Pneumocystis carinii cell wall biosynthesis kinase gene CBK1 is an environmentally responsive gene that complements cell wall defects of cbk-deficient yeast.
Pneumocystis species remain an important cause of life-threatening pneumonia in immunocompromised hosts, including those with AIDS. Responses of the organism to environmental cues both within the lung and elsewhere have been poorly defined. Herein, we report the identification of a cell wall biosynthesis kinase gene (CBK1) homologue in Pneumocystis carinii, isolated by differential display PCR, that is expressed optimally at physiological pH (7 to 8) as opposed to more acidic environments. Expression of Pneumocystis CBK1 was also induced by contact with lung epithelial cells and extracellular matrix. Translation of this gene revealed extensive homology to other fungal CBK1 kinases. Pneumocystis CBK1 expression was equal in the cyst and trophic life forms of the organisms. We further demonstrate that Pneumocystis CBK1 expressed in cbk1 Delta Saccharomyces cerevisiae cells restored defective cell wall separation during proliferation. Consistent with this, Pneumocystis CBK1 expression also stimulated transcription of the CTS1 chitinase in cbk1 Delta mutant yeast cells, an event necessary for cell wall separation. In addition, Pneumocystis CBK1 cDNA supported normal mating projection formation in response to alpha-factor in the cbk1 Delta yeast cells. Site-directed mutations of serine-303 and threonine-494, potential regulatory phosphorylation sites in Pneumocystis CBK1, abolished mating projection formation, indicating a role for these amino acid residues in CBK1 activity. These findings indicate that Pneumocystis CBK1 is an environmentally responsive gene that may function in signaling pathways necessary for cell growth and mating.
Thoracic Diseases Research Unit, Department of Medicine, Mayo Clinic, Rochester, MN 55905, USA.
Theodore J
TJ
Kottom
Andrew H
AH
Limper
D000595:Amino Acid Sequence; D000818:Animals; D002448:Cell Adhesion; D002473:Cell Wall / Q000235:genetics / Q000378:metabolism*; D003470:Culture Media; D004777:Environment; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005816:Genetic Complementation Test; D018869:Heat-Shock Response; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D045363:Pneumocystis carinii / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D017422:Sequence Analysis, DNA
D003470:Culture Media; D005656:Fungal Proteins; D017346:Protein Serine-Threonine Kinases