Fig. 3.

Import of tRNAs in Plasmodium sporozoites. (A) P. berghei, P. yoelii, and P. falciparum WT sporozoites were incubated with (+) or without (−) E. coli tRNA^Val and were subjected to FISH. Due to high sequence similarities between mammalian and plasmodial tRNAs, E. coli tRNA^Val was chosen as a template so that a specific FISH probe could be designed [3′ end-labeled with Texas Red (TxRd)] that does not cross-hybridize with any Plasmodium endogenous tRNAs. On tRNA import, about 80% of sporozoites (WT P. berghei, P. yoelii, and P. falciparum) hybridized the fluorescent probe. (Scale bars, 2 µm.) (B) Radioactive tRNAs from mouse Hepa1-6 cells were cosedimented with both infectious and noninfectious P. berghei sporozoites in the absence (Left) and presence (Center and Right) of RNase A. Following extracellular RNase treatment, intracellular sporozoite endogenous tRNAs were undamaged (Right), and exogenously added radiolabeled tRNAs were observed within living sporozoites only (Center), indicating tRNA import had occurred. Additional bands correspond to tRNA aggregation resulting from phenol extraction.