CHARACTERIZATION OF RECOMBINANT HUMAN APO-B-48-CONTAINING LIPOPROTEINS IN RAT HEPATOMA MCA-RH7777 CELLS TRANSFECTED WITH APO-B-48 CDNA OVEREXPRESSION OF APO-B-48 DECREASES SYNTHESIS OF ENDOGENOUS APO-B-100

We studied the effect of overexpression of apolipoprotein (ape) B-48 o n the synthesis and secretion of endogenous apoB-100 in rat hepatoma M cA-RH7777 cell lines stably transfected with human apoB-48 cDNA under the control of the cytomegalovirus promoter. Three cell lines that sec rete 40 to 60 ng human apoB . mg cell protein(-1) . h(-1) were used. T he recombinant human apoB-48 exhibited physicochemical characteristics (buoyant density, 1.06 to 1.21 g/mL; beta-electrophoretic mobility an d diameters, 16 to 20 nm) indistinguishable from those of endogenous r at apoB-48. Overexpression of the recombinant human apoB-48 resulted i n a 50% decrease in the secretion of endogenous apoB-100 but did not a ffect the secretion of apoE or apoA-I. Several possible mechanisms for the decreased secretion of apoB-100 were evaluated. First, recruitmen t of lipids into lipoproteins was shown to be unaffected since no majo r changes in the physicochemical properties of apoB-100-containing lip oproteins were observed. Second, the intracellular degradation of apoB -100 was not altered as the intracellular retention half-time and secr etion efficiency remained unaffected by apoB-48 overexpression. Third, the posttranslational regulatory mechanisms for apoB-100 remained nor mal, as demonstrated by a twofold increase in apoB-100 secretion after supplementation with oleic acid. Unexpectedly, a 35% to 50% decrease in the steady-state synthesis of endogenous apoB-100 was observed in a poB-48-transfected cells compared with control cells. These data sugge sted that decreased secretion of apoB-100 was secondary to decreased s ynthesis. The decreased apoB-100 synthesis was not due to decreased st eady-state levels of rat apoB-100 mRNA. These results suggest that ove rexpression of recombinant human apoB-48 may interfere with posttransc riptional events, possibly at the translation-translocation level, and decrease translational yield of apoB-100. These posttranscriptional e vents prior to the complete synthesis of the apoB-100 polypeptide can be important in the control of apoB-100 secretion.