Borrelia burgdorferi rel Is Responsible for Generation of Guanosine-3′-Diphosphate-5′-Triphosphate and Growth Control

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Abstract
The global transcriptional regulator (p)ppGpp (guanosine-3′-diphosphate-5′-triphosphate and guanosine-3′,5′-bisphosphate, collectively) produced by the relA and spoT genes in Escherichia coli allows bacteria to adapt to different environmental stresses. The genome of Borrelia burgdorferi encodes a single chromosomal rel gene (BB0198) ( B. burgdorferi rel [ rel Bbu ]) homologous to relA and spoT of E. coli . Its role in (p)ppGpp synthesis, bacterial growth, and modulation of gene expression has not been studied in detail. We constructed a rel Bbu deletion mutant in an infectious B. burgdorferi 297 strain and isolated an extrachromosomally complemented derivative of this mutant. The mutant did not synthesize rel Bbu mRNA, Rel Bbu protein, or (p)ppGpp. This synthesis was restored in the complemented derivative, confirming that rel Bbu is necessary and sufficient for (p)ppGpp synthesis and degradation in B. burgdorferi . The rel Bbu mutant grew well during log phase in complete BSK-H but reached lower cell concentrations in the stationary phase than the wild-type parent, suggesting that (p)ppGpp may be an important factor in the ability of B. burgdorferi to adapt to stationary phase. Deletion of rel Bbu did not eliminate the temperature-elicited OspC shift, nor did it alter bmp gene expression or B. burgdorferi antibiotic susceptibility. Although deletion of rel Bbu eliminated B. burgdorferi virulence for mice, which was not restored by complementation, we suggest that rel Bbu -dependent accumulation of (p)ppGpp may be important for in vivo survival of this pathogen.