IHF- and SeqA-binding sites, present in plasmid cloning vectors, may significantly influence activities of promoters.
Escherichia coli Integration Host Factor (IHF) regulates transcription from some bacterial and phage promoters by affecting DNA topology. Here we demonstrate that IHF affects transcription from bacteriophage lambdapR promoter and the ptac promoter located on plasmids that contain IHF-binding sites. The IHF consensus sites are abundant and they can bind the IHF protein as shown in in vitro studies. The SeqA protein has a role in the complex regulation of pR activity, together with other factors altering DNA topology. Down-regulation of the transcription from ptac in the absence of IHF, together with IHF- and SeqA-mediated effects on pR, suggest that DNA topology cannot be underestimated when assessing in vivo promoters' activity. This may have a significant impact on experiments employing recombinant genes cloned in plasmids and on choosing appropriate plasmid vectors.
Lyzeń
Robert
R
Department of Molecular Biology, University of Gdańsk, Kładki 24, 80-822 Gdansk, Poland.
Kochanowska
Maja
M
Wegrzyn
Grzegorz
G
Szalewska-Pałasz
Agnieszka
A
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D010582:Bacteriophage lambda / Q000235:genetics; D001665:Binding Sites; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors / Q000235:genetics*; D034982:Integration Host Factors / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D034982:Integration Host Factors; C089746:SeqA protein, E coli