Protein Synthesis Attenuation by Phosphorylation of eIF2α Is Required for the Differentiation of Trypanosoma cruzi into Infective Forms

Chagas' disease is a potentially life-threatening illness caused by the unicellular protozoan parasite Trypanosoma cruzi. It is transmitted to humans by triatomine bugs where T. cruzi multiplies and differentiates in the digestive tract. The differentiation of proliferative and non-infective epimastigotes into infective metacyclic trypomastigotes (metacyclogenesis) can be correlated to nutrient exhaustion in the gut of the insect vector. In vitro, metacyclic-trypomastigotes can be obtained when epimastigotes are submitted to nutritional stress suggesting that metacyclogenesis is triggered by nutrient starvation. The molecular mechanism underlying such event is not understood. Here, we investigated the role of one of the key signaling responses elicited by nutritional stress in all other eukaryotes, the inhibition of translation initiation by the phosphorylation of the eukaryotic initiation factor 2 alpha (eIF2 alpha), during the in vitro differentiation of T. cruzi. Monospecific antibodies that recognize the phosphorylated Tc-eIF2 alpha form were generated and used to demonstrate that parasites subjected to nutritional stress show increased levels of Tc-eIF2 alpha phosphorylation. This was accompanied by a drastic inhibition of global translation initiation, as determined by polysomal profiles. A strain of T. cruzi overexpressing a mutant Tc-eIF2 alpha, incapable of being phosphorylated, showed a block on translation initiation, indicating that such a nutritional stress in trypanosomatids induces the conserved translation inhibition response. In addition, Tc-eIF2 alpha phosphorylation is critical for parasite differentiation since the overexpression of the mutant eIF2 alpha in epimastigotes abolished metacyclogenesis. This work defines the role of eIF2 alpha phosphorylation as a key step in T. cruzi differentiation.