TIMING OF REPLICATION OF DIFFERENTIALLY TRANSCRIBED GENES IN SYRIAN-HAMSTER EMBRYO FIBROBLASTS

Syrian hamster embryo cell lines have been used as models of neoplasti c progression in vitro. Changes in phenotype and biological properties have been observed in these cell lines in association with modulation of gene transcription. We explored this natural evolution of cells in culture to investigate the reported relationship between transcriptio nal activity and replication in early S phase. In this study we used t wo nontumorigenic cell lines that had either retained (supB+) or lost (supB-) the ability to suppress tumorigenicity of malignantly transfor med hamster fibroblasts (BP6T). In association with the loss of suppre ssor gene function, supB cells have downregulated the expression of th e H19 and tropomyosin-I (TM-I) genes, which are actively transcribed i n supB+ cells. Synchronous populations of supB+ and supB- cells were p ulse-labeled with [H-3]thymidine and bromodeoxyuridine at 1-h interval s during S phase; the replicating DNA was isolated by centrifugation i n cesium chloride gradients and hybridized to P-32 labeled gene probes . No correlation was found between the timing of gene replication and the status of expression of these two genes. TM-I replicated during th e first hour and H19 replicated between the second and third hours of the s phase in the expressing and nonexpressing cell lines. Immunoglob ulin gene sequences, known to be late-replicating infibroblasts, repli cated at the end of the S phase. These results suggest that downregula tion of transcription is not always accompanied by a concomitant chang e in time of gene replication from early to late S phase. (C) 1993 Wil ey-Liss, Inc.