cAMP-dependent Signaling in Trypanosoma cruzi

EDREIRA, M; JÄGER, AV; MILD, JG; MC CORMACK, B; PEREYRA, E; ALTSCHULER, DL; DE GAUDENZI, JG

Bariloche

Congreso; The Second South American Spring Symposium in Signal Transduction and Molecular Medicine; 2012

cAMP has been implicated as a secondary messenger in a wide range of cellular processes. Effectors include PKA and Epac. In contrast to mammalians, little is known about cAMP mechanism of action in T. cruzi. Several studies suggested the involvement of cAMP in T. cruzi life cycle. Even PKA has been cloned and characterized, a genomic sequence for Epac has not been found. Early studies demonstrated a negative role for cAMP in cell proliferation. However, inhibition of PKA (the only known T. cruzi cAMP effector up-to-date) instead of the expected rescue, led also to epimastigote death. This result strongly suggests the existence of other biologically active cAMP effectors. In order to identify new cAMP effectors, we carried out in silico studies using the predicted T. cruzi proteome. A combination of different search methods let us identify 27 proteins with putative cAMP binding domains. Phylogenetic analysis made from coding sequence alignments showed that these proteins are segregated into two main branches: one containing protein kinases and the other gathering hypothetical proteins with no assigned function. Meanwhile, phylogenetic analysis of the cAMP binding domains presented a more homogeneous distribution consistent with the conservation of a nucleotide-binding domain. Thus, we could identify conserved cAMP binding domain sequences, in hypothetical proteins different from PKA. Part of the in silico hits were cloned and express in bacteria, in order to experimentally validate these potential new proteins as novel cAMP effectors.