Proteomic identification of a sporozoite-specific antigen using HDAC3 inhibitor-treated tachyzoites as surrogate.

The apicomplexan parasite has a complex life cycle. Access to sexual stages and sporozoite-containing oocysts, essential for studying the parasite's environmental transmission, is limited and requires animal experiments with cats. Thus, alternatives and resource-efficient methods are needed. Several molecular factors and transcriptional switches responsible for differentiation have been identified in recent years. In tachyzoites, drug-induced inhibition of the histone deacetylase HDAC3, or genetic depletion of transcription factors regulating HDAC3, leads to the expression of genes that are specific to sexual stages and oocysts. Here, we applied this concept and showed that the commercially available HDAC3 inhibitor apicidin could be used to identify the hitherto unknown antigen of the sporozoite-specific monoclonal antibody G1/19 in tachyzoites. Using mass spectrometry of immunoprecipitated G1/19 target protein from apicidin-treated cultures, we identified it as SporoSAG. In addition, for the much less abundant sporozoite-specific protein LEA860, apicidin treatment was still sufficient to induce a detectable protein level in immunofluorescence microscopy. We also discuss further applications and the limitations of this approach. This allows to overcome issues with the paucity of material of sexual stages and oocysts from to some extent without the need for cat-derived material.