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Very-low-density lipoprotein binding to the apolipoprotein E receptor 2 is enhanced by lipoprotein lipase, and does not require apolipoprotein E.

ABSTRACT: The apolipoprotein (apo)E receptor 2 (apoER2) is a recently cloned member of the low-density lipoprotein (LDL) receptor (LDLR) family, showing a high homology with both the LDLR and the very-low-density lipoprotein (VLDL) receptor (VLDLR). In the present study, the binding characteristics of the apoER2 with respect to apoE and lipoprotein lipase (LPL) were investigated. VLDL was isolated from both apoE-deficient mice and mice expressing the human APOE2 (Arg(158)-->Cys) and APOE3-Leiden isoforms on an Apoe(-/-),Ldlr(-/-) double knock-out background. apoE-rich rabbit beta-VLDL was used as a positive control for binding. Binding experiments performed with Chinese hamster ovary cells expressing the human apoER2 showed that the receptor was able to bind VLDL containing either of the apoE isoforms, as well as the apoE-deficient VLDL. Hence, in contrast with the VLDLR, the apoER2 is not strictly dependent on apoE for VLDL binding. Since LPL has been shown to enhance the binding of lipoproteins to several members of the LDLR family, including the LDLR-related protein, VLDL receptor, gp330 and the LDLR itself, VLDL binding experiments were performed in the presence of LPL. Addition of LPL resulted in a significant increase in apoER2 binding for all VLDL fractions used in this study. In conclusion, lipoprotein binding of VLDL to the apoER2 is enhanced in the presence of LPL, and is not restricted to apoE-containing lipoproteins.

SUBMITTER: Tacken PJ

PROVIDER: S-EPMC1220966 | biostudies-other | 2000 Apr

REPOSITORIES: biostudies-other

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Project description:We have shown previously that human apolipoprotein (apo)C1 transgenic mice exhibit hyperlipidaemia, due primarily to an impaired clearance of very-low-density lipoprotein (VLDL) particles from the circulation. In the absence of at least the low-density-lipoprotein receptor (LDLR), it was shown that APOC1 overexpression in transgenic mice inhibited the hepatic uptake of VLDL via the LDLR-related protein. In the present study, we have now examined the effect of apoC1 on the binding of lipoproteins to both the VLDL receptor (VLDLR) and the LDLR. The binding specificity of the VLDLR and LDLR for apoC1-enriched lipoprotein particles was examined in vivo through adenovirus-mediated gene transfer of the VLDLR and the LDLR [giving rise to adenovirus-containing (Ad)-VLDLR and Ad-LDLR respectively] in APOC1 transgenic mice, LDLR-deficient (LDLR-/-) mice and wild-type mice. Remarkably, Ad-VLDLR treatment did not reduce hyperlipidaemia in transgenic mice overexpressing human APOC1, irrespective of both the level of transgenic expression and the presence of the LDLR, whereas Ad-VLDLR treatment did reverse hyperlipidaemia in LDLR-/- and wild-type mice. On the other hand, Ad-LDLR treatment strongly decreased plasma lipid levels in these APOC1 transgenic mice. These results suggest that apoC1 inhibits the clearance of lipoprotein particles via the VLDLR, but not via the LDLR. This hypothesis is corroborated by in vitro binding studies. Chinese hamster ovary (CHO) cells expressing the VLDLR (CHO-VLDLR) or LDLR (CHO-LDLR) bound less APOC1 transgenic VLDL than wild-type VLDL. Intriguingly, however, enrichment with apoE enhanced dose-dependently the binding of wild-type VLDL to CHO-VLDLR cells (up to 5-fold), whereas apoE did not enhance the binding of APOC1 transgenic VLDL to these cells. In contrast, for binding to CHO-LDLR cells, both wild-type and APOC1 transgenic VLDL were stimulated upon enrichment with apoE. From these studies, we conclude that apoC1 specifically inhibits the apoE-mediated binding of triacylglycerol-rich lipoprotein particles to the VLDLR, whereas apoC1-enriched lipoproteins can still bind to the LDLR. The variability in specificity of these lipoprotein receptors for apoC1-containing lipoprotein particles provides further evidence for a regulatory role of apoC1 in the delivery of lipoprotein constituents to different tissues on which these receptors are located.

Project description:The function of apolipoprotein (apo) C1 in vivo is not clearly defined. Because transgenic mice overexpressing human apoC1 show elevated triacylglycerol (TG) levels [Simonet, Bucay, Pitas, Lauer and Taylor (1991) J. Biol. Chem. 266, 8651-8654], an as yet unknown role for apoC1 in TG metabolism has been suggested. Here we investigated directly the effect of the complete absence of apoC1 on very-low-density lipoprotein (VLDL)-TG lipolysis, clearance and production, by performing studies with the previously generated apoC1-deficient mice. On a sucrose-rich, low fat/low cholesterol (LFC) diet, apoC1-deficient mice accumulate in their circulation VLDL particles, which contain relatively lower amounts of lipids when compared with VLDL isolated from control mice. Lipolysis assays in vitro on VLDL from apoC1-deficient and control mice showed no differences in apparent K(m) and Vmax values (0.27 +/- 0.06 versus 0.24 +/- 0.03 mmol of TG/litre and 0.40 +/- 0.03 versus 0.36 +/- 0.03 mmol of non-esterified fatty acid (NEFA)/min per litre respectively). To correct for potential differences in the size of the VLDL particles, the resulting K(m) values were also expressed relative to apoB concentration. Under these conditions apoC1-deficient VLDL displayed a lower, but not significant, K(m) value when compared with control VLDL (3.44 +/- 0.71 versus 4.44 +/- 0.52 mmol of TG2/g apoB per litre). VLDL turnover studies with autologous injections of [3H]TG-VLDL in vivo showed that the VLDL fractional catabolic rate (FCR) was decreased by up to 50% in the apoC1-deficient mice when compared with control mice (10.5 +/- 3.4 versus 21.0 +/- 1.2/h of pool TG). No significant differences between apoC1-deficient and control mice were observed in the hepatic VLDL production estimated by Triton WR139 injections (0.19 +/- 0.02 versus 0.21 +/- 0.05 mmol/h of TG per kg) and in the extra-hepatic lipolysis of VLDL-TG (4.99 +/- 1.62 versus 3.46 +/- 1.52/h of pool TG) in vivo. Furthermore, [125I]VLDL-apoB turnover experiments in vivo also showed a 50% decrease in the FCR of VLDL in apoC1-deficient mice when compared with control mice on the LFC diet (1.1 +/- 0.3 versus 2.1 +/- 0.1/h of pool apoB). When mice were fed a very high fat/high cholesterol (HFC) diet, the VLDL-apoB FCR was further decreased in apoC1-deficient mice (0.4 +/- 0.1 versus 1.4 +/- 0.4/h of pool apoB). We conclude that, in apoC1-deficient mice, the FCR of VLDL is reduced because of impaired uptake of VLDL remnants by hepatic receptors, whereas the production and lipolysis of VLDL-TG is not affected.

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Very-low-density lipoprotein binding to the apolipoprotein E receptor 2 is enhanced by lipoprotein lipase, and does not require apolipoprotein E.

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