was expressed at a rather low level. The same binding factor(s) can bind to both silencer I and II regions, although its affinity for silencer II is much higher than that for silencer I.