Fig 3.

ER stress represses global translation initiation in Toxoplasma. (A) Equal amounts of lysate from parasites treated with tunicamycin (TUN) or vehicle (DMSO) were separated in a 4 to 12% polyacrylamide gel and transferred for Western blotting with antibodies that specifically recognize TgIF2α phosphorylated at Ser-71 (TgIF2α-P) or total TgIF2α protein. (B) Polysome profiles were generated from parasites exposed to 10 μM tunicamycin (TUN) or no treatment (NT) for 1 h. Fractions containing free RNA, ribosomal subunits, and the 80S monoribosomes were collected and pooled into a single tube (termed free ribosomes and monosomes). A second pool of fractions containing mRNAs engaged with three or more ribosomes (large polysomes) were collected for subsequent microarray analysis.