Investigating CovRS activation within skin microenvironments to drive heterogenicity of Streptococcus pyogenes gene expression

摘要 化脓性链球菌感染是美国医疗保健系统的主要负担， 全球范围内。大多数感染是轻微的和浅表的，如咽炎和脓疱病。然而，快速的疾病 进展可能导致危及生命的侵袭性感染，如坏死性筋膜炎和中毒性休克。 综合症。此外，康复的患者经常遭受感染后的慢性并发症，包括急性 风湿性心脏病。化脓性链球菌感染和感染后并发症总共导致超过 全球每年有60万人死亡。目前，还没有针对化脓性链球菌的疫苗，也没有大量的 发病率和死亡率突出表明迫切需要替代治疗战略。与大多数病原体不同，S。 化脓菌通过表达促进免疫逃避的毒力因子在炎性环境中茁壮成长， 这通常与侵袭性感染有关。大约15%的基因组，包括大多数毒力 因子，由一个称为CovRS(或CsrRs)的两组分系统进行调节。CovRS检测到主肽LL- 37，这是毒力因子调节的有效信号。然而，目前还不清楚这种变化是如何发生的 在LL-37中，由于细胞死亡导致的可用性改变了感染进展期间的CovRS调节。此外， 化脓性链球菌的不同毒素在CovRs“开/关”活性状态下表达，但在感染过程中都是必不可少的。 这表明处于不同CovR“开/关”状态的遗传相同细菌的亚群可能会释放 在感染的不同阶段和不同的微环境中的不同毒素 在发病机制中起作用。我假设异质性的CovRS调控是由L1-1的变异控制的。 37水平由于炎性细胞死亡，允许产生化脓性链球菌所需的异源毒素 感染。我将通过1)使用寄主和病原体内的遗传学技术来验证我的假设 炎性细胞死亡影响IL-37的释放和CovRS的调节；2)单细胞的显微镜观察 分析局部IL-37浓度对工程化脓性链球菌CovRS信号的影响 记者菌株。总的来说，这些研究将提供对化脓性链球菌的致病机制的洞察。 诱导炎症并确定延缓疾病进展的新药物靶点。

ABSTRACT Streptococcus pyogenes infections are a major burden to the healthcare system in the United States and globally. Most infections are mild and superficial, such as pharyngitis and impetigo. However, rapid disease progression may result in life-threatening invasive infections, such as necrotizing fasciitis and toxic shock syndrome. Furthermore, recovered patients often suffer post-infection chronic complications, including acute rheumatic heart disease. Altogether, S. pyogenes infections and post-infection complications result in more than 600,000 global deaths annually. Currently, there is no vaccine against S. pyogenes and high instances of morbidity and mortality highlight the urgent need for alternative treatment strategies. Unlike most pathogens, S. pyogenes thrives in inflammatory environments by expressing virulence factors that promote immune evasion, which is often associated with invasive infections. Approximately 15% of the genome, including most virulence factors, is regulated by a two-component system called CovRS (or CsrRS). CovRS detects the host peptide LL- 37, which functions as a potent signal for virulence factor regulation. However, it remains unclear how changes in LL-37 availability due to cell death alters CovRS regulation during the progression of an infection. Further, distinct toxins of S. pyogenes are expressed in CovRS “on/off” activity states, yet all are essential during infection. This indicates that sub-populations of genetically identical bacteria in different CovRS “on/off” states may release varying toxins during different stages of infection and within distinct microenvironments that collectively contribute to pathogenesis. I hypothesize that heterogenous CovRS regulation is controlled by variations of LL- 37 levels due to inflammatory cell death, allowing heterogenous toxin production necessary for S. pyogenes infections. I will test my hypothesis by 1) using genetics techniques within the host and pathogen to identify how inflammatory cell death impacts LL-37 release and CovRS regulons, and 2) using microscopy for single cell analysis to examine how local LL-37 concentration contributes to CovRS signaling in engineered S. pyogenes reporter strains. Collectively, these studies will provide insight into the mechanisms responsible for S. pyogenes- induced inflammation and identify novel drug targets that stall disease progression.