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Pneumocystis carinii contains a functional cell-division-cycle Cdc2 homologue
C F Thomas1, R A Anders, M P Gustafson
1Division of Pulmonary, Critical Care and Internal Medicine, Mayo Clinic and Foundation, Rochester, Minnesota 55905, USA.
Abstract:
Pneumocystis carinii causes life-threatening pneumonia in immunocompromised patients. The inability to culture P. carinii has hampered basic investigations of the organism's life cycle, limiting the development of new therapies directed against it. Recent investigations indicate that P. carinii is a fungus phylogenetically related to other ascomycetes such as Schizosaccharomyces pombe. The cell cycles of S. pombe and homologous fungi are carefully regulated by cell-division-cycle molecules (cdc), particularly cell-division-cycle 2 (Cdc2), a serine-threonine kinase with essential activity at the G1 restriction point and for entry into mitosis. Antibodies to the proline-serine-threonine-alanine-isoleucine-arginine (PSTAIR) amino-acid sequence conserved in Cdc2 proteins specifically precipitated, from P. carinii extracts, a molecule with kinase activity consistent with a Cdc2-like protein. Cdc2 molecules exhibit differential activity throughout the life cycle of the organisms in which they occur. In accord with this, the P. carinii Cdc2 showed greater specific activity in P. carinii trophic forms (trophozoites) than in spore-case forms (cysts). In addition, complete genomic and complementary DNA (cDNA) sequences of P. carinii Cdc2 were cloned and found to be most closely homologus to the corresponding sequences of other pathogenic fungi. The function of P. carinii cdc2 cDNA was further documented through its ability to complement the DNA of mutant strains of S. pombe with temperature-sensitive deficiencies in Cdc2 activity. The P. carinii cdc2 cDNA restored normal Cdc2 function in these mutant strains of S. pombe, and promoted fungal proliferation. These studies represent the first molecular analysis of the cell-cycle-regulatory machinery in P. carinii. Further understanding of P. carinii's life cycle promises novel insights for preventing and treating the intractable infection it causes in immunocompromised patients.
Insights
Researchers identified a cell-division-cycle 2 (Cdc2) protein in Pneumocystis carinii, crucial for fungal growth. This discovery offers new therapeutic targets for treating P. carinii pneumonia in immunocompromised individuals.
Area of Science:
- Mycology
- Molecular Biology
- Medical Microbiology
Background:
- Pneumocystis carinii causes severe pneumonia in immunocompromised patients.
- Limited understanding of P. carinii's life cycle hinders new therapy development.
- P. carinii is a fungus related to other ascomycetes, with cell cycles regulated by cell-division-cycle (cdc) molecules.
Purpose of the Study:
- To investigate the cell-cycle regulation in P. carinii.
- To identify and characterize a Cdc2-like protein in P. carinii.
- To explore the potential of P. carinii cdc2 as a therapeutic target.
Main Methods:
- Used antibodies against the conserved PSTAIR sequence to precipitate a Cdc2-like protein from P. carinii extracts.
- Assayed kinase activity of the precipitated protein.
- Cloned and sequenced the genomic and cDNA of P. carinii cdc2.
- Complemented temperature-sensitive mutant strains of Schizosaccharomyces pombe with P. carinii cdc2 cDNA.
Main Results:
- A protein with kinase activity consistent with Cdc2 was identified in P. carinii.
- P. carinii Cdc2 showed higher activity in trophic forms than in cyst forms.
- The cloned P. carinii cdc2 cDNA restored normal cell cycle function and promoted proliferation in S. pombe mutants.
- P. carinii cdc2 is homologous to Cdc2 proteins in other pathogenic fungi.
Conclusions:
- This study provides the first molecular analysis of cell-cycle regulation in P. carinii.
- P. carinii Cdc2 is a functional homolog of S. pombe Cdc2.
- Understanding P. carinii's cell cycle machinery offers potential for novel therapeutic strategies against P. carinii infections.