Coping with stress: How Reactive Oxygen and Nitrogen Species alter the transcriptional landscape of the Lyme disease spirochete Borrelia burgdorferi

Abstract

Borrelia burgdorferi, the Lyme disease spirochete, modulates expression of distinct sets of lipoprotein-encoding genes to successfully infect its tick vector Ixodes scapularis and various mammalian hosts. Lipoprotein expression is controlled by a limited number of transcription factors in response to environmental challenges encountered during infection, including changes in pH, temperature, osmolarity, and nutrient availability. Previous work from our laboratory demonstrated B. burgdorferi encounters host-derived reactive oxygen species (ROS) and reactive nitrogen species (RNS) during infection of I. scapularis ticks, which we hypothesize also serve as signals sensed by B. burgdorferi to modulate gene expression. We tested this hypothesis by determining genes differentially regulated by B. burgdorferi in response to sublethal concentrations of ROS-producing hydrogen peroxide (H2O2) and the RNS-generating compound spermine NONOate (sper/NO) with and without a pH shift via RNA-seq and reverse transcriptase-quantitative PCR (RT-qPCR). We probed for translational changes in targeted proteins via western blotting. Genes regulated by RpoS (including those associated with mammalian infection such as ospC, dbpA, and bba66) were downregulated in response to sublethal concentrations of H2O2 (25 µM) and sper/NO (0.625 mM) coupled with a pH shift—the expression patterns were less defined in the absence of a pH shift. Additionally, we observed elevated expression of bbd18 by RT-qPCR and increased concentrations of BBD18 protein through western blot analysis in B. burgdorferi exposed to H2O2 and sper/NO with and without a pH shift, but NO more strongly induced BBD18 as compared to H2O2 regardless of pH shift or lack thereof. These observations, coupled with our data, suggest ROS and RNS serve as signals sensed by B. burgdorferi to navigate the enzootic cycle. As tight regulation of RpoS and its associated genes enable B. burgdorferi to complete the enzootic cycle, ROS/RNS could serve as crucial negative regulating signals when the bacteria transitions between the mammalian host and tick vector—there is a lot of published data about what signals stimulate the RpoS cascade, but it is still not known what signals cause suppression of the RpoS cascade to our knowledge. With that in mind, because RpoS is essential for mammalian infection, understanding the mechanisms behind negative regulation of RpoS and its associated genes may provide targets for future Lyme disease intervention and treatment strategies.