Enhancement of the Binding of Triglyceride-rich Lipoproteins to the Very Low Density Lipoprotein Receptor by Apolipoprotein E and Lipoprotein Lipase

Abstract

The low-density lipoprotein (LDL) receptor plays a crucial role in cholesterol metabolism. A related protein, designated the very low density lipoprotein (VLDL) receptor, that specifically binds apolipoprotein (apo) E has recently been characterized and shown to be expressed in heart, muscle and adipose tissue and the human monocyte-macrophage cell line THP-1. The VLDL receptor binds and internalizes VLDL and intermediate density lipoprotein from Watanabe heritable hyperlipidemic (WHHL) rabbits as well as beta-migrating VLDL from cholesterol-fed rabbits but not LDL from WHHL rabbits. Chinese hamster ovary (CHO) cells transfected with the rabbit VLDL receptor cDNA have now been shown to bind or internalize VLDL (d < 1.006 g/ml) isolated from fasted normolipidemic human subjects with lower affinity than WHHL-VLDL or rabbit beta-VLDL. However, binding and internalization were markedly enhanced when fasted human VLDL was preincubated with either recombinant human apoE (3/3) or lipoprotein lipase (LPL) in CHO cells overexpressing the rabbit or human VLDL receptor. CHO cells transfected with both the rabbit VLDL receptor cDNA and the human LPL cDNA effectively bound, internalized, and degraded fasted human VLDL without pretreatment. Treatment of heparinase reduced the effect of LPL-mediated binding at 4 degrees C, but the inhibitory effect was lower at 37 degrees C. Pseudomonas LPL also enhanced the binding of human fasted VLDL to the VLDL receptor at 37 degrees C in CHO cells overexpressing the human VLDL receptor. Taken together, LPL causes the enhancement of triglyceride-rich lipoproteins binding to the VLDL receptor via both the formation of bridge between lipoproteins and heparan sulfate proteoglycans and its lipolytic effect. Ligand blot analysis showed that the apparent molecular mass of the VLDL receptor is 118 kDa, which is smaller than that of the LDL receptor. These results indicate that the VLDL receptor recognizes both triglyceride-rich lipoproteins that are also relatively rich in apoE, as well as the remnants of triglyceride-rich lipoproteins after catabolism and the interaction with heparan sulfate proteoglycans by LPL. The VLDL receptor may thus function as a receptor for remnants of triglyceride-rich lipoproteins in extrahepatic tissues.

Very low density lipoprotein receptor binds apolipoprotein E2/2 as well as apolipoprotein E3/3

“…Human VLDL obtained from fasted subjects has a significantly reduced ability to bind to the VLDL receptor compared with rabbit 13-VLDL. But apoE and LPL, which are secreted from macrophages, enhanced the binding of human VLDL to the VLDL receptor [12], supporting the hypothesis that the VLDL receptor pathway may contribute to foam cell formation.…”

“…Plasmids encoding the human VLDL receptor (ph-VLR2) or pSV2-neo, which contains the neomycin resistance gene, were transfected to ldlA-7 cells (mutant Chinese hamster ovary cells that lack LDL receptors) which was kindly provided by Dr. Monty Krieger (Department of Biology, Massachusetts Institute of Technology, Cambridge, MA) using lipofectin as described previously [1][2][3]9,12]. Positive cell clones for the human VLDL receptor were designated VLDL-R cells.…”

“…Cellular protein was measured according to the method of Lowry et al [11]. To exclude the effect of LRP (LDL receptor-related protein), specific activity was determined by subtracting the radioactivity associated with Neo cells from that associated with VLDL-R cells [12]. All binding assays were performed in triplicate.…”

“…receptor Human fasted [12~I]VLDL from normolipidemic subjects has been shown to bind the VLDL receptor with lower affinity compared with 13-VLDL from cholesterol-fed rabbits [12]. Competitