Pathogen-host interactions in Lyme disease interrogated at Single Cell Level

莱姆病是美国最常见的节肢动物传播疾病。目前还没有疫苗

Lyme disease is the most common arthropod-borne illness in the US. Currently, there are no vaccines available to prevent human Lyme disease and antibacterial therapy is the treatment of choice. Borrelia burgdorferi, the agent of Lyme disease, is maintained in nature through an enzootic cycle involving a tick vector and a variety of vertebrate hosts. B. burgdorferi undergoes extensive remodeling of its cell envelope in response to environmental cues prevalent in these highly divergent hosts. The molecular interactions between the borrelial envelope or its major determinants, such as peptidoglycan (PG) and surface-exposed lipoproteins, with host cells play a critical role in the survival and infectivity of B. burgdorferi during different stages of its infectious cycle. Moreover, B. burgdorferi is an extreme auxotroph lacking metabolic pathways for biosynthesis of essential biomolecules and is therefore a highly host-adapted pathogen. In addition, the peptidoglycan (PG) of B. burgdorferi has a unique composition with L-Ornithine as the branching amino acid in its pentapeptide cross-linking the N-acetyl glucosamine and N-acetyl muramic acid polymers, which is distinct from the pentapeptides present in the PG of Gram-negative and Gram-positive bacterial pathogens. Two mutants strains of B. burgdorferi lacking Borrelia host-adaptation Regulator (BadR) or Borrelia host-adaptation Protein (BadP) hyper- and hypo-express lipoproteins, respectively, serving as ideal genetic tools to interrogate pathogen-host cell interactions mediated by differences in levels of surface-exposed lipoproteins. The long- term objective of the proposed research is to disrupt the complex lifestyle of Lyme spirochetes within ticks and mammalian hosts to reduce incidence of Lyme disease. In this proposal, we will test the central hypothesis that remodeling of borrelial cell envelope alters host antimicrobial responses influencing survival of B. burgdorferi in different hosts. The central hypothesis will be tested with two specific aims: Specific Aim 1: To determine signaling effects of borrelial PG/lipoproteins on tick and mammalian cells using single cell RNA-Seq (scRNA- Seq) analysis. Large arrays of transcriptional data will be analyzed and validated to identify pathways and cellular phenotypes that distinguish response of single cells from transmission, reservoir and dead-end hosts exploiting the advantages of using small numbers of host cells, spirochetes and minuscule levels of purified borrelial determinants. Role of Peptidoglycan Recognition Proteins and Hippo signaling pathway in modulating Bb infection will be interrogated. Specific Aim 2: To define effects of intact spirochetes defective in PG/lipoprotein composition on primary host cells at single-cell level. Mutant spirochetes defective in PG stability/synthesis or with altered levels of lipoproteins will be used to define their interactions with host cells using RNA-Seq analysis and further validated for colonization capabilities using the experimental mouse models of Lyme disease. The impact of this innovative study of pathogen-host interactions at single-cell level will enable identification of novel signaling pathways critical for colonization/transmission of Lyme spirochetes.