Comparative transcriptome profiling of virulent and non-virulent Trypanosoma cruzi underlines the role of surface proteins during infection

Abstract

Trypanosoma cruzi, the protozoan that causes Chagas disease, has a complex life cycle involving several morphologically and biochemically distinct stages that establish intricate interactions with various insect and mammalian hosts. It has also a heterogeneous population structure comprising strains with distinct properties such as virulence, sensitivity to drugs, antigenic profile and tissue tropism. We present a comparative transcriptome analysis of two cloned T. cruzi strains that display contrasting virulence phenotypes in animal models of infection: CL Brener is a virulent clone and CL-14 is a clone that is neither infective nor pathogenic in in vivo models of infection. Gene expression analysis of trypomastigotes and intracellular amastigotes harvested at 60 and 96 hours post-infection (hpi) of human fibroblasts revealed large differences that reflect the parasite’s adaptation to distinct environments during the infection of mammalian cells, including changes in energy sources, oxidative stress responses, cell cycle control and cell surface components. While extensive transcriptome remodeling was observed when trypomastigotes of both strains were compared to 60 hpi amastigotes, differences in gene expression were much less pronounced when 96 hpi amastigotes and trypomastigotes of CL Brener were compared. In contrast, the differentiation of the avirulent CL-14 from 96 hpi amastigotes to extracellular trypomastigotes was associated with considerable changes in gene expression, particularly in gene families encoding surface proteins such as trans-sialidases, mucins and the mucin associated surface proteins (MASPs). Thus, our comparative transcriptome analysis indicates that the avirulent phenotype of CL-14 may be due, at least in part, to a reduced or delayed expression of genes encoding surface proteins that are associated with the transition of amastigotes to trypomastigotes, an essential step in the establishment of the infection in the mammalian host. Confirming the role of members of the trans-sialidase family of surface proteins for parasite differentiation, transfected CL-14 constitutively expressing a trans-sialidase gene displayed faster kinetics of trypomastigote release in the supernatant of infected cells compared to wild type CL-14.

Transmigration of Trypanosoma cruzi trypomastigotes through 3D cultures resembling a physiological environment

“…Trypanosoma cruzi presents a high genetic heterogeneity and, currently, T. cruzi strains are classified into six clusters or discrete typing units (DTUs), named TcI to TcVI [28]. We selected CL Brener (DTU TcVI) and SylvioX10 (DTU TcI) strains, of high and low virulence, respectively, and whose biologically distinctive behavior in experimental models of T. cruzi infection is well characterized [8,29–31].…”

ATrypanosoma cruziZinc Finger protein that controls expression of epimastigote specific genes and affects metacyclogenesis

“…We have previously reported transcriptome profiling analyses comparing trypomastigotes and intracellular amastigotes from T. cruzi, revealing distinct virulence phenotypes and essential factors involved in the parasite's adaptation to the mammalian host (Belew et al, 2017). Extending our RNA-seq data analyses to include the epimastigote stage ( Supplementary Table 1), we compared here the transcriptome profiles obtained from tissue culture-derived trypomastigotes and intracellular amastigotes obtained 60 hours post-infection (hpi) with the transcriptome obtained from in vitro cultured epimastigotes of the CL Brener cloned strain.…”

“…Because post-transcriptional mechanisms mediated by RBPs play a major role in controlling gene expression in T. cruzi, we searched for RBPs that may act as regulators of epimastigote genes by comparing transcript levels of all RBP genes between epimastigotes and the two other forms of the parasite. Although 253 T. cruzi sequences encoding RBPs were previously identified in the CL Brener genome (Belew et al, 2017), homology searches based on protein domains that included additional motifs known to be present in RBPs resulted in the identification of 297 sequences that correspond to 175 RBP genes ( Supplementary Table 4). As indicated in our previous work, because the CL Brener strain has a hybrid genome (El-Sayed et al, 2005), for most genes, we identified two distinct alleles, corresponding to the Esmeraldo and the non-Esmeraldo haplotypes.…”

“…Differential gene expression analyses were performed between epimastigotes samples and tissue cultured trypomastigotes or intracellular amastigotes samples from the RNA-Seq data published by Belew et al (2017). A total of 22014 genes passed the low counts filter (applied to the raw counts of all 25099 genes deposited in CL Brener genome v.38) and the differential expression analysis was performed using the DESeq2 package (Love et al, 2014).…”

Evolution of protein trafficking in kinetoplastid parasites: Complexity and pathogenesis

“…In the insect stage, T. brucei remodels its entire surface, characterized by the replacement of VSGs by procyclins that are important for persistence within the insect vector . In contrast, T. cruzi invades the cells of its host, and its own surface is dominated by mucin‐like molecules that defend the parasite from immune and proteolytic attack and are persistent through the life cycle . There are also multiple additional proteins present within the T. cruzi mucin coat performing adhesion functions during invasion of host cells.…”