G. Guadiz et al., THE CARBOXYL-TERMINUS OF PNEUMOCYSTIS-CARINII GLYCOPROTEIN-A ENCODES A FUNCTIONAL GLYCOSYLPHOSPHATIDYLINOSITOL SIGNAL SEQUENCE, The Journal of biological chemistry, 273(40), 1998, pp. 26202-26209
Pneumocystis carinii pneumonia is a hallmark disease associated with A
IDS. An abundant glycoprotein, termed gpA, on the surface of P. carini
i is considered an important factor in host-parasite interactions. The
primary structure of ferret P. carinii gpA contains a carboxyl-termin
al sequence characteristic of a signal for glycosylphosphatidylinosito
l (GPI) anchors. Here we report the capacity for this gpA carboxyl seq
uence to direct attachment of a secreted protein, human growth hormone
(hGH), to the membranes of COS cells. A control fusion protein (hGHDA
F37) was obtained which, under the direction of the GPI signal from de
cay accelerating factor, directs hGR cell surface expression. A constr
uct (phGH2-1A30) was created similar to hGHDAF37 by fusing hGH to the
putative GPI signal sequence encoded in the terminal 30 residues from
a ferret P. carinii gpA cDNA clone. By indirect immunofluorescent stai
ning, hGH was detected on the surface of COS cells transfected with ph
GH2-1A30; this surface location was confirmed by confocal laser cytome
try. Metabolic labeling with [H-3]ethanolamine and subsequent immunopu
rification of hGH from cells transfected with phGH2-1A30 confirmed tha
t a lipid moiety characteristic of a conventional GPI anchor was linke
d covalently to hGH, and cell surface hGH2-1A30 fusion protein was sen
sitive to enzymatic cleavage by phosphatidylinositol-phospholipase C.
Furthermore, hGH2-1A30 recombinant protein cofractionated with 5'-nucl
eotidase, a classical GPI-anchored membrane marker. Together, these re
sults indicate that the carboxyl-terminal residues of ferret P. carini
i gpA constitute a biologically functional GPI consensus domain, thus
providing a potential mechanism for antigenic variation of P. carinii
gpA during P. carinii pneumonia.