Regulation of pap pilin phase variation

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Title Regulation of pap pilin phase variation
Publication Type dissertation
School or College School of Medicine
Department Pathology
Author Blyn, Lawrence Bruce
Date 1991-06
Description The regulation of expression of the pyelonephritis-associated pili (pap) pilin gene in Escherichia coli was investigated using a single copy lacZYA fusion system. The expression of pap-encoded pili is under the control of a heritable phase variation mechanism. Pap phase-variation is transcriptionally regulated and responsive to changes in the carbon source used for growth and the incubation temperature. Pilin phase variation occurs without DNA rearrangement of pap DNA sequences. Analysis of the DNA regulatory region upstream of the papBA promoter showed that there are two nearly perfect 27 base pair inverted repeats separated by 89 base pairs. Each inverted repeat contains a single Dam (deoxyadenosine methylase) methylation site (GATC1028 and GATC1130 sites). Using a pap-lac gene fusion, as well as primer extension analysis of papB mRNA, it was found that the dam gene product is required for pap transcription. The methylation states of the GATC028 and GATC1130 sites in DNAs isolated from transcriptionally phase on and phase off cells were different as evidenced by Southern blot experiments. The GATC1028 site was found to be unmethylated only in DNA isolated from phase on populations. Conversely, GATC1130 sites were unmethylated in DNA isolated from phase off populations. E. coli mutants were isolated that failed to protect both the GATC1028 and GATC 1130 sites from methylation by Dam. The defective gene(s) in these mutants was cloned and designated as mbf for methylation blocking factor. The mbf gene is located at 19.6 min on the E. coli chromosome. In mbf+ E. coli the GATC1130 site is umethylated (the phase off methylation state) in the absence of any pap-encoded proteins. Methylation protection of GATC1028 (the phase on methylation state) requires both mbf and papI, a regulatory gene within the pap operon, indicating that pap pilin phase variation requires the action of at least two genes, papI and mbf. A model is presented for pap pilin phase variation which predicts that the GATC methylation state regulates Mbf and PapI binding to pap DNA and that binding of Mbf and PapI inhibits the methylation of GATC sites by Dam methylase.
Type Text
Publisher University of Utah
Subject Uropathogenic coli; Methylationstates
Subject MESH Fimbriae, Bacterial; Escherichia coli; Microbiology; Gene Expression
Dissertation Institution University of Utah
Dissertation Name PhD
Language eng
Relation is Version of Digital reproduction of "The regulation of pap pilin phase variation." Spencer S. Eccles Health Sciences Library. Print version of "The regulation of pap pilin phase variation." available at J. Willard Marriott Library Special Collection. QR6.5 1991 .B58.
Rights Management © Lawrence Bruce Blyn.
Format application/pdf
Format Medium application/pdf
Format Extent 3,841,747 bytes
Identifier undthes,4872
Source Original: University of Utah Spencer S. Eccles Health Sciences Library (no longer available).
Master File Extent 3,841,816 bytes
ARK ark:/87278/s6j9688r
Setname ir_etd
ID 191934
Reference URL https://collections.lib.utah.edu/ark:/87278/s6j9688r
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