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CALCIUM 247
on sequence homologies. Susceptibility of
PfATP 4 and TcSCA to cyclopiazonic acid
suggest that this inhibitor might be a useful
tool for disrupting Ca 2 homeostasis in these
organisms. Whether or not overexpression
of the SERCA pumps changes the level of Ca 2
stored in the ER and alters cell function is
not known. Nonetheless, the expression level
of PfATP 4 varies during the Plasmodium life
cycle, with the maximum level reached in merozoites
compared to ring stages. In Leishmania,
changes in the expression level of Lmaa 1 have
also been detected. Here, overexpression correlates
with increased virulence. However, Ca 2
storage within the ER could not be measured.
In procyclic T. brucei, a tenfold overexpression
of TBA1 was achieved with no effect on parasite
growth rate or basal levels of [Ca 2 ] i . The
expression levels of TBA1 and TcSCA are constant
throughout the life cycle.
Ca 2 release channels of the ER and
regulation by phospholipid metabolites
During the signal process, stored Ca 2 in the
ER is released through specific receptors that
also function as channels. The receptors are
divided into two related families, depending
upon whether release is activated by binding
to inositol 1,4,5 trisphosphate (InsP 3 ) or to the
fungal toxin ryanodine. In addition, both receptor
families open in response to an elevation in
cytoplasmic Ca 2 concentration. This phenomenon
is known as Ca 2 -induced Ca 2 release
(CICR). The process is biphasic since too much
external Ca 2 can close the channel and prevent
further release. Ca 2 release through the InsP 3
receptor is also quantal. In other words, discrete
amounts of Ca 2 are released in response
to increasing amounts of agonists. Structurally,
the receptor families are highly related in their
carboxyl termini, the region that spans the
membrane and forms the functional channel.
The amino terminus of each is variable and
binds to the different channel agonists. Along
with its distribution on the cytoplasmic face of
the ER, the InsP 3 receptor is also found on the
nuclear matrix side of the nuclear envelope.
In mammalian cells, stored Ca 2 within the
nuclear envelope has been detected with
sequestered fura-2. Presence of the InsP 3 receptor
might represent a regulated route for the
direct movement of Ca 2 into the nucleus.
Ca 2 movement into the nucleus has been
measured with targeted aequorin in T. brucei
procyclic forms. However, the Ca 2 appeared
to pass through nuclear pores. Whether the
nuclear envelope of parasites can modulate
nuclear Ca 2 levels is not known. In cells that
are not electrically coupled (a model that may
approximate the situation in pathogenic
protozoa), depletion of the ER Ca 2 pool is associated
with regulation of store-operated Ca 2
channels (SOCs) in the plasma membrane.
This phenomenon of Ca 2 gating is referred to
as capacitative Ca 2 entry.
Because the ER Ca 2 channels can be regulated
by InsP 3 , the metabolism of phosphatidylinositol
(4,5) bisphosphate is of broad
interest. Additionally, phosphorylated forms
of phosphatidylinositol can serve to recruit
proteins that contain pleckstrin homology (PH)
domains. Interestingly, two proteins that are
thought to utilize PH domains as membrane
anchors lack these motifs in the parasite
homologs. These proteins include phospholipase
C- from T. cruzi and protein kinase B
from Plasmodium.
In T. cruzi, phosphatidylinositol and its
metabolites have been detected by TLC and
HPLC. The amount of InsP 3 increases upon
exposure of permeabilized cells to Ca 2 , or upon
treatment of intact cells with the cholinergic
agonist carbachol. Plasmodium also varies the
amount of phosphatidylinositol (4,5) bisphosphate
in the membranes, with the highest
amounts detected in later stages of the asexual
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA