Characterization of Antigenic Determinants of Borrelia burgdorferi Shared by Other Bacteria

Coleman, James L.; Benach, Jorge L.

doi:10.1093/infdis/165.4.658

Cited by 66 publications

(57 citation statements)

References 48 publications

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“…Monoclonal antibodies kindly provided by other investigators included the following: anti-FlaB (H9724) by A. Barbour (University of California, Irvine, CA), anti-FlaA by B. Johnson (Centers for Disease Control and Prevention, Atlanta, GA), and anti-DnaK by J. Benach (SUNY, Stony Brook, NY). Specific reactivity to B. burgdorferi FlaA, FlaB, and DnaK has been reported previously (4,11,20,39,40).…”

Section: Methodssupporting

confidence: 68%

CheX Is a Phosphorylated CheY Phosphatase Essential forBorrelia burgdorferiChemotaxis

Motaleb¹,

Miller

Li³

et al. 2005

J Bacteriol

112

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Motility and chemotaxis are believed to be important in the pathogenesis of Lyme disease caused by the spirochete Borrelia burgdorferi. Controlling the phosphorylation state of CheY, a response regulator protein, is essential for regulating bacterial chemotaxis and motility. Rapid dephosphorylation of phosphorylated CheY (CheY-P) is crucial for cells to respond to environmental changes. CheY-P dephosphorylation is accomplished by one or more phosphatases in different species, including CheZ, CheC, CheX, FliY, and/or FliY/N. Only a cheX phosphatase homolog has been identified in the B. burgdorferi genome. However, a role for cheX in chemotaxis has not been established in any bacterial species. Inactivating B. burgdorferi cheX by inserting a flgB-kan cassette resulted in cells (cheX mutant cells) with a distinct motility phenotype. While wild-type cells ran, paused (stopped or flexed), and reversed, the cheX mutant cells continuously flexed and were not able to run or reverse. Furthermore, swarm plate and capillary tube chemotaxis assays demonstrated that cheX mutant cells were deficient in chemotaxis. Wild-type chemotaxis and motility were restored when cheX mutant cells were complemented with a shuttle vector expressing CheX. Furthermore, CheX dephosphorylated CheY3-P in vitro and eluted as a homodimer in gel filtration chromatography. These findings demonstrated that B. burgdorferi CheX is a CheY-P phosphatase that is essential for chemotaxis and motility, which is consistent with CheX being the only CheY-P phosphatase in the B. burgdorferi chemotaxis signal transduction pathway.Bacteria move toward or away from environments that are favorable or unfavorable, respectively, to enhance their survival (reviewed in references 5, 63, and 65). When this movement is in response to chemicals, the process is termed chemotaxis. Flagella or periplasmic flagella, depending upon their location in a cell, are responsible for locomotion in many species of bacteria. Regulation of flagellar rotation and chemotaxis has been studied most extensively in Escherichia coli and Salmonella enterica serovar Typhimurium, and phosphorylation of the response regulator CheY plays an important role in regulating the swimming pattern of cells (reviewed in references 5, 8, 12, 60, 63, and 65). The concentration of phosphorylated CheY (CheY-P) determines whether a cell runs or tumbles. In the absence of attractants, the concentration of CheY-P is relatively high, and CheY-P diffuses to and binds the flagellar switch protein FliM, switching flagellar rotation from a default counterclockwise (CCW) state to a clockwise (CW) rotation. CW rotation of one or more flagella disrupts flagellar bundles, causing cells to tumble and reorient direction during the next run (37, 64). Although CheY-P autodephosphorylates, E. coli CheZ is required for efficient CheY-P dephosphorylation, allowing rapid responses to the environment (53). Thus, functionally reducing CheY-P in null mutants of cheA (encoding the protein that transfers phosphate to CheY) or cheY resu...

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Section: Methodssupporting

confidence: 68%

CheX Is a Phosphorylated CheY Phosphatase Essential forBorrelia burgdorferiChemotaxis

Motaleb¹,

Miller

Li³

et al. 2005

J Bacteriol

112

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“…The following antibodies were kindly provided by other investigators: Blair (University of Utah, Salt Lake City, UT). The specific reactivity of these antibodies to B. burgdorferi CheX, CheY3, Pfs, FlaB, MotB, FliG1, FliG2, and FliM has been reported previously (16,50,52,56,58,60,74,82).…”

Section: Methodssupporting

confidence: 53%

Analysis of the HD-GYP Domain Cyclic Dimeric GMP Phosphodiesterase Reveals a Role in Motility and the Enzootic Life Cycle of Borrelia burgdorferi

et al. 2011

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HD-GYP domain cyclic dimeric GMP (c-di-GMP) phosphodiesterases are implicated in motility and virulence in bacteria. Borrelia burgdorferi possesses a single set of c-di-GMP-metabolizing enzymes, including a putative HD-GYP domain protein, BB0374. Recently, we characterized the EAL domain phosphodiesterase PdeA. A mutation in pdeA resulted in cells that were defective in motility and virulence. Here we demonstrate that BB0374/PdeB specifically hydrolyzed c-di-GMP with a K m of 2.9 nM, confirming that it is a functional phosphodiesterase. Furthermore, by measuring phosphodiesterase enzyme activity in extracts from cells containing the pdeA pdeB double mutant, we demonstrate that no additional phosphodiesterases are present in B. burgdorferi. pdeB single mutant cells exhibit significantly increased flexing, indicating a role for c-di-GMP in motility. Constructing and analyzing a pilZ pdeB double mutant suggests that PilZ likely interacts with chemotaxis signaling. While virulence in needle-inoculated C3H/HeN mice did not appear to be altered significantly in pdeB mutant cells, these cells exhibited a reduced ability to survive in Ixodes scapularis ticks. Consequently, those ticks were unable to transmit the infection to naïve mice. All of these phenotypes were restored when the mutant was complemented. Identification of this role of pdeB increases our understanding of the c-di-GMP signaling network in motility regulation and the life cycle of B. burgdorferi.

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“…Monoclonal and polyclonal antibodies that were kindly provided by other investigators included the following: anti-FlaB (H9724) by A. G. Barbour (University of California, Irvine, CA), anti-FlaA by B. Johnson (Centers for Disease Control and Prevention, Atlanta, GA), anti-DnaK by J. Benach (SUNY, Stony Brook, NY), and polyclonal anti-CheA by P. Matsumura (University of Illinois, Chicago, IL). The specific reactivities of those antibodies to B. burgdorferi FlaB, FlaA, DnaK, and CheA2, respectively, have been reported previously (1,13,33,40,43). The intensity of the CheY3 protein bands in wild-type and complemented cheY3 ϩ was measured using Alpha View spot densitometry (Version 3.0.2.0; Alpha Innotech Corporations).…”

Section: Ry3-f (Ggatccattcaaaagactacaattgc) and Ry3-r (Aagcttt Ttaacamentioning

confidence: 95%

CheY3 of Borrelia burgdorferi Is the Key Response Regulator Essential for Chemotaxis and Forms a Long-Lived Phosphorylated Intermediate

et al. 2011

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Spirochetes have a unique cell structure: These bacteria have internal periplasmic flagella subterminally attached at each cell end. How spirochetes coordinate the rotation of the periplasmic flagella for chemotaxis is poorly understood. In other bacteria, modulation of flagellar rotation is essential for chemotaxis, and phosphorylation-dephosphorylation of the response regulator CheY plays a key role in regulating this rotary motion. The genome of the Lyme disease spirochete Borrelia burgdorferi contains multiple homologues of chemotaxis genes, including three copies of cheY, referred to as cheY1, cheY2, and cheY3. To investigate the function of these genes, we targeted them separately or in combination by allelic exchange mutagenesis. Whereas wild-type cells ran, paused (flexed), and reversed, cells of all single, double, and triple mutants that contained an inactivated cheY3 gene constantly ran. Capillary tube chemotaxis assays indicated that only those strains with a mutation in cheY3 were deficient in chemotaxis, and cheY3 complementation restored chemotactic ability. In vitro phosphorylation assays indicated that CheY3 was more efficiently phosphorylated by CheA2 than by CheA1, and the CheY3-P intermediate generated was considerably more stable than the CheY-P proteins found in most other bacteria. The results point toward CheY3 being the key response regulator essential for chemotaxis in B. burgdorferi. In addition, the stability of CheY3-P may be critical for coordination of the rotation of the periplasmic flagella.Spirochetes are a group of motile bacteria that have a unique morphology and means of motility. On the surface of the spirochete is an outer membrane, which is often referred to as the outer membrane sheath. Within this outer membrane are the cell cylinder and the periplasmic flagella. The periplasmic flagella reside between the outer membrane and the cell cylinder. Because of its medical importance and recent advances in genetic manipulation, we have focused on the Lyme disease spirochete Borrelia burgdorferi to analyze spirochete motility and chemotaxis (for recent reviews, see references 10, 26, 34, and 68). This spirochete is relatively long (10 to 20 m) and thin (0.31 m) and has a flat-wave morphology, and motility is generated by rotation of the periplasmic flagella (11,14,24,25,33,42). Approximately 7 to 11 periplasmic flagella are subterminally attached at each cell end (30), and recent electron cryotomography analysis indicates that these periplasmic flagella form elegant ribbons that wrap clockwise (CW) around the cell cylinder (11). Not only are the periplasmic flagella involved in motility, but these organelles have a skeletal function that in part dictates the flat-wave shape of the cell (10,14,26,35,40,53,68). Thus, mutants that lack periplasmic flagella are nonmotile and have a rod-shaped morphology (35,40,53). Motility is accomplished by backward-moving flat waves along the cell body. These waves are generated by the coordinated rotation of the rigid periplasmic flagella as the...

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Characterization of Antigenic Determinants of Borrelia burgdorferi Shared by Other Bacteria

Cited by 66 publications

References 48 publications

CheX Is a Phosphorylated CheY Phosphatase Essential forBorrelia burgdorferiChemotaxis

CheX Is a Phosphorylated CheY Phosphatase Essential forBorrelia burgdorferiChemotaxis

Analysis of the HD-GYP Domain Cyclic Dimeric GMP Phosphodiesterase Reveals a Role in Motility and the Enzootic Life Cycle of Borrelia burgdorferi

CheY3 of Borrelia burgdorferi Is the Key Response Regulator Essential for Chemotaxis and Forms a Long-Lived Phosphorylated Intermediate

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