The Small heat shock protein 20 palmitoylation is important for proper sub-cellular localization in Toxoplasma gondii

DE NAPOLI, MG; MORENO SN; ANGEL, SO; CORVI, MM

Ottawa

Congreso; The eleventh international congress on toxoplasmosis; 2011

Toxoplasma gondii expresses five different stress-induced small heat shock proteins (HSPs) that localize to different organelles within the parasite. Among them, TgHsp20 presents chaperone activity and is found to localize at the inner membrane complex (IMC) facing the plasma membrane. In spite of this, TgHsp20 does not contain any signal peptide, transmembrane domain or hydrophobic regions. An “in silico” analysis suggested that TgHSP20 could be palmitoylated and this could explain its localization. Metabolic labeling followed by immunoprecipitation confirmed that this protein is post-translationally modified by palmitic acid. As such, different stable transfectants expressing mutant versions of TgHSP20 on putative palmitoylation sites were generated. Metabolic labeling assays of these constructs suggested that all three cysteine residues are palmitoylated “in vivo”. In addition, indirect immuno-fluorescence assays demonstrated that the cysteine residues are important for proper sub-cellular localization. Triple mutant TgHSP20C3,4,160S presents a cytoplasmic localization in non-dividing parasites but localizes to the IMC of daughter cells. All these results suggest that TgHSP20 palmitoylation is not important for its association to the IMC during parasite replication but it is important for correct localization in mature tachyzoites. Furthermore, putative TgHSP20 palmitoylation sites are conserved in other HSP20 in Apicomplexas, suggesting an important role for this modification on HSP20.