Transcriptional and phenotypical heterogeneity of Trypanosoma cruzi cell populations

Abstract

Trypanosoma cruzi has a complex life cycle comprising pools of cell populations which circulate among humans, vectors, sylvatic reservoirs and domestic animals. Recent experimental evidence has demonstrated the importance of clonal variations for parasite population dynamics, survival and evolution. By limiting dilution assays, we have isolated seven isogenic clonal cell lines derived from the Pan4 strain of T. cruzi. Applying different molecular techniques, we have been able to provide a comprehensive characterization of the expression heterogeneity in the mucin-associated surface protein (MASP) gene family, where all the clonal isogenic populations were transcriptionally different. Hierarchical cluster analysis and sequence comparison among different MASP cDNA libraries showed that, despite the great variability in MASP expression, some members of the transcriptome (including MASP pseudogenes) are conserved, not only in the life-cycle stages but also among different strains of T. cruzi. Finally, other important aspects for the parasite, such as growth, spontaneous metacyclogenesis or excretion of different catabolites, were also compared among the clones, demonstrating that T. cruzi populations of cells are also phenotypically heterogeneous. Although the evolutionary strategy that sustains the MASP expression polymorphism remains unknown, we suggest that MASP clonal variability and phenotypic heterogeneities found in this study might provide an advantage, allowing a rapid response to environmental pressure or changes during the life cycle of T. cruzi.

The C-terminal region of Trypanosoma cruzi MASPs is antigenic and secreted via exovesicles

“…The sequencing of the different isolated C-term clones and western blot analysis of trypomastigote EVs using anti-MASP C-term antibodies revealed the simultaneous co-expression of several members of this family in EVs, which is in agreement with the high diversity of expression of MASP proteins in T. cruzi 48 . Thus, we can conclude that the humoral response to the C-term region is not due to the expression of individual MASP members but to the simultaneous expression of numerous MASP proteins, which in turn allows presumption that the molar ratio of the antigen exposed to the immune system is very high.…”

High-resolution profiling of linear B-cell epitopes from mucin-associated surface proteins (MASPs) of Trypanosoma cruzi during human infections

“…In the case of MASP antigenic motifs, which are differentially distributed among different members of a large family of genes, a second assumption should be made: a rather similar probability of expression of different ‘alleles’ during the infection. Concurrent expression of multiple MASP variants has been extensively shown both in vitro and in infected mouse models [ 15 , 17 – 22 ]. Moreover, it is worth noting that several positive sera recognized multiple MASP motifs (including combinations of not co-occurring motifs according to our analysis in Fig 3D ).…”

“…Due to the particular type of gene expression regulation in trypanosomatids, in which nearly all protein-coding genes are arrayed in long polycistronic transcription units [ 16 ], this genomic disposition hinted at a possible co-expression of MASP and TcMUC genes on the surface of trypomastigotes. Subsequent transcriptomic and proteomic studies supported this assumption and unveiled distinctive MASP expression features such as concurrent expression of multiple ‘alleles’ in a single parasite population, and variations in the subset of members preferentially transcribed among trypomastigotes from a single population [ 15 , 17 – 22 ].…”

Benznidazole induces in vitro anaerobic metabolism in Trypanosoma cruzi epimastigotes

“…the metacyclogenesis have been described such as morphology, infectivity and gene expression [10][11][12], the differences in the energetic metabolism throughout the culture growth have not yet been explored. Shah-Simpson et al [13] described the glucose metabolism in epimastigotes, metacyclic trypomastigotes and amastigotes determining the differences in excretion of pyruvate, acetate and alanine.…”

“…It has been described that T. cruzi spontaneously performs metacyclogenesis in vitro at the stationary phase, around 9-11 days of culture [10]. Although several aspects of…”