title: Ubiquitous promoter-localization of essential virulence regulators in Francisella tularensis.
abstract: Francisella tularensis is a Gram-negative bacterium whose ability to replicate within macrophages and cause disease is strictly dependent upon the coordinate activities of three transcription regulators called MglA, SspA, and PigR. MglA and SspA form a complex that associates with RNA polymerase (RNAP), whereas PigR is a putative DNA-binding protein that functions by contacting the MglA-SspA complex. Most transcription activators that bind the DNA are thought to occupy only those promoters whose activities they regulate. Here we show using chromatin immunoprecipitation coupled with high-throughput DNA sequencing (ChIP-Seq) that PigR, MglA, and SspA are found at virtually all promoters in F. tularensis and not just those of regulated genes. Furthermore, we find that the ability of PigR to associate with promoters is dependent upon the presence of MglA, suggesting that interaction with the RNAP-associated MglA-SspA complex is what directs PigR to promoters in F. tularensis. Finally, we present evidence that the ability of PigR (and thus MglA and SspA) to positively control the expression of genes is dictated by a specific 7 base pair sequence element that is present in the promoters of regulated genes. The three principal regulators of virulence gene expression in F. tularensis therefore function in a non-classical manner with PigR interacting with the RNAP-associated MglA-SspA complex at the majority of promoters but only activating transcription from those that contain a specific sequence element. Our findings reveal how transcription factors can exert regulatory effects at a restricted set of promoters despite being associated with most or all. This distinction between occupancy and regulatory effect uncovered by our data may be relevant to the study of RNAP-associated transcription regulators in other pathogenic bacteria.
authors.affiliation: Division of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Ramsey
authors.affiliation: Division of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Melisa L
authors.initials: ML
authors.lastname: Osborne
authors.affiliation: Department of Genetics and Waksman Institute, Rutgers University, Piscataway, New Jersey, United States of America.
authors.forename: Irina O
authors.initials: IO
authors.lastname: Vvedenskaya
authors.affiliation: Division of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Cathy
authors.initials: C
authors.lastname: Su
authors.affiliation: Department of Genetics and Waksman Institute, Rutgers University, Piscataway, New Jersey, United States of America.
authors.forename: Bryce E
authors.initials: BE
authors.lastname: Nickels
authors.affiliation: Division of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Simon L
authors.initials: SL
authors.lastname: Dove
mesh_terms: D047369:Chromatin Immunoprecipitation; D018274:Electroporation; D005604:Francisella tularensis / Q000235:genetics* / Q000472:pathogenicity*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D005798:Genes, Bacterial; D059014:High-Throughput Nucleotide Sequencing; D015151:Immunoblotting; D011401:Promoter Regions, Genetic / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*; D014774:Virulence / Q000235:genetics
chemical_list: D014157:Transcription Factors
references.citation: Mol Microbiol. 2000 Aug;37(4):687-95
references.pmid: 10972792
references.citation: Cell. 2001 Mar 23;104(6):901-12
references.pmid: 11290327
references.citation: Biochem Biophys Res Commun. 2002 Mar 8;291(4):979-86
references.pmid: 11866462
references.citation: Mol Microbiol. 2002 Mar;43(6):1471-91
references.pmid: 11952899
references.citation: Mol Microbiol. 2002 Jan;43(2):355-70
references.pmid: 11985714
references.citation: FEMS Microbiol Lett. 2002 Sep 24;215(1):53-6
references.pmid: 12393200
references.citation: Mol Microbiol. 2003 Jun;48(6):1621-31
references.pmid: 12791143
references.citation: Infect Immun. 2003 Oct;71(10):5940-50
references.pmid: 14500514
references.citation: Proc Natl Acad Sci U S A. 2004 Mar 23;101(12):4246-9
references.pmid: 15010524
