THE CARBOXYL-TERMINUS OF PNEUMOCYSTIS-CARINII GLYCOPROTEIN-A ENCODES A FUNCTIONAL GLYCOSYLPHOSPHATIDYLINOSITOL SIGNAL SEQUENCE

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.