Genetic profiling of the isoprenoid and sterol biosynthesis pathway genes of Trypanosoma cruzi.

Raúl O Cosentino,Fernán Agüero,Mauricio Martins Rodrigues

doi:10.1371/journal.pone.0096762

Abstract

In Trypanosoma cruzi the isoprenoid and sterol biosynthesis pathways are validated targets for chemotherapeutic intervention. In this work we present a study of the genetic diversity observed in genes from these pathways. Using a number of bioinformatic strategies, we first identified genes that were missing and/or were truncated in the T. cruzi genome. Based on this analysis we obtained the complete sequence of the ortholog of the yeast ERG26 gene and identified a non-orthologous homolog of the yeast ERG25 gene (sterol methyl oxidase, SMO), and we propose that the orthologs of ERG25 have been lost in trypanosomes (but not in Leishmanias). Next, starting from a set of 16 T. cruzi strains representative of all extant evolutionary lineages, we amplified and sequenced ∼24 Kbp from 22 genes, identifying a total of 975 SNPs or fixed differences, of which 28% represent non-synonymous changes. We observed genes with a density of substitutions ranging from those close to the average (∼2.5/100 bp) to some showing a high number of changes (11.4/100 bp, for the putative lathosterol oxidase gene). All the genes of the pathway are under apparent purifying selection, but genes coding for the sterol C14-demethylase, the HMG-CoA synthase, and the HMG-CoA reductase have the lowest density of missense SNPs in the panel. Other genes (TcPMK, TcSMO-like) have a relatively high density of non-synonymous SNPs (2.5 and 1.9 every 100 bp, respectively). However, none of the non-synonymous changes identified affect a catalytic or ligand binding site residue. A comparative analysis of the corresponding genes from African trypanosomes and Leishmania shows similar levels of apparent selection for each gene. This information will be essential for future drug development studies focused on this pathway.

Highlights

Trypanosoma cruzi, a protozoan parasite of the order Kinetoplastida, is the causative agent of Chagas Disease, a neglected disease that is endemic in South America, affecting in excess of 8 million people [1]
Filling pathway holes: genes involved in sterol biosynthesis in T. cruzi
To analyze the genetic diversity of the T. cruzi sterol biosynthesis pathway (SBP) we decided to sequence all enzymes of the pathway, starting from enzymes that produce the terpenoid backbone precursors, and going down to the last enzyme that produces ergosterol as a product

Summary

Introduction

Trypanosoma cruzi, a protozoan parasite of the order Kinetoplastida, is the causative agent of Chagas Disease, a neglected disease that is endemic in South America, affecting in excess of 8 million people [1]. There are no general agreement regarding the estimated time of divergence of the six T. cruzi lineages; recent estimations suggest that major lineages (excluding the hybrid lineages TcV and TcVI) diverged 1–4 millions of years ago [9,10] while the hybrid lineages emerged much more recently (less than 1 million years ago, according to Flores-Lopez and Machado [9], and within the last 60,000 years, according to Lewis MD, et al [10]) This divergence was accompanied with a diversification of phenotypic and biological properties. Several investigations suggest that the observed diversity in host preference, cell tropism and drug susceptibility might be properties of different strains and/or lineages [11,12,13,14]

Objectives

Methods

Results

Discussion

Conclusion