Extent of BrdUrd-induced prolactin gene amplification in GH cells

The thymidine analog, 5-bromodeoxyuridine (BrdUrd), induces prolactin (Prl) synthesis and Prl gene amplification in a subclone of GH cells (rat pituitary tumor cells in culture). Withdrawal of drug treatment reverses both processes. Our previous results show that Prl gene amplification is associated with an event involving extrachromosomal DNA. The results presented in this report show that a 20-kb length of DNA, including all the coding sequences of the Prl gene, is amplified following BrdUrd treatment of this strain of GH cells. Amplification of DNA sequences extends about 3 kb upstream from the 5' end and about 7 kb downstream from the 3' end of the Prl gene. The DNA sequences immediately adjacent to these 5'-and 3'-flanking regions of the Prl gene are not affected by BrdUrd treatment. These results suggest that BrdUrd-induced amplification of Prl gene coding and neighboring sequences is restricted to within 3 kb of the 5' end and within 7 kb of the 3' end of the structural gene Analysis of low-molecular-weight extrachromosomal DNA isolated by Hirt's procedure from drug-treated cells reveals a similar pattern of amplification of the Prl gene and its flanking sequences.