Profiling of Host Immune Responses to Leishmania Parasites and Phlebotomine Arthropod Vectors via Investigations of microRNA Regulation and Acquired Immunity

Leishmaniasis is a group of sand fly vector-borne parasitic diseases which exhibit clinical pathologies specific to the species of causative intracellular parasites. Visceral and cutaneous leishmaniasis can be caused Leishmania donovani and Leishmania major parasites, respectively. Macrophages (MPs) and Dendritic Cells (DCs) are host cell types for Leishmania. These parasites employ immune evasion strategies in order to infect their vertebrate hosts that may include advantageous microenvironments resulting from the effects of sand fly salivary components at the site of inoculation in the host, or parasite influences on host cell transcriptional and translational molecular pathways. The host:vector interface was examined through analyses of anti-sand fly salivary gland proteins using blood plasma antibodies from persons residing in differentially endemic regions of Phlebotomus papatasi vectors. Five major salivary proteins were immunogenic, which permitted the clustering of blood donors based on antibody specificity corresponding to the endemic characteristics of their native regions. The host:pathogen interface was examined through investigations of host microRNA (miRNA) expression and quantitative associations with predicted transcript targets. Mature miRNAs are short non-coding single stranded RNAs that associate with Argonaute 2 (AGO2) in humans. The miRNAs complement to RNA transcripts to block protein translation. An analysis of mature miRNA expression in MPs and DCs infected with L. donovani or L. major found that most miRNAs were down-regulated compared to uninfected cells. However, L. donovani-infected DC miRNAs were, on average, more highly expressed compared to L. major-infected DCs. A second temporal study was performed with L. donovani and L. major-infected DCs, where mature miRNAs were co-immunoprecipitated with AGO2 and quantified by RNAseq. Concurrently, gene transcript expression from paired samples was assessed by microarrays. The multi-dimensional datasets were compared through miRNA:target prediction guided-correlations. In examining mixed regulatory circuits among significant correlations, miR-155 was found to target SMAD1. Target reporter experiments confirmed miR-155's ability to target SMAD1, while quantitative Western blot analysis found a greater down-regulation of SMAD1 protein in L. donovani infection samples compared to L. major samples. The work presented herein makes substantial contributions to our understanding of the host cell immune responses to both Leishmania parasites and their sand fly vectors.