A FYVE‐containing unusual cyclic nucleotide phosphodiesterase from Trypanosoma cruzi

Abstract

Cyclic‐nucleotide‐specific phosphodiesterases (PDEs) are key players in the intracellular signaling pathways of the important human pathogen Trypanosoma cruzi. We report herein the identification of an unusual PDE from this protozoal organism. This enzyme, TcrPDEC, is a member of the class I PDEs, as determined from the presence of a characteristic signature sequence and from the conservation of a number of functionally important amino acid residues within its catalytic domain. Class I PDEs include a large number of PDEs from eukaryotes, among them all 11 human PDE families. Unusually for an enzyme of this class, TcrPDEC contains a FYVE‐type domain in its N‐terminal region, followed by two closely spaced coiled‐coil domains. Its catalytic domain is located in the middle of the polypeptide chain, whereas all other class I enzymes contain their catalytic domains in their C‐terminal parts. TcrPDEC can complement a PDE‐deficient yeast strain. Unexpectedly for a kinetoplastid PDE, TcrPDEC is a dual‐specificity PDE that accepts both cAMP and cGMP as its substrates.

Chemical Validation of Phosphodiesterase C as a Chemotherapeutic Target in Trypanosoma cruzi , the Etiological Agent of Chagas' Disease

“…PDE-deficient Saccharomyces cerevisiae strain PP5 transformed with the construct containing the entire full-length open reading frame of TcrPDEC was obtained from Thomas Seebeck, University of Bern, Bern, Switzerland. The cells were grown and maintained in SC-Leu medium (15).…”

“…In general, PDEs have unique N-terminal regulatory domains, and the catalytic domain is located near their C terminus (16). TcrPDEC is the only trypanosome PDE identified to date that is capable of hydrolyzing cyclic GMP (cGMP), although it prefers cAMP as a substrate (15). Additionally, TcrPDEC is unusual in that its N-terminal region contains a FYVE-type domain, a functional domain that has not been found in any PDE so far (2,15).…”

“…This pathway is terminated by the action of a T. cruzi cAMP phosphodiesterase (PDE) C (TcrPDEC) (28) that is located in the CVC (A. Schoijet, K. Miranda, L. C. Soares Medeiros, W. de Souza, M. Flawia, H. Torres, R. Docampo, and G. Alonso, unpublished results). This is a novel and rather unusual PDE in which, unlike all other class I PDEs, the catalytic domain is localized in the middle of the polypeptide chain (2,15). In general, PDEs have unique N-terminal regulatory domains, and the catalytic domain is located near their C terminus (16).…”

“…TcrPDEC is the only trypanosome PDE identified to date that is capable of hydrolyzing cyclic GMP (cGMP), although it prefers cAMP as a substrate (15). Additionally, TcrPDEC is unusual in that its N-terminal region contains a FYVE-type domain, a functional domain that has not been found in any PDE so far (2,15). It should be possible to design inhibitors that are selective for the trypanosome PDEs and do not cause toxic effects owing to actions on mammalian PDEs.…”

Hydrolysis products of cAMP analogs cause transformation ofTrypanosoma bruceifrom slender to stumpy-like forms

“…In earlier studies, etazolate showed antiproliferative effects, causing slender-to-stumpy transformation at concentrations of 1-2 M (12). However, recent work characterizing T. brucei PDE families (16,17,27,28) showed that etazolate only modestly inhibits most T. brucei PDEs. Inhibition of total PDE activity in T. brucei whole-cell lysates in the presence of etazolate was measured, and etazolate was found to inhibit PDE activity with an IC 50 of 810 Ϯ 10 M [at 1 M cAMP (29), n ϭ 3] (Fig.…”