Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

A polyomic approach to understand the molecular basis of virulence in Leishmania (#445)

Richard Burchmore 1 , Abduabset Kabli 1 , Kathryn Crouch 1 , Michael Barrett 1
  1. University of Glasgow, Glasgow, SCOTLAND, United Kingdom

Leishmania have the ability to subvert the host immune system and adopt sophisticated strategies to develop and survive within the mammalian host. Leishmania parasites survive within macrophages, inside a parasitophorous vacuole. The molecular communication between host and parasite decides the outcome of infection, but is incompletely understood. We have compared genotype and phenotype of an attenuated Leishmania mexicanaline with a virulent, isogenic wild type precursor. We aim to identify key virulence factors and to explore the potential of the attenuated line as a vaccine candidate.

This study was conducted on promastigotes of Leishmania mexicanaand involvedcomparative polyomics approaches to identify the molecules that contribute to Leishmania virulence. Log phase promastigotes of wild-type and gentamicin-attenuated (H-line) were grown in parallel in media containing 10% FBS. For comparative proteomic analysis, protein extracts were labelled using 6-plex TMT and analyzed with LC-MS/MS. For metabolomics, metabolites were extracted with Chloroform/ Methanol/ Water (1:3:1) and analyzed with LC-MS. For transcriptomics, RNA was isolated and converted into a library of cDNA molecules for cluster generation and DNA sequencing. We found 18 proteins differentially expressed in attenuated Leishmania (FC ≥1.5 and FDR ≤0.05) and 26 identified metabolites (FC ≥1.5 and FDR≤0.05), whereas transcriptomics data found 481 transcripts were differentially expressed (FC ≥1.5 and FDR ≤0.05). Most of the differentially expressed proteins and transcripts were metabolic enzymes, and the majority of the differentially expressed metabolites were substrates that involved in nucleotide, carbohydrate and amino acid metabolisms. Correlation of polyomics datasetsreveals that nucleotide metabolism (pyrimidine and purine) is significantly altered in gentamicin-attenuated Leishmania. Furthermore, nucleobase and nucleoside transporters were significantly down regulated in proteomics analysis. Modulation of gene expression may relate to gentamicin selection. ΔNT3 cells became more sensitive to allopurinol (purine analogue) compared to wild-type cells (EC50= 141.1), suggesting that this may contribute to Leishmania virulence.