title: Cis-acting effects of sequences within 2.4-kb upstream of the human c-myc gene on autonomous plasmid replication in HeLa cells.
abstract: We have used density shift analysis to monitor the autonomous replicating sequence (ARS) activity of plasmids containing various DNA fragments from the 5'-flanking region of the human c-myc gene. The ARS activity of certain of these plasmids implied that structures in the c-myc DNA could be recognized for the initiation of replication in the absence of chromosomal integration. The plasmid pNeo.Myc-2.4 contains 2.4 contains 2.4 kb of c-myc 5'-flanking DNA, and replicated semiconservatively as a circular extrachromosomal element. Deletion derivatives of pNeo.Myc-2.4 containing either of two nonoverlapping regions of c-myc DNA semiconservatively incorporated bromodeoxyuridine into discrete populations of heavy-light supercoiled molecules to roughly the same extent as the chromosomal DNA in the same cultures. Some constructs displayed lower ARS activity, implying that distinct cis-acting sequences in the c-myc 5'-flanking DNA may independently affect DNA replication. The ARS activity of two separate c-myc sequences suggests that replication initiation signals are redundant in the c-myc origin. The smallest c-myc insert that displayed substantial ARS activity was 930 bp long and contained three 10/11 matches to the yeast ARS consensus and several additional features found in eukaryotic replication origins.
authors.affiliation: Department of Biochemistry and Molecular Biology, Wright State University, Dayton, OH 45435, USA.
authors.forename: C
authors.initials: C
authors.lastname: McWhinney
authors.forename: S E
authors.initials: SE
authors.lastname: Waltz
authors.forename: M
authors.initials: M
authors.lastname: Leffak
mesh_terms: D001483:Base Sequence; D001973:Bromodeoxyuridine; D002877:Chromosomes, Human / Q000502:physiology; D016384:Consensus Sequence; D004261:DNA Replication; D004273:DNA, Neoplasm / Q000235:genetics; D005809:Genes, Regulator; D016259:Genes, myc; D006367:HeLa Cells; D006801:Humans; D010957:Plasmids; D015183:Restriction Mapping; D017384:Sequence Deletion
chemical_list: D004273:DNA, Neoplasm; D001973:Bromodeoxyuridine