E. coli SSB activates N4 virion RNA polymerase promoters by stabilizing a DNA hairpin required for promoter recognition.
Bacteriophage N4 virion RNA polymerase transcription of double-stranded promoter-containing DNAs requires supercoiled template and E. coli single-stranded DNA-binding protein (EcoSSB); other single-stranded DNA-binding proteins cannot substitute. The DNA determinants of virion RNA polymerase binding at the promoter comprise a small template-strand hairpin. The requirement for EcoSSB is surprising, since single-stranded DNA-binding proteins destabilize hairpin structures. DNA footprinting of EcoSSB on wild-type and mutant promoters indicates that EcoSSB stabilizes the template-strand hairpin owing to the hairpin-loop sequences. Other single-stranded DNA-binding proteins destabilize the promoter hairpin, explaining the specificity of EcoSSB activation. We conclude that EcoSSB activates transcription by providing the appropriate DNA structure for polymerase binding. The existence of small hairpins stable to single-stranded protein binding suggests a novel mechanism that provides structural determinants for specific recognition in single-stranded DNA transactions by an otherwise nonspecific DNA-binding protein.
Glucksmann-Kuis
M A
MA
Department of Molecular Genetics and Cell Biology, University of Chicago, Illinois 60637, USA.
Dai
X
X
Markiewicz
P
P
Rothman-Denes
L B
LB
D001483:Base Sequence; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004268:DNA-Binding Proteins / Q000502:physiology*; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D003850:Deoxyribonuclease I / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D017902:Podoviridae / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D014158:Transcription, Genetic / Q000235:genetics
D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases; D003850:Deoxyribonuclease I