Trypomastigote Small Surface Antigen ablation causes infection impairment in Trypanosoma cruzi

BALOUZ, VIRGINIA; CRUZ-BUSTOS, TERESA; ROBELLO, CARLOS; BUSCAGLIA, CARLOS ANDRÉS; CÁMARA, MARÍA DE LOS MILAGROS; MASIP, YAMIL EZEQUIEL; DOCAMPO, ROBERTO; RODRIGUEZ, MATÍAS EXEQUIEL; BERNÁ, LUISA; TEKIEL, VALERIA

Woods Hole

Congreso; XXX Molecular Parasitology Meeting; 2019

Trypanosoma cruzi is the etiological agent of Chagas disease. tssa (Trypomastigote Small Surface Antigen) is a multicopy mucin-like gene coding for a glycoprotein displayed on the surface of infective trypomastigote forms that shows amino acids polymorphisms among parasite isolates. These polymorphisms correlate with differential antibody responses in T. cruzi-infected humansand differential adhesion towards non-macrophagic cell monolayers. The TSSA variant present in TcII, TcV and TcVI DTUs has been characterized as a parasite adhesin, engaging surface receptor(s) and inducing signaling pathways on the host cell as a prerequisite for trypomastigote internalization. Most interestingly, trypomastigotes over-expressing TSSA displayed improvedadhesion and infectivity towards non-macrophagic cell lines in vitro. To get further insights into the functional significance of TSSA, we applied CRISPR/Cas9 technique in the RA strain (TcVI) to obtain TSSA-KO parasites. After antibiotic selection, epimastigotes were cloned and genotypified by PCR and PacBIO WGS. tssa gene tandem was successfully ablated. Clones of interest werecycled in vitro and the lack of expression of TSSA protein in trypomastigotes was confirmed by Western blot, IFA and flow cytometry. TSSA-KO clones were significantly less infective in nonmacrophagic cell monolayers and in a three-dimensional cell array than control cell lines in in vitro infection assays. Interestingly, TSSA-KO parasites showed an attenuated phenotype in vivo evidenced by a decreased parasitemia and mortality in BALB/c mice than control cell lines. Altogether, our results shed new light on the interaction between T. cruzi and the host cell. Elucidation of this interaction and the molecules and signals involved is essential for the discovery of new drugable targets and development of new tools to fight against Chagas Disease.