FIG 6.

Impaired stress-induced LdeIF2α phosphorylation hampers promastigote-to-amastigote transformation and reduces parasite viability. (A) Phase-contrast images showing morphological changes during transformation from promastigotes (Pro) to amastigotes (Ama) in the absence or presence of GSK2606414. (B) Polysome profiles of L. donovani promastigotes grown under unstressed conditions and axenic amastigotes and promastigotes subjected to 48 h of high temperature and low pH (37°C and pH 5.5) with GSK2606414 (pH+T+GSK) or without GSK2606414 treatment. (C) Western blot analysis of L. donovani protein lysates with monoclonal anti-A2 antibody to determine the expression levels of the A2 protein in unstressed promastigotes or promastigotes subjected to 48 h of high temperature and low pH (37°C and pH 5.5, respectively) (pH+T) and promastigotes subjected to 48 h of high temperature and low pH (37°C and pH 5.5, respectively) along with GSK2606414. LdeIF2α and α-tubulin were used as loading controls in the same experiment. Data shown here are representative of data from three independent experiments with similar results. (D) Percent cell viability of L. donovani promastigotes subjected to combined exposure to high-temperature (37°C) and low-pH (pH 5.5) stress or high-temperature (37°C) and low-pH (pH 5.5) stress with GSK2606414 or in the presence of GSK2606414 only, monitored by MTT survival assays at 48 h.