Abstract
The unorthodox two-component sensor protein BvgS ofBordetella pertussis contains several interesting sequence motifs of unknown functional relevance, such as a histidine motif in its output domain, which is conserved among several unorthodox sensor proteins, a putative nucleotide binding site [Walker box type A] in its linker region, and a region in its periplasmic domain with significant homology to the TonB protein ofEscherichia coli. We investigated potential functions of these sequences by constructingB. pertussis strains that express mutant BvgS derivatives. The His1172 residue in the output domain was exchanged for Gln, and the Walker motif was mutated either by the replacement of Lys625 by Arg, or of Gly624 by Val and Lys625 by Leu. To analyse the TonB motif, the periplasmic domain of BvgS was replaced with the corresponding domain of EvgS, anE. coli sensor that is highly homologous to BvgS but lacks the similarity with TonB. All mutations except the conservative Lys/Arg exchange in the Walker box caused the inactivation of BvgS, indicating the functional importance of the conserved motifs. The activity of the mutant proteins could be restored by complementation in trans with various separately expressed, truncated parts of BvgS. Mutations in the BvgS receiver domain could be complemented not only by a construct expressing the wild-type receiver and output domains, but also by the derivative containing the His-Gln exchange. Therefore, the histidine motif, although important for BvgS function, is not essential for complementation of BvgS mutants. The mutations in the Walker box and in the periplasmic domain could be complemented by a truncated BvgS derivative lacking the receiver and output domains. The characterization of a spontaneous revertant of the strain expressing the originally inactive EvgS/BvgS hybrid protein revealed the presence of a mutation in the BvgS linker region, conferring constitutive activity on the protein. As TonB energizes transport processes across the outer membrane ofE. coli, the strain expressing the constitutive EvgS/BvgS hybrid protein lacking the TonB motif was used in preliminary investigations of a possible direct involvement of BvgS in transport processes.
Similar content being viewed by others
References
Arico B, Miller JF, Roy CR, Stibitz S, Monack D, Falkow S, Gross R, Rappuoli R (1989) Sequences required for expression ofBordetella pertussis virulence factors share homology with prokaryotic signal transduction proteins. Proc Natl Acad Sci USA 86: 6671–6675
Bordet J, Gengou O (1909) L'endotoxine coquelucheuse. Ann Inst Pasteur (Paris) 23: 415–419
Braun V (1995) Energy-coupled transport and signal transduction through the gram-negative outer membrane via TonB-ExbB-ExbD-dependent receptor proteins. FEMS Microbiol Rev 16: 295–307
Beier D, Schwarz B, Fuchs TM, Gross R (1995) In vivo characterization of the unorthodox BvgS two-component sensor protein ofBordetella pertussis. J Mol Biol 248: 596–610
Carbonetti NH, Fuchs TM, Patamawenu AA, Irish TJ, Deppisch H, Gross R (1994) Effect of mutations causing overexpression of RNA polymeraseα subunit on regulation of virulence factors inBordetella pertussis. J Bacteriol 176: 7267–7273
Chang C, Kwok SF, Bleecker AB, Meyerowitz EM (1993)Arabidopsis ethylene-response geneETR1: similarity of product to two-component regulators. Science 262: 539–544
Charles TC, Nester EW (1993) A chromosomally encoded two-component sensory tranduction system is required for virulence ofAgrobacterium tumefaciens. J Bacteriol 175: 6614–6625
Delepelaire P (1994) PrtD, the integral membrane ATP-binding cassette component of theErwinia chrysanthemi metalloprotease secretion system, exhibits a secretion signal-regulated ATPase activity. J Biol Chem 269: 27952–27957
Giardina PC, Foster LA, Musser JM, Akerley BJ, Miller JF, Dyer DW (1995)bvg repression of alcaligin synthesis inBordetella bronchiseptica is associated with phylogenetic lineage. J Bacteriol 177: 6058–6063
Glaser P, Ladant D, Sezer O, Pichot F, Ullmann A, Danchin A (1988) The calmodulin-sensitive adenylate cyclase ofBordetalla pertussis: cloning and expression inEscherichia coli. Mol Microbiol 2: 19–30
Goyard S, Bellalou J, Mireau H, Ullmann A (1994) Mutations in theBordetella pertussis bvgS gene that confer altered expression of thefhaB gene inEscherichia coli. J Bacteriol 176: 5163–5166
Gross R (1993) Signal transduction in human and animal pathogens. FEMS Microbiol Rev 104: 301–326
Gross R, Rappuoli R (1988) Positive regulation of pertussis toxin expression. Proc Natl Acad Sci USA 85: 3913–3917
Gross R, Rappuoli R (1989) Pertussis toxin promoter sequences involved in modulation. J Bacteriol 171: 4026–4030
Gross R, Arico B, Rappuoli R (1989) Families of bacterial signal transduction proteins. Mol Microbiol 3: 1661–1667
Ishige K, Nagasawa S, Tokishita S, Mizuno T (1994) A novel device of bacterial signal transducers. EMBO J 13: 5195–5202
