A REPLICATION-ENHANCING ELEMENT WITH TRANSCRIPTIONAL SILENCER ACTIVITY IN AUTONOMOUSLY REPLICATING HUMAN CHROMOSOMAL FRAGMENTS

We have identified specific nucleotide sequences involved in autonomou s replication of human chromosomal fragments in human cells. Nested de letion analysis of a 10.2-kb long human chromosomal fragment showed th at replication efficiency of the fragment was reduced to about 50% by loss of a short specific segment. Deletions outside the segment reduce d the replication efficiency depending on their lengths. By introducin g linker substitutions, we found that the distinct segment required fo r the efficient replication consisted of an 18-bp sequence, named REE1 (Replication Enhancing Element 1). Single or tandem copies of REE1 al one had no significant replication activity, but they stimulated repli cation of human chromosomal DNA fragments. We found, in addition, that the REE1 sequence inserted at a site 2.7 kb upstream of the SV40 earl y promoter caused repression of transcription from the promoter, sugge sting that REE1 had a transcriptional silencer activity. introduction of linker-substitutions into the REE1 indicated that the nucleotide se quences required for the repression of transcription were the same as those for enhancement of replication. Thus, REE1 is responsible for bo th enhancement of replication and repression of transcription.