Identification, characterization and subcellular localization of TcPDE1, a novel cAMP-specific phosphodiesterase from Trypanosoma cruzi

Abstract

Compartmentalization of cAMP phosphodiesterases plays a key role in the regulation of cAMP signalling in mammals. In the present paper, we report the characterization and subcellular localization of TcPDE1, the first cAMP-specific phosphodiesterase to be identified from Trypanosoma cruzi. TcPDE1 is part of a small gene family and encodes a 929-amino-acid protein that can complement a heat-shock-sensitive yeast mutant deficient in phospho-diesterase genes. Recombinant TcPDE1 strongly associates with membranes and cannot be released with NaCl or sodium cholate, suggesting that it is an integral membrane protein. This enzyme is specific for cAMP and its activity is not affected by cGMP, Ca2+, calmodulin or fenotiazinic inhibitors. TcPDE1 is sensitive to the phosphodiesterase inhibitor dipyridamole but is resistant to 3-isobutyl-1-methylxanthine, theophylline, rolipram and zaprinast. Papaverine, erythro-9-(2-hydroxy-3-nonyl)-adenine hydrochloride, and vinpocetine are poor inhibitors of this enzyme. Confocal laser scanning of T. cruzi epimastigotes showed that TcPDE1 is associated with the plasma membrane and concentrated in the flagellum of the parasite. The association of TcPDE1 with this organelle was confirmed by subcellular fractionation and cell-disruption treatments. The localization of this enzyme is a unique feature that distinguishes it from all the trypanosomatid phosphodiesterases described so far and indicates that compartmentalization of cAMP phosphodiesterases could also be important in these parasites.

Biological and Structural Characterization of Trypanosoma cruzi Phosphodiesterase C and Implications for Design of Parasite Selective Inhibitors

“…TcrPDEA is a cAMP-specific PDE with a high K m and is not inhibited by isobutylmethylxanthine, papaverine, or theophylline (10). The highly similar TcrPDEB1 and TcrPDEB2 are cAMP-specific and contain two GAF domains upstream of their catalytic domains (11,14,17). TcrPDEC contains a FYVE-type domain (18) at its N terminus and was shown to hydrolyze both cAMP and cGMP with similar K m values (13).…”

A FYVE‐containing unusual cyclic nucleotide phosphodiesterase from Trypanosoma cruzi

“…A cAMP‐specific phosphodiesterase (PDE) activity has been demonstrated in T. cruzi by various laboratories [12,13]. Recently, the first cyclic‐nucleotide‐specific PDE from T. cruzi has been identified and characterized at the molecular level [14]. This enzyme, TcPDE1 is entirely cAMP‐specific, and it is located along the flagellum.…”

Pde1 Phosphodiesterase Modulates Cyclic AMP Levels through a Protein Kinase A-Mediated Negative Feedback Loop in Cryptococcus neoformans

“…Pde2 is a high-affinity PDEase that is represented in organisms ranging from yeast to mammals (7). In contrast, Pde1, the low-affinity PDEase, is less well characterized but is found in a range of organisms, including S. cerevisiae, Schizosaccharomyces pombe, Candida albicans, Dictyostelium discoideum, Vibrio fischeri, Leishmania mexicana, Trypanosoma brucei, and Trypanosoma cruzi (9,11,14,22,27,28,34,39).…”