Regulated export of a secretory protein from the ER of the hepatocyte: a specific binding site retaining C-reactive protein within the ER is downregulated during the acute phase response.
The half-time for secretion of the plasma protein C-reactive protein (CRP) by the hepatocyte decreases markedly in association with its increased synthesis during the acute phase response to tissue injury (Macintyre, S., D. Samols, and I. Kushner. 1985. J. Biol. Chem. 260:4169-4173). In studies in which subcellular fractions were prepared from cells incubated under pulse-chase conditions, CRP was found to be preferentially retained within the ER of normal hepatocytes, but secreted relatively efficiently in cells prepared from rabbits undergoing the acute phase response. On the basis of the detergent-dependency of specific binding of radiolabeled CRP, as well as EM visualization of biotinylated CRP identified with peroxidase-conjugated streptavidin, CRP was found to bind to the lumenal surface of permeabilized rough microsomes, while no binding was detected in Golgi fractions. As judged by both kinetic and equilibrium binding studies, rough microsomes from control rabbits were found to have two classes of specific binding sites for CRP; a high affinity site (Kd = 1 nM, Bmax = 1 pmol CRP/mg microsomal protein) as well as a much lower affinity (Kd = 140 nM) site. In contrast, only the lower affinity class was detected in microsomes isolated from rabbits undergoing the acute phase response. On nitrocellulose blots probed with radiolabeled CRP a 60-kD protein, distinct from BiP, was detected in extracts of rough microsomes isolated from control rabbits, but not in Golgi fractions or rough microsomes from stimulated animals. These findings correlate with previous observations of changes in secretion kinetics of CRP and are consistent with the hypothesis that the intracellular sorting of CRP could be rerouted by downregulation of a specific ER binding site during the acute phase response.
Macintyre
S S
SS
Case Western Reserve University, MetroHealth Medical Center, Cleveland, Ohio 44109-1998.
D000209:Acute-Phase Proteins / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002097:C-Reactive Protein / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology; D002458:Cell Fractionation; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D003840:Deoxycholic Acid / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure; D007700:Kinetics; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008854:Microscopy, Electron; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D011817:Rabbits; D011971:Receptors, Immunologic / Q000302:isolation & purification / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
D000209:Acute-Phase Proteins; C071581:CRP protein, human; D011971:Receptors, Immunologic; D003840:Deoxycholic Acid; D002097:C-Reactive Protein; D002118:Calcium