trans

268 INTRACELLULAR SIGNALING
a different MAPK (Hog1). MAPKKKs are usually
diverse in structure, allowing them to interact
with different groups of proteins. The regulatory
regions of MAPKKK may include pleckstrin
homology (PH) domains, leucine-zipper dimerization
sequences, src homology 3 domains
(SH3), or association sites for GTP-binding
proteins. This heterogeneity of MAPKKK regulatory
domains allows MAPK modules to
respond to an assortment of stimuli.
Downstream of MAPK activation is the regulation
of gene expression brought about by
phosphorylation of a variety of transcription
factors such as Ets1, c-Jun, c-myc, Elk1, ATF-2,
Max, p53 and NFAT4, among many. Although
control of gene expression through phosphorylation
is an important function of MAPKs,
the pool of active MAPK does not completely
translocate to the nucleus. Active MAPKs also
regulate cytoplasmic proteins such as p90 srk S6
kinase, MAPKAP kinase, or carbamoyl phosphate
synthetase. The phosphorylation site of
MAPK substrates usually contains a serine or
threonine followed by proline at the P1 position,
hence MAPKs are known as prolinedirected
kinases. Sometimes a proline at the
P2 site is also present. Additional domains
also are involved in substrate recognition by
MAPKs: the D domain composed of basic
and hydrophobic residues, and the FXFP
sequence. It would be an important and interesting
endeavor to characterize downstream
processes of MAPK-mediated signaling pathways
in protozoan parasites.
MAP kinase activation in protozoan parasites
MAPK homologs have been identified in
many parasites including Plasmodium,
Trypanosoma, Leishmania, and Giardia. How
these structures might be activated is not
known. While tyrosine kinase activity is present
in these organisms, only indirect evidence
is available to suggest catalytic receptors
might also be present. Serpentine receptors
have not been identified. Heterotrimeric
G-protein activity has been hinted at with
antibodies, labeled GTP analogs or ADPribosylation
with cholera toxin or pertussis
toxin. Indirect evidence for G exists in T.
brucei, T. cruzi, P. falciparum and T. gondii.
However, no genes corresponding to G have
been cloned. Nonetheless, the treatment of
asexual malaria parasites with cholera toxin
induces gametocytogenesis, implicating G-
protein-coupled signals in the sexual differentiation
process. Ras-mediated signaling is an
important component of MAPK activation.
Genes coding for Ras have been identified in
Entamoeba. Two isoforms of Entamoeba Ras,
EhRas1 and EhRas2, are 47% similar to human
Ras. Both EhRas1 and EhRas2 carry the signature
cysteine-aliphatic-aliphatic-any amino
acid (CAAX) prenylation sequence, and the
prenylation of EhRas1 has been observed
in vitro. Inhibitors of prenyl transferases are
lethal to T. brucei and P. falciparum. Protein
prenylation has also been demonstrated in
Giardia, T. brucei, and P. falciparum.
MAP kinase pathways in apicomplexans
In T. gondii, studies with heterologous ERK
antibodies and those using gel kinase assays
suggest the presence of MAPK-like proteins in
cell homogenates. In P. falciparum two putative
MAPK homologs have been identified. PfMAP-
1 contains MAPK conserved sequences, including
the TXY activation motif and a large
carboxy terminus extension of highly charged
tetra- or octapeptide repeats. Recombinant
PfMAP-1 autophosphorylates both the tyrosine
and threonine residues in this motif,
increasing catalytic activity to a small extent.
The 826 amino acid residue PfMAP-1 has the
potential to generate a 100 kDa protein. Interestingly,
Western blot analysis indicated the
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA