Project: Research project

Project Details


Pneumocystis carinii causes severe pneumonia in immunocompromised patients
with AIDS. Recent investigations have advanced our understanding of host-
organism interactions during P. carinii pneumonia. These studies indicate
that attachment of P. carinii to alveolar epithelial cells is an integral
event which promotes completion of the organism's life cycle. Continuous
culture of P. carinii in a cell-free environment continues to be elusive.
However, the organism can multiply on cultured respiratory epithelial
cells. Attachment of P. carinii trophozoites to epithelial cells occurs by
the close apposition of the organism's cell membrane with the membrane of
the host. Further studies demonstrate that the attachment of P. carinii to
host cells is largely potentiated by the adhesive proteins vitronectin
(VN) and fibronectin (FN) present in the alveolar space. However, the role
of these attachment mechanisms in promoting completion of the organism's
life cycle have not been fully defined. In addition, the attachment of P.
carinii to the host, also suppresses division of respiratory epithelial
cells, thereby slowing repair of lung injury. The mechanisms by which P.
carinii attachment promotes completion of the organism's life cycle, while
simultaneously impairing the cell cycle of the host are largely unknown.
The eukaryotic cell cycle is carefully regulated by a group of cell
division cycle (cdc) molecules including cyclins and cyclin-dependent
kinases. Of critical importance is the cdc2 gene which encodes a
serine/threonine kinase. The timing of cdc2 activation is controlled in
part by the cdc25 gene which activates the cdc2 gene product in late G2.
Homologues of cdc2 and cdc25 have been identified in a variety of
eukaryotic organisms from Schizosaccharomyces pombe, a fungus closely
related to P. carinii, through man. To date, very little has been learned
about cell cycle control molecules in P. carinii. Recently, our laboratory
has demonstrated that P. carinii possesses a protein containing the PSTAIR
amino-acid sequence conserved in cdc2 gene products in all eukaryotes. We
hypothesize that attachment of P. carinii to respiratory epithelial cells
promotes life cycle completion and proliferation of P. carinii. We propose
that P. carinii attachment by VN and FN-mediated mechanisms alters the
expression and/or activity of homologous cdc2 and cdc25 gene products in
P. carinii. We further postulate that P. carinii attachment also inhibits
the cell cycle of host epithelial cells, through altered activity of the
mammalian cyclins and cyclin dependent kinases. We will evaluate whether
attachment of P. carinii to respiratory epithelial cells through the
adhesive proteins VN and FN promotes life cycle progression in the
organism. Further, we will identify specific P. carinii cell cycle control
molecules using a complementation cloning strategy. Mutants of S. pombe
which lack cdc2 and cdc2S have been derived and successfully used to clone
homologous molecules from other species. Therefore, we will use
complementation to clone the P. carinii homologues of cdc2 and cdc25.
Next, the expression and activity of these cell cycle control-molecules
will be studied in purified populations of P. carinii trophozoites and
cysts. Finally, we will determine the mechanisms by which P. carinii
attachment to epithelial cells mediates alterations in the cell cycles of
both the organism and the host promises to yield new therapeutic insights
in P. carinii pneumonia.
Effective start/end date9/30/958/31/10


  • Medicine(all)