Iuchi S, Lin ECC (1992) Mutational analysis of signal transduction by ArcB, a membrane sensor protein for anaerobic repression of operons involved in the central aerobic pathways inEscherichia coli. J Bacteriol 174: 3972–3980
Kunkel TA, Roberts JD, Zakour RA (1987) Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol 154: 367–383
Lacey BW (1960) Antigenic modulation ofBordetella pertussis. J Hyg 58: 57–93
Leininger E, Roberts M, Kenimer JG, Charles IG, Fairweather N, Novotny P, Brennan J (1991) Pertactin, an Arg-Gly-Asp-containingBordetella pertussis surface protein that promotes adherence of mammalian cells. Proc Natl Acad Sci USA 88: 345–349
Locht C, Keith, JM (1986) Pertussis toxin gene: nucleotide sequence and genetic organization. Science 232: 1258–1264
Manetti R, Arico B, Rappuoli R, Scarlato V (1994) Mutations in the linker region of BvgS abolish response to signals for the regulation of the virulence factors inBordetella pertussis. Gene 150: 123–127
Miller JF, Mekalanos JJ, Falkow S (1989) Coordinate regulation and sensory transduction in the control of bacterial virulence. Science 243: 916–922
Miller JF, Johnson SA, Black WJ, Beattie DT, Mekalanos JJ, Falkow S (1992) Isolation and analysis of constitutive sensory transduction mutations in theBordetella pertussis bvgS gene. J Bacteriol 174: 970–979
Morales VM, Backman A, Bagdasarian M (1991) A series of wide-host-range low-copy-number vectors that allow direct screening for recombinants. Gene 97: 39–47
Nicosia A, Perugini M, Franzini C, Casagli MC, Borri MG, Antoni G, Almoni M, Neri P, Ratti G, Rappuoli R (1986) Cloning and sequencing of the pertussis toxin genes: operon structure and gene duplication. Proc Natl Acad Sci USA 83: 4631–4635
Ota IM, Varshavsky A (1993) A yeast protein similar to bacterial two-component regulators. Science 262: 4631–4635
Parkinson JS (1995) Genetic approaches for signalling pathways and proteins. In: Hoch JA, Silhavy TJ (eds) Two-component signal transduction. ASM Press, Washington DC, pp 9–23
Parkinson JS, Kofoid EC (1992) Communication modules in bacterial signaling proteins. Annu Rev Genet 26: 71–112
Relman DA, Domenighini M, Tuomanen E, Rappuoli R, Falkow S (1989) Filamentous hemagglutinin ofBordetella pertussis: nucleotide sequence and crucial role in adherence. Proc Natl Acad Sci USA 86: 2637–2641
Roy CR, Falkow S (1991) Identification ofBordetella pertussis regulatory sequences required for transcriptional activation of thefhaB gene and autoregulation of thebvgAS operon. J Bacteriol 173: 2385–2392
Sambrook J, Fritsch EF, Maniatis T (1992) Molecular cloning: a laboratory manual (2nd edn). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
Scarlato V, Arico B, Prugnola A, Rappuoli R (1991) Sequential activation and environmental regulation of virulence genes inBordetella pertussis. EMBO J 10: 3971–3975
Simon R, Priefer U, Pühler A (1983) A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in gram-negative bacteria. Bio/Technology 1: 37–45
Stainer DW, Scholte MJ (1970) A simple chemically defined medium for the production of phase IBordetella pertussis. J Gen Microbiol 63: 211–220
Stephens KM, Roush C, Nester E (1995)Agrobacterium tumefaciens VirB11 protein requires a consensus nucleotide-binding site for function in virulence. J Bacteriol 177: 27–36
Stibitz S, Yang MS (1991) Subcellular location and immunological detection of proteins encoded by thevir locus ofBordetella pertussis. J Bacteriol 173: 4288–4296
Stock J, Ninfa AJ, Stock AM (1989) Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiol Rev 53: 450–490
Tabor S, Richardson CC (1985) A bacteriophage T7 RNA polymerase-promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci USA 82: 1074–1078
Uhl MA, Miller JF (1994) Autophosphorylation and phosphotransfer in theBordetella pertussis BvgAS signal transduction cascade. Proc Natl Acad Sci USA 91: 1163–1167
Utsumi R, Katayama S, Taniguchi M, Horie T, Ikeda M, Igaki S, Nakagawa H, Miwa A, Noda M (1993) Newly identified genes involved in the signal transduction ofEscherichia coli K-12. Gene 140: 73–77
Walker JE, Sarasta M, Runswick MJ, Gay NJ (1982) Distantly related sequences in the alpha- and beta-subunit of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J 1: 945–951
Weiss AA, Falkow S (1984) Genetic analysis of phase change inBordetella pertussis. Infect Immun 43: 263–269
Weiss AA, Hewlett EL (1986) Virulence factors ofBordetella pertussis. Annu Rev Microbiol 40: 661–686
Willems R, Paul A, van der Heide HGJ, ter Avest AR, Mooi FR (1990) Fimbrial phase variation inBordetella pertussis: a novel mechanism for transcriptional regulation. EMBO J 9: 2803–2809
Author information
Authors and Affiliations
Additional information
Communicated by W. Arber
Rights and permissions
About this article
Cite this article
Beier, D., Deppisch, H. & Gross, R. Conserved sequence motifs in the unorthodox BvgS two-component sensor protein ofBordetella pertussis . Molec. Gen. Genet. 252, 169–176 (1996). https://doi.org/10.1007/BF02173217
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02173217