Biology Reference
In-Depth Information
TcrPDEC is weakly inhibited by etazolate, trequinsin, and dipyridamole (IC
50
values in the 1
m
M range), while it is highly resistant to many other PDE inhibitors,
including rolipram, cilostamide, zardaverin, IBMX, and papaverine.
PDED
. All kinetoplastid genomes contain one copy of this putative PDE. The
DNA sequence predicts a polypeptide of about 670 amino acids length containing a
class 1 catalytic domain at the C-terminus. Currently, only the sequences are known
and no experimental data are available in any species.
2.2 Biological Function
PDEA
. Homozygous deletion of TbrPDEA has only minor effects on cell growth
in culture. A slight increase in the steady-state cAMP concentration is observed in
the deletion mutant, and generation time is prolonged by about 20%. However, the
deletion mutants remain fully infectious for tsetse flies (Gong et al.
2001
). The
currently available information suggests that TbrPDEA is a minor activity in both
bloodstream and procyclic trypanosomes and that it is not essential for cell viabil-
ity. Its biological role might rather consist of the modulation of cAMP signals,
analogous to what has been suggested for the high-
K
m
class 2 PDE of
S. cerevisiae
(Ma et al.
1999
).
PDEB
. In striking contrast, TbrPDEB1 and TbrPDEB2 are essential enzymes in
T. brucei
. The lack of success in early attempts to knock out these genes in
bloodstream form
T. brucei
indicated that they might be required for cell survival
(Zoraghi and Seebeck
2002
). The development of a genetically encoded inducible
RNA interference system in
T. brucei
enabled reexamination of the biological role
of the two enzymes; RNAi against either of the two enzymes does not produce an
observable phenotype. However, simultaneous RNAi against both PDEs leads
to complete cell lysis within 48 h after induction of RNAi (Oberholzer et al.
2007
). Simultaneous RNAi, but not RNAi against the individual enzymes, results
in a dramatic increase (100-fold) in steady-state intracellular cAMP concentration
within a few hours of inducing RNAi. RNAi against TbrPDEB1 and TbrPDEB2
does not increase the mRNA levels of the other three PDEs, TbrPDEA, TbrPDEC,
or TbrPDED, indicating that the expression of these PDEs is not subject to control
by cAMP.
Further experiments demonstrated that TbrPDEB1 and TbrPDEB2 can function-
ally complement each other, despite the fact that their subcellular localization is
clearly different (Oberholzer et al.
2007
; Kunz et al.
2009a
). This finding is
somewhat at variance with current concepts of neatly localized cAMP microdo-
mains and precisely anchored members of the signaling cascade. Rather, it suggests
that maintenance of the overall steady-state concentration of cAMP by a PDE
located anywhere in the cell is sufficient for preserving many of the basic functions
of the cell. This conclusion is also supportedbythecomplementationofPDE-deficient
yeast cells by various heterologous PDEs (Kunz et al.
2004
; Johner et al.
2006
;