references.citation: Nat Rev Microbiol. 2004 Jan;2(1):57-65
references.pmid: 15035009
references.citation: Infect Immun. 2004 Jun;72(6):3204-17
references.pmid: 15155622
references.citation: J Bacteriol. 2004 Oct;186(19):6430-6
references.pmid: 15375123
references.citation: J Mol Biol. 2004 Nov 12;344(1):1-10
references.pmid: 15504398
references.citation: Nat Rev Microbiol. 2004 Dec;2(12):967-78
references.pmid: 15550942
references.citation: Appl Environ Microbiol. 2004 Dec;70(12):7511-9
references.pmid: 15574954
references.citation: Nat Genet. 2005 Feb;37(2):153-9
references.pmid: 15640799
references.citation: J Biol Chem. 2005 Apr 29;280(17):17380-91
references.pmid: 15735307
references.citation: J Bacteriol. 2005 Sep;187(17):6166-74
references.pmid: 16109958
references.citation: Mol Cell. 2005 Nov 11;20(3):357-66
references.pmid: 16285918
references.citation: Infect Immun. 2006 Dec;74(12):6642-55
references.pmid: 17000729
references.citation: Ann N Y Acad Sci. 2007 Jun;1105:122-37
references.pmid: 17395722
references.citation: Ann N Y Acad Sci. 2007 Jun;1105:1-29
references.pmid: 17395726
references.citation: PLoS Pathog. 2007 Jun;3(6):e84
references.pmid: 17571921
references.citation: Infect Immun. 2008 Aug;76(8):3473-80
references.pmid: 18559431
references.citation: Proc Natl Acad Sci U S A. 2008 Dec 2;105(48):18947-52
references.pmid: 19028873
references.citation: Nat Methods. 2008 Sep;5(9):829-34
references.pmid: 19160518
references.citation: Cell Microbiol. 2009 Jul;11(7):1128-50
references.pmid: 19388904
references.citation: Microbiology. 2009 Aug;155(Pt 8):2560-72
references.pmid: 19443547
references.citation: Bioinformatics. 2009 Aug 15;25(16):2078-9
references.pmid: 19505943
references.citation: Cell. 2009 Jul 10;138(1):146-59
references.pmid: 19596241
references.citation: BMC Microbiol. 2009 Oct 12;9:216
references.pmid: 19821974
references.citation: PLoS Pathog. 2009 Oct;5(10):e1000641
references.pmid: 19876386
references.citation: Mol Microbiol. 2009 Dec;74(6):1459-70
references.pmid: 20054881
references.citation: Bioinformatics. 2010 Mar 15;26(6):841-2
references.pmid: 20110278
references.citation: Infect Immun. 2010 May;78(5):2189-98
references.pmid: 20231408
references.citation: Bioinformatics. 2011 Apr 1;27(7):1017-8
references.pmid: 21330290
references.citation: Front Microbiol. 2010 Dec 28;1:138
references.pmid: 21687806
references.citation: Mol Cell. 2011 Jun 24;42(6):817-25
references.pmid: 21700226
references.citation: Nat Methods. 2012 Mar 04;9(4):357-9
references.pmid: 22388286
references.citation: Brief Bioinform. 2013 Mar;14(2):178-92
references.pmid: 22517427
references.citation: Genes Dev. 2012 Jul 1;26(13):1498-507
references.pmid: 22751503
references.citation: BMC Microbiol. 2012 Oct 11;12:231
references.pmid: 23051860
references.citation: Proc Natl Acad Sci U S A. 2013 Jul 30;110(31):12619-24
references.pmid: 23858468
references.citation: PLoS One. 2013 Aug 22;8(8):e72329
references.pmid: 23991093
references.citation: PLoS Pathog. 2013;9(9):e1003582
references.pmid: 24039572
references.citation: Microbes Infect. 2013 Oct 25;:null
references.pmid: 24513705
references.citation: Curr Opin Microbiol. 2014 Jun;19:97-105
references.pmid: 25024085
references.citation: J Bacteriol. 2014 Oct;196(19):3516-26
references.pmid: 25070738
references.citation: Cell Host Microbe. 2014 Aug 13;16(2):227-236
references.pmid: 25070807
references.citation: Gene. 1989 Apr 15;77(1):51-9
references.pmid: 2744487
references.citation: Proc Int Conf Intell Syst Mol Biol. 1994;2:28-36
references.pmid: 7584402
references.citation: J Bacteriol. 1994 Jul;176(13):4081-91
references.pmid: 8021189
references.citation: Cell. 1996 Aug 9;86(3):495-501
references.pmid: 8756731
references.citation: Nature. 1997 Apr 10;386(6625):627-30
references.pmid: 9121589
references.citation: J Bacteriol. 1998 Feb;180(4):793-800
references.pmid: 9473031
references.citation: Cell. 1998 Mar 6;92(5):597-600
references.pmid: 9506513
references.citation: Mol Microbiol. 1998 Jul;29(1):247-59
references.pmid: 9701818
references.citation: Res Microbiol. 1997 May;148(4):289-303
references.pmid: 9765808