The Intergenic Small Non-Coding RNA ittA is Required for Optimal Infectivity and Tissue Tropism in Borrelia burgdorferi
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ABSTRACTPost-transcriptional regulation via small regulatory RNAs (sRNAs) has been implicated in diverse regulatory processes in bacteria, including virulence. One class of sRNAs, termedtrans-acting sRNAs, can affect the stability and/or the translational efficiency of regulated transcripts. In this study, we utilized a collaborative approach that employed data from infection with theBorrelia burgdorferiTn library, coupled with Tn-seq, together with borrelial sRNA and total RNA transcriptomes, to identify an intergenictrans-acting sRNA, which we designate here asittAforinfectivity-associated andtissue-tropic sRNA locusA. The genetic inactivation ofittAresulted in a significant attenuation in infectivity, with decreased spirochetal load in ear, heart, skin and joint tissues. In addition, theittAmutant did not disseminate to peripheral skin sites or heart tissue, suggesting a role forittAin regulating a tissue-tropic response. RNA-Seq analysis determined that 19 transcripts were differentially expressed in theittAmutant relative to its genetic parent, includingvraA, bba66,ospDandoms28(bba74). Subsequent proteomic analyses also showed a significant decrease of OspD and Oms28 (BBA74) proteins. To our knowledge this is the first documented intergenic sRNA that alters the infectivity potential ofB. burgdorferi.AUTHOR SUMMARYLyme disease is a tick-borne infection mediated by the spirochetal bacterium,Borrelia burgdorferi, that is responsible for greater than 300,000 infections in the United States per year. As such, additional knowledge regarding how this pathogen modulates its regulatory armamentarium is needed to understand howB. burgdorferiestablishes and maintains infection. The identification and characterization of small, non-coding RNA molecules in living systems, designated as sRNAs, has recalibrated how we view post-transcriptional regulation. Recently, over 1,000 sRNAs were identified inB. burgdorferi. Despite the identification of these sRNAs, we do not understand how they affect infectivity orB. burgdorferipathogenesis related outcomes. Here, we characterize theittA B. burgdorferisRNA and show that it is essential for optimal infection using murine experimental infection as our readout. We also track the effect of this sRNA on the transcriptional and proteomic profile as the first step in providing mechanistic insight into how this important sRNA mediates its regulatory effect.
