TcTASV: A Novel Protein Family in Trypanosoma cruzi Identified from a Subtractive Trypomastigote cDNA Library

Abstract

BackgroundThe identification and characterization of antigens expressed in Trypanosoma cruzi stages that parasitize mammals are essential steps for the development of new vaccines and diagnostics. Genes that are preferentially expressed in trypomastigotes may be involved in key processes that define the biology of trypomastigotes, like cell invasion and immune system evasion.Methodology/Principal FindingsWith the initial aim of identifying trypomastigote-specific expressed tags, we constructed and sequenced an epimastigote-subtracted trypomastigote cDNA library (library TcT-E). More than 45% of the sequenced clones of the library could not be mapped to previously annotated mRNAs or proteins. We validated the presence of these transcripts by reverse northern blot and northern blot experiments, therefore providing novel information about the mRNA expression of these genes in trypomastigotes. A 280-bp consensus element (TcT-E element, TcT-Eelem) located at the 3′ untranslated region (3′ UTR) of many different open reading frames (ORFs) was identified after clustering the TcT-E dataset. Using an RT-PCR approach, we were able to amplify different mature mRNAs containing the same TcT-Eelem in the 3′ UTR. The proteins encoded by these ORFs are members of a novel surface protein family in T. cruzi, (which we named TcTASV for T. cruzi Trypomastigote, Alanine, Serine and Valine rich proteins). All members of the TcTASV family have conserved coding amino- and carboxy-termini, and a central variable core that allows partitioning of TcTASV proteins into three subfamilies. Analysis of the T. cruzi genome database resulted in the identification of 38 genes/ORFs for the whole TcTASV family in the reference CL-Brener strain (lineage II). Because this protein family was not found in other trypanosomatids, we also looked for the presence of TcTASV genes in other evolutionary lineages of T. cruzi, sequencing 48 and 28 TcTASVs members from the RA (lineage II) and Dm28 (lineage I) T. cruzi strains respectively. Detailed phylogenetic analyses of TcTASV gene products show that this gene family is different from previously characterized mucin (TcMUCII), mucin-like, and MASP protein families.Conclusions/SignificanceWe identified TcTASV, a new gene family of surface proteins in T. cruzi.

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

“…Much of the genome of T. cruzi is composed of repeated elements, among which the following multigene surface families are found: TS, mucin, MASP, dispersed gene family-1, gp63, TcTSAV or TcSMP 18 39 40 41 . The antigenicity of proteins belonging to the TS and mucin families has been previously described, and recent reports showed that peptides from the hypervariable MASP region could elicit IgM and IgG responses in experimental infections 30 .…”

Towards High-throughput Immunomics for Infectious Diseases: Use of Next-generation Peptide Microarrays for Rapid Discovery and Mapping of Antigenic Determinants

“…Peptides from the same protein were synthesized at randomized positions within a single sector. The chip contains additional 2386 unique peptides corresponding to eight proteins not accounted within the protein groups defined above (and of which 2 proteins resulted seropositive), including members of the T. cruzi TASV protein family ( 30 , 31 ) and 21 epitopes of high prevalence in healthy humans (not associated to Chagas disease, identified in the IEDB with a search for linear B-cell epitopes from any source with sero-prevalence > = 50%, assayed in at least 20 subjects). The sequences of these 532 proteins (457 T. cruzi proteins, 54 neo-proteins and the additional 21 short peptidic epitopes of high prevalence in healthy humans) are available in supplemental Table S4 .…”

Advances and challenges towards a vaccine against Chagas disease

“…54 Similarly, a novel family of surface proteins has been recently identified that seems to be specific for T. cruzi. 55 Taken together, these studies may thus lead to some promising antigens with important potential as novel vaccine candidates.…”