Hypothetical Protein BB0569 Is Essential for Chemotaxis of the Lyme Disease Spirochete Borrelia burgdorferi

Abstract

The Lyme disease spirochete Borrelia burgdorferi has five putative methyl-accepting chemotaxis proteins (MCPs). In this report, we provide evidence that a hypothetical protein, BB0569, is essential for the chemotaxis of B. burgdorferi. While BB0569 lacks significant homology to the canonical MCPs, it contains a conserved domain (spanning residues 110 to 170) that is often evident in membrane-bound MCPs such as Tar and Tsr of Escherichia coli. Unlike Tar and Tsr, BB0569 lacks transmembrane regions and recognizable HAMP and methylation domains and is similar to TlpC, a cytoplasmic chemoreceptor of Rhodobacter sphaeroides. An isogenic mutant of BB0569 constantly runs in one direction and fails to respond to attractants, indicating that BB0569 is essential for chemotaxis. Immunofluorescence, green fluorescent protein (GFP) fusion, and cryo-electron tomography analyses demonstrate that BB0569 localizes at the cell poles and is required for chemoreceptor clustering at the cell poles. Protein cross-linking studies reveal that BB0569 forms large protein complexes with MCP3, indicative of its interactions with other MCPs. Interestingly, analysis of B. burgdorferi mcp mutants shows that inactivation of either mcp 2 or mcp 3 reduces the level of BB0569 substantially and that such a reduction is caused by protein turnover. Collectively, these results demonstrate that the domain composition and function of BB0569 are similar in some respects to those of TlpC but that these proteins are different in their cellular locations, further highlighting that the chemotaxis of B. burgdorferi is unique and different from the Escherichia coli and Salmonella enterica paradigm. IMPORTANCESpirochete chemotaxis differs substantially from the Escherichia coli and Salmonella enterica paradigm, and the basis for controlling the rotation of the bundles of periplasmic flagella at each end of the cell is unknown. In recent years, Borrelia burgdorferi, the causative agent of Lyme disease, has been used as a model organism to understand spirochete chemotaxis and its role in infectious processes of the disease. In this report, BB0569, a hypothetical protein of B. burgdorferi, has been investigated by using an approach of genetic, biochemistry, and cryo-electron tomography analyses. The results indicate that BB0569 has a distinct role in chemotaxis that may be unique to spirochetes and represents a novel paradigm. Chemotaxis allows bacteria to swim toward favorable environments or away from harmful ones by modulation of their swimming behavior (1, 2). The molecular mechanisms involved in bacterial chemotaxis have been extensively studied in two prototype organisms, Escherichia coli and Salmonella enterica (for recent reviews, see references 3-5). The chemotaxis signaling apparatus works as a supramolecular unit that is composed of three major components: methyl-accepting chemotaxis proteins (MCPs), the histidine kinase CheA, and the response regulator CheY (6, 7). MCPs sense various environmental and intracellular signals and control the acti...

A chemosensory-like histidine kinase is dispensable for chemotaxis in vitro but regulates the virulence of Borrelia burgdorferi through modulating the stability of RpoS

“…The current evidence has revealed that all of the chemotaxis genes in the cheA 2 cluster are required for chemotaxis [11][12][13]. In contrast, the genes in the cheW 2 cluster that have been studied (with the exception of bb0569, which encodes for an atypical MCP [14]) are not involved in chemotaxis in vitro. For instance, cheW 2 and cheY 2 mutants have a similar in vitro phenotype as the wild type with no apparent defect in chemotactic function [11,13,15,16].…”

Borrelia burgdorferi Keeps Moving and Carries on: A Review of Borrelial Dissemination and Invasion

“…The number of chemotaxis homologs encoded in the B. burgdorferi genome is greater and far more complex than other motile pathogens, such as E. coli or S. enterica ( 13 , 14 ). Additional genes, csrA and bb0569 , influence the motility regulation of B. burgdorferi ( 58 , 59 ). Carbon storage regulator, CsrA, is involved in posttranscriptional regulation of flagellar genes and contributes to cell morphology ( 59 , 60 ).…”

“…Carbon storage regulator, CsrA, is involved in posttranscriptional regulation of flagellar genes and contributes to cell morphology ( 59 , 60 ). BB0569, annotated as a hypothetical protein, has sequence similarity to MCPs and was found to localize to the poles of the borrelial cell and important for motility and chemotaxis ( 58 ). B. burgdorferi motility is regulated throughout the enzootic cycle with the distinct behavior of intermittent swimming in the tick vector that includes periods of a non-motile state relative to the persistent swimming observed in the murine host ( 44 , 61 ).…”

Fluorescent Proteins, Promoters, and Selectable Markers for Applications in the Lyme Disease Spirochete Borrelia burgdorferi

“…Constitutive gene expression is mostly limited to the use of very strong promoters. Moreover, apart from a few exceptions ( 14 – 19 ), fluorescent protein reporters have primarily been used as gene expression reporters or as cellular stains for in vivo localization of the spirochete ( 13 ). Yet, fluorescent proteins have many more uses which have transformed the field of cell biology ( 20 ).…”