Cell surface proteoglycans modulate net synthesis and secretion of macrophage apolipoprotein E.
Using a macrophage cell line that constitutively expresses a human apolipoprotein E (apoE) cDNA, we have investigated the post-translational metabolism of endogenously produced apoE. Inhibition of lysosomal or cysteine proteases led to significant inhibition of apoE degradation but did not increase apoE secretion, indicating that cellular degradation is not limiting for apoE secretion in macrophages. Treatment of macrophages with inhibitors of proteoglycan synthesis (4-methylumbelliferyl-beta-D-xyloside) or sulfation (sodium chlorate) enhanced the release of apoE from cells and significantly attenuated the increase in secretion produced by incubation with phosphatidylcholine vesicles (PV). These observations suggested that a significant fraction of the apoE retained by cells (and released by incubation with PV) was associated with proteoglycans. Treatment of cells with exogenous heparinase led to a greater than 4-fold increase in apoE secretion and similarly attenuated the response to PV, suggesting that apoE was trapped in an extracellular proteoglycan matrix. This conclusion was confirmed in studies showing that PV could enhance the release of apoE from cells during an incubation at 4 degrees C, but this enhanced release was abolished in proteoglycan-depleted cells. Incubation with lactoferrin at 4 or 37 degrees C produced a similar decrement in cellular apoE, again indicating the existence of a cell surface pool of apoE. Pulse-chase studies showed that the apoE trapped in the proteoglycan matrix was susceptible to rapid cellular degradation such that net synthesis of apoE (secreted plus cell-associated) was increased significantly in proteoglycan-depleted cells compared with control cells as early as 45 min during a chase period.
Lucas
M
M
Department of Medicine, Rush Medical College, Chicago, Illinois 60612, USA.
Mazzone
T
T
D001057:Apolipoproteins E / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D019763:Heparin Lyase; D006801:Humans; D007781:Lactoferrin / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D008954:Models, Biological; D010713:Phosphatidylcholines / Q000494:pharmacology; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection
D001057:Apolipoproteins E; D018076:DNA, Complementary; D010713:Phosphatidylcholines; D011480:Protease Inhibitors; D011509:Proteoglycans; D007781:Lactoferrin; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase