Localization of PfCysRS isoforms using GFP fusions in transgenic

Plasmodium parasites. Live-cell epifluorescence microscopy of P. falciparum transfected using pGlux.1 vector for episomal expression of GFP fused to the two Pf CysRS mRNA isoforms: PfCysRS-A 1–153 GFP (A) and Pf CysRS-B 1–70 GFP (B) in live cells. In (A), both parasites are early trophozoite stage, and localization of GFP is to a single punctum, consistent with the size and position of the apicoplast. The redblood cell (RBC) and the parasite (P) are indicated. (B) An early trophozoite (top panels) and late trophozoite (bottom panels) with GFP dispersed throughout the cytosol. (C) Immunofluorescence assays of the parasites indicated, PfCysRS-A 1–153 GFP (top panels) and PfCysRS-B 1–70 GFP (bottom panels), using antibodies against GFP and ACP, an apicoplast marker. The PfCysRS-A 1–153 GFP signal specifically co-localizes with the apicoplast marker, whereas the PfCysRS-B 1–70 GFP signal is distributed throughout the cytosol. For (A)–(C), all scale bars indicate 5 µm. Pham JS, Sakaguchi R, Yeoh LM, De Silva NS, McFadden GI, Hou YM, Ralph SA. A dual-targeted aminoacyl-tRNA synthetase in Plasmodium falciparum charges cytosolic and apicoplast tRNACys. Biochem J. 2014 458(3):513-23