trans

272 INTRACELLULAR SIGNALING
cells with 1N2K (1 nucleus, 2 kinetoplasts) produce
1N1K daughter cells and a zoid (0N1K).
However, in bloodstream forms the absence of
TbCYC6 results in blocked mitosis and cytokinesis
without affecting kinetoplast replication
and segregation. These results suggest developmental
stage-specific checkpoint control
mechanisms in T. brucei.
CDKs and cyclins in Plasmodium
Similar to the trypanosomatids, several CDK
homologs have been cloned from P. falciparum.
However, the physiological functions of these
kinases are not clear yet. PfPK5 contains a PST-
TIRE motif instead of PSTAIRE and is the most
likely CDK-like kinase candidate to drive the
Plasmodium cell cycle. PfPK5 phosphorylates
histone H1 in vitro and localizes to the nucleus
at the beginning of nuclear division. Interestingly,
PfPK5 shows robust activation by human
cyclin H and p25 (CDK5-activating cyclin). The
activation by cyclin H is surprising, since cyclin
H is thought to be a partner of CDK-activating
kinase CDK7 and not CDK1. Furthermore, p25
and cyclin H are equally effective in stimulating
PfPK5 activity. It is widely known that p25 is
quite specific about its preference for CDK5.
In addition, cyclin H association also activates
CDK5. These results provide strong evidence
that PfPK5 is an ortholog of CDK5 and is
promiscuous for its cyclin partner, in common
with yeast Pho85 kinases. A P. falciparum
cyclin homolog PfCYC1 also strongly stimulates
PfPK5 activity. Similarly, a Xenopus protein
RINGO (rapid inducer of G 2 /M progression in
oocytes) also strongly stimulates PfPK5 activity.
RINGO has no sequence similarity to cyclins,
but can bind and activate Cdc2. In contrast,
human CDK inhibitor (CKI) p21 CIP1 inhibits
PfPK5 activity in vitro.
Another CDK-related kinase in P. falciparum,
PfPK6, contains a SKCILRE sequence in place
of PSTAIRE. PfPK6 has characteristics of both
CDKs and MAPKs, localizes mainly to the
cytoplasm, and has a preference for Mn 2 .
However, unlike PfPK5, PfPK6 appears to be a
cyclin-independent kinase and shows strong
autophosphorylation and histone H1 kinase
activity in the absence of a cyclin. The IC 50 values
of PfPK6 to the CDK inhibitor rescovitine
are close to MAPKs rather than CDKs. PfPK6
appears to interact with translational elongation
factor-1 (EF-1). EF-1 can interact with
the cytoskeleton by binding and bundling actin
filaments and microtubules, suggesting that
PfPK6, rather than directly regulating cell-cycle
transition, may have a secondary role in cellcycle
control through its involvement in translation
regulation or cytoskeletal organization.
Additional CDK-related kinases in Plasmodium
include Pfcrk1, which contains an
AMTSLRE motif similar to PITSLRE in p58 GTA .
PfCRK1 has been implicated in sexual stage
development as the transcript accumulates
in gametocytes. PfMRK uses NFVLLRE and is
similar to NRTALRE from the CDK-activating
kinase (CAK) CDK7. A complex between PfMRK
and human cyclin H stimulates histone H1
kinase activity. Interestingly, PfMRK/human
cyclin H does not activate PfPK5 suggesting
that either PfMRK has a substrate preference
different from PfPK5 or that it is not a CAK. All
CDK-like kinases from P. falciparum autophosphorylate,
unlike CDKs from other species.
In an effort to identify parasite intracellular
targets of the purine CDK inhibitors, purvalanol
was coupled to an agarose matrix. Surprisingly,
casein kinase 1 (CK1) was the main protein
that bound to the matrix in extracts of P. falciparum,
L. mexicana, T. cruzi, and T. gondii, in
contrast to results from metazoans. Predicted
amino acid sequences from P. falciparum and
T. cruzi CK1 genes show strong identity with
known sequence isoforms, and recombinant
proteins showed properties characteristic of
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA