Modulation of Borrelia burgdorferi Stringent Response and Gene Expression during Extracellular Growth with Tick Cells

Abstract

Borrelia burgdorferi N40 multiplied extracellularly when it was cocultured with tick cells in L15BS medium, a medium which by itself did not support B. burgdorferi N40 growth. Growth of B. burgdorferi N40 in the presence of tick cells was associated with decreased production of (p)ppGpp, the stringent response global regulator, a fourfold decrease in relA/spoT mRNA, an eightfold net decrease in bmpD mRNA, and a fourfold increase in rpsL-bmpD mRNA compared to growth of B. burgdorferi in BSK-H medium. As a result, the polycistronic rpsL-bmpD mRNA level increased from 3 to 100% of the total bmpD message. These observations demonstrate that there are reciprocal interactions between B. burgdorferi and tick cells in vitro and indicate that the starvation-associated stringent response mediated by (p)ppGpp present in B. burgdorferi growing in BSK-H medium is ameliorated in B. burgdorferi growing in coculture with tick cell lines. These results suggest that this system can provide a useful model for identifying genes controlling interactions of B. burgdorferi with tick cells in vitro when it is coupled with genetic methods to isolate and complement B. burgdorferi mutants.Borrelia burgdorferi, the cause of Lyme disease, modulates its gene expression in the presence of host cells. As an example of this modulation, the relative expression of the ospA and ospC genes is altered as B. burgdorferi passes from a tick to a mammalian host (32); a similar down-modulation-up-modulation of OspA and OspC is observed during coculture of B. burgdorferi with tick cells in vitro (27). This oscillation in gene expression may be mediated by a regulatory loop involving changes in the expression of 54 and S , suggesting that B. burgdorferi growing in the presence of tick cells may undergo a series of as-yet-uncharacterized global regulatory changes (36). It also suggests that study of the reciprocal interactions between B. burgdorferi and tick cells could illuminate aspects of gene modulation in B. burgdorferi and the genes and gene products involved (17,36).Flexible adaptation by bacterial pathogens to rapidly changing host environments requires the interplay of regulators generated in response to signals triggered by environmental stimuli. One such adaptive response is the stringent response, a global response triggered by nutritional stress when protein synthesis is blocked by amino acid starvation (10, 13, 34). The stringent response causes up-regulation of protein degradation and amino acid synthesis, down-regulation of nucleic acid and protein synthesis, and alteration of many cellular functions, including transcription, translation, replication, transport, and metabolism, and it is accompanied by slow growth (10, 34). In Escherichia coli, the stringent response involves RelA, a synthase, and SpoT, a hydrolase, which synthesize and degrade, respectively, ppGpp and pppGpp [(p)ppGpp], the alarmon (master regulator, second message) that is the proximate mediator of this response (9, 10). In some bacterial pathogens, such as Mycobacterium t...

Localization of BmpA on the Exposed Outer Membrane ofBorrelia burgdorferiby Monospecific Anti-Recombinant BmpA Rabbit Antibodies

“…Chromosomally encoded BmpA and the other Bmp proteins appear to be less prone to recombination than plasmid-encoded proteins (7), and the temperature-and pH-independent expression of bmpA mRNA (15) and BmpA (J. J. Shin and F. C. Cabello, unpublished data) suggest that its expression is not controlled by the RpoNRpoS regulatory pathway, which appears to control expression of some plasmid-encoded lipoproteins (25,46). However, previous experiments in our laboratory have indicated that modulation of the expression of the bmp gene cluster is achieved by growing B. burgdorferi cells in the presence of tick cells (5), suggesting that the expression of these genes may be modulated by alternative stimuli and pathways. The surface location of BmpA lipoprotein, the growth inhibition of B. burgdorferi by anti-rBmpA antibodies, the conservation of the bmpA gene in different B. burgdorferi sensu lato strains (20), and the consistent detection of bmpA expression and BmpA at different B. burgdorferi cell densities, temperatures, and pH and in different B. burgdorferi sensu lato strains (15 and Shin and Cabello, burgdorferi.…”

Tick cell lines: tools for tick and tick-borne disease research

“…Tick cells provided an important environment for studies on stage-specific gene transcription and protein expression in B. burgdorferi; co-cultivation of spirochaetes with IDE8 and ISE6 cells influenced temperature-dependent outer surface protein expression associated with increased infectivity of spirochaetes for the mammalian host [40] and modulated transcription of genes involved with the starvation-associated stringent response [41]. Similarly, temperature-dependent protein expression was observed in R. rickettsii propagated in IDE2 and Dermacentor albipictus DALBE3 cells at both 28 8C and 34 8C [42]; the antigen profile in tick cells at 34 8C was similar to that seen in mammalian cells, whereas several proteins of unknown function present at the higher temperature were apparently not expressed at 28 8C.…”

ppGpp Conjures Bacterial Virulence

“…The alarmone likely contributes to nutritional adaptation, since during the growth of B. burgdorferi in medium with serum or in the presence of tick cells, the levels of both ppGpp and spoT (called rel Bbu ) mRNAs decrease (34). Conversely, spoT transcription is induced when spirochetes are cultured in medium supplemented with tick saliva (47).…”