Primary structure of the Plasmodium vivax crk2 gene and interference of the yeast cell cycle upon its conditional expression.
The cdc2 gene product, a 34-kDa protein kinase, plays a universal role in the M phase of the eukaryotic cell cycle. To study the cell cycle regulation in malarial parasites, we have characterized a cdc2-related gene from the most widely distributed human malaria, Plasmodium vivax (Pvcrk2). The full-length Pvcrk2 revealed 90--99% homology with Crk2 proteins from other Plasmodium species and approximately 60% homology with p34(cdc2) proteins from higher eukaryotes. We used the temperature-sensitive Schizosaccharomyces pombe cdc2 mutant (cdc2-33(ts)) for gene complementation studies. Expression of the full-length 33-kDa PvCrk2 protein, a truncated 27-kDa version, and two chimeric proteins in which we exchanged the N- and C-terminal regions of PvCrk2 with their S. pombe counterparts at the restrictive temperature in the mutant cdc2-33(ts) did not complement the cell cycle defect. However, conditional expression of the Pvcrk2 genes or the chimera containing the C terminus from Spcdc2 in mutant cdc2-33(ts) cells produced cell-cycle-arrested phenotypes only in the induced state and at the permissive temperature. Our results thus provide the first compelling genetic evidence that the plasmodial Crk2 gene product(s) is capable of interfering with the well-conserved eukaryotic cell cycle machinery.
Sperança
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MA
Departamento de Parasitologia, Instituto de Ciências Biomédicas, 05508-900 São Paulo, SP, Brazil.
Vinkenoog
R
R
Ocampos
M
M
Fischer
K
K
Janse
C J
CJ
Waters
A P
AP
del Portillo
H A
HA
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle / Q000502:physiology*; D003001:Cloning, Molecular; D018844:Cyclin-Dependent Kinases / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D018076:DNA, Complementary / Q000737:chemistry; D005786:Gene Expression Regulation; D015723:Gene Library; D005816:Genetic Complementation Test; D006801:Humans; D016780:Malaria, Vivax / Q000469:parasitology; D008969:Molecular Sequence Data; D010966:Plasmodium vivax / Q000166:cytology / Q000235:genetics*; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics*; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012568:Schizosaccharomyces / Q000166:cytology* / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
C077833:CRK protein, protozoan; D018076:DNA, Complementary; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D011494:Protein Kinases; D042846:CDC2-CDC28 Kinases; D018844:Cyclin-Dependent Kinases