The 78 kDa glucose-regulated protein (GRP78/BIP) is expressed on the cell membrane, is released into cell culture medium and is also present in human peripheral circulation.
In human rabdomiosarcoma cells (TE671/RD) chronic exposure to 500 nM thapsigargin (a powerful inhibitor of the endoplasmic reticulum Ca2+-ATPases) resulted in the induction of the stress protein GRP78/BIP. Making use of the surface biotinylation method, followed by the isolation of the GRP78 using ATP-agarose affinity chromatography, it was found that a fraction of the thapsigargin-induced GRP78 is expressed on the cell surface. The presence of GRP78 on the membrane of thapsigargin-treated cells was confirmed by fractionation of cell lysates into a soluble and a membrane fraction, followed by Western blot analysis with an anti-GRP78 antibody. It was also found that conspicuous amounts of GRP78 are present in the culture medium collected from thapsigargin-treated cultures. This extracellular GRP78 originates mostly by an active release from intact cells and does not result solely from the leakage of proteins from dead cells. Moreover, small amounts of circulating, free GRP78 and naturally-occurring anti-GRP78 autoantibodies were detected in the peripheral circulation of healthy human individuals.
Delpino
Andrea
A
Regina Elena Cancer Institute, Center for Experimental Research, Via delle Messi d'Ora 156 00158 Roma, Italy. castelli@ifo.it
Castelli
Mauro
M
D001323:Autoantibodies / Q000097:blood; D002352:Carrier Proteins / Q000097:blood / Q000276:immunology / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D003470:Culture Media / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006360:Heat-Shock Proteins; D006801:Humans; D008297:Male; D018832:Molecular Chaperones / Q000097:blood / Q000276:immunology / Q000378:metabolism*; D012208:Rhabdomyosarcoma / Q000188:drug therapy / Q000378:metabolism; D019284:Thapsigargin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D001323:Autoantibodies; D002352:Carrier Proteins; D003470:Culture Media; D000091342:Endoplasmic Reticulum Chaperone BiP; D004791:Enzyme Inhibitors; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D019284:Thapsigargin