Gp96/GRP94 is a putative high density lipoprotein-binding protein in liver.
We have previously shown that three high density lipoproteins (HDL)-binding proteins in liver, of 90, 110 and 180 kDa, are structurally related. In this study, these proteins are identified as gp96/GRP94. This protein is known to occur as a homodimer and has a dual subcellular localization: it is both an endoplasmic reticulum resident protein, where it is supposed to act as a chaperonin, and a plasma membrane protein, whose significance is unknown. In ultrastructural studies the plasma membrane localization of the homodimeric form was verified. The 90-kDa protein was abundantly present at the membranes of the endosomal/lysosomal vesicles as well as at the apical hepatocyte membranes, comprising the bile canaliculi. The monomeric protein is scarcely present at the basolateral membrane of the hepatocytes, but could be demonstrated in coated pits, suggesting involvement in receptor-mediated endocytosis. Labeling of the endoplasmic reticulum was virtually absent. Gp96/GRP94 was transiently expressed in COS-1 cells. However, the expressed protein was exclusively localized in the endoplasmic reticulum. Transfection with constructs in which the C-terminal KDEL sequence had been deleted, resulted in plasma membrane localized expression of protein, but only in an extremely low percentage of cells. In order to evaluate the HDL-binding capacities of this protein, stably transfected cells were generated, using several cell types. It appeared to be difficult to obtain a prolonged high level expression of gp96. In these cases, however, a marked increase of HDL-binding activity compared with the control cells could be observed.
de Crom
R
R
The Medical Genetics Center, Department of Cell Biology and Genetics, Erasmus University Rotterdam, Dr. Molenwaterplein 50, P.O. Box 1738, 3000 DR, Rotterdam, The Netherlands.decrom@ch1.fgg.eur.nl
van Haperen
R
R
Janssens
R
R
Visser
P
P
Willemsen
R
R
Grosveld
F
F
van der Kamp
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D013552:Swine; D014162:Transfection
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; C087410:Hdlbp protein, rat; D008565:Membrane Proteins; C018551:glucose-regulated proteins