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

Background The Trypanosoma cruzi genome bears a huge family of genes and pseudogenes coding for Mucin-Associated Surface Proteins (MASPs). MASP molecules display a ‘mosaic’ structure, with highly conserved flanking regions and a strikingly variable central and mature domain made up of different combinations of a large repertoire of short sequence motifs. MASP molecules are highly expressed in mammal-dwelling stages of T. cruzi and may be involved in parasite-host interactions and/or in diverting the immune response. Methods/Principle findings High-density microarrays composed of fully overlapped 15mer peptides spanning the entire sequences of 232 non-redundant MASPs (~25% of the total MASP content) were screened with chronic Chagasic sera. This strategy led to the identification of 86 antigenic motifs, each one likely representing a single linear B-cell epitope, which were mapped to 69 different MASPs. These motifs could be further grouped into 31 clusters of structurally- and likely antigenically-related sequences, and fully characterized. In contrast to previous reports, we show that MASP antigenic motifs are restricted to the central and mature region of MASP polypeptides, consistent with their intracellular processing. The antigenicity of these motifs displayed significant positive correlation with their genome dosage and their relative position within the MASP polypeptide. In addition, we verified the biased genetic co-occurrence of certain antigenic motifs within MASP polypeptides, compatible with proposed intra-family recombination events underlying the evolution of their coding genes. Sequences spanning 7 MASP antigenic motifs were further evaluated using distinct synthesis/display approaches and a large panel of serum samples. Overall, the serological recognition of MASP antigenic motifs exhibited a remarkable non normal distribution among the T. cruzi seropositive population, thus reducing their applicability in conventional serodiagnosis. As previously observed in in vitro and animal infection models, immune signatures supported the concurrent expression of several MASPs during human infection. Conclusions/Significance In spite of their conspicuous expression and potential roles in parasite biology, this study constitutes the first unbiased, high-resolution profiling of linear B-cell epitopes from T. cruzi MASPs during human infection.
Author summary Trypanosomatids belong to an ancient eukaryotic lineage that cause devastating diseases in humans and livestock. The elucidation of the genomes of Trypanosoma cruzi, Trypanosoma brucei and Leishmania major established a landmark in the study of these relevant protozoa. In particular, the genome of T. cruzi, the agent of Chagas disease, evidenced a marked expansion and diversification of gene families likely involved in parasite-host interplay. MASPs (for mucin-associated surface proteins) define the second largest T. cruzi gene family, with ~1400 genes (and pseudogenes) coding for surface glycoproteins involved in parasite infectivity and immune evasion. MASP deduced products bear highly conserved sorting signals on their N- and C-terminal regions that direct and anchor them to the parasite plasma membrane. Conversely, the central and mature domain, the only one displayed on the parasite surface is strikingly variable and exhibit a ‘mosaic-like’ structure made up of different combinations of a large repertoire of short sequence motifs. Few studies have analyzed MASP antigenicity so far, and they were carried out using either low throughput approaches or animal infection models. By means of exquisite peptide micro-arrays technology followed by conventional serological methods we herein present the first non-biased, high-throughput profiling of linear B-cell epitopes from a representative pool (n = 232) of T. cruzi MASPs during human infection. Our findings are discussed in terms of MASP evolution, expression profile and applicability in Chagas disease serodiagnosis.