Signal transduction in Pneumocystis carinii: Characterization of the genes (pcg1) encoding the alpha subunit of the G protein (PCG1) of Pneumocystis carinii carinii and Pneumocystis carinii ratti

Abstract

Pneumocystis carinii is a eukaryotic organism that causes pneumonia in immunocompromised hosts. The cell biology and life cycle of the organism are poorly understood primarily because of the lack of a continuous in vitro cultivation system. These limitations have prevented investigation of the organism's infectious cycle and hindered the rational development of new antimicrobial therapies and implementation of measures to prevent exposure to the organism or transmission. The interaction of P. carinii with its host and its environment may be critical determinants of pathogenicity and life cycle. Signal transduction pathways are likely to be critical in regulating these processes. G proteins are highly conserved members of the pathways important in many cellular events, including cell proliferation and environmental sensing. To characterize signal transduction pathways in P. carinii, we cloned a G-protein alpha subunit (G-α) of P. carinii carinii and P. carinii ratti by PCR amplification and hybridization screening. The gene encoding the G-α was present in single copy on a 450-kb chromosome of P. c. carinii and on a 420-kb chromosome of P. c. ratti. The 1,062-bp G-α open reading frame is interrupted by nine introns. The predicted polypeptide showed 29 to 53% identity with known fungal G-α proteins with greatest homology to Neurospora crassa Gna-2. Northern (RNA) blot analysis and immunoprecipitation demonstrated expression of the G-α mRNA and protein in P. carinii isolated from heavily infected animals. Some alteration in the level of transcription was noted in short-term maintenance in starvation or rich medium. Characterization of signal transduction in P. carinii will permit a better understanding of the reproductive capacity and other cellular processes in this family of organisms that cannot be cultured continuously.