Tn7 recognizes transposition target structures associated with DNA replication using the DNA-binding protein TnsE.
We report that the bacterial transposon Tn7 selects targets by recognizing features associated with DNA replication using the transposon-encoded DNA-binding protein TnsE. We show that Tn7 transposition directed by TnsE occurs in one orientation with respect to chromosomal DNA replication, indicating that a structure or complex involved in DNA replication is likely to be a critical determinant of TnsE insertion. We find that mutant TnsE proteins that allow higher levels of transposition also bind DNA better than the wild-type protein. The increased binding affinity displayed by the TnsE high-activity mutants indicates that DNA binding is relevant to transposition activity and suggests that TnsE interacts directly with target DNAs. In vitro, TnsE interacts preferentially with certain DNA structures, indicating a mechanism for the TnsE-mediated orientation and insertion preference. The pattern of TnsE-mediated insertion events around the Escherichia coli chromosome provides insight into how DNA replication forks proceed in vivo.
Peters
J E
JE
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Craig
N L
NL
D000483:Alleles; D000595:Amino Acid Sequence; D015153:Blotting, Western; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication; D004251:DNA Transposable Elements / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C424904:TnsE protein, E coli; D004247:DNA