Figure 6. Effect of histidine supplementation on puncta formation and GFP-TbAtg8.1 levels.

T. brucei procyclic forms expressing GFP-TbAtg8.1 were incubated for 2 h in gHBSS, supplemented with histidine, glutamine or valine as indicated. Fluorescent puncta were analyzed and quantified as puncta per cell (A) and cells containing one or more puncta (B). The results are from two independent experiments, in which at least 100 cells per incubation condition were analyzed. Error bars indicate standard errors of the mean. (C) Immunoblots of parasite lysates obtained after incubation of cells in the absence or presence of amino acids and probed with antibodies against GFP (lower panel), or eukaryotic elongation factor 1A (upper panel) as loading control. Each lane contains 5×10⁶ cell equivalents. Molecular mass markers are indicated in kDa in the margin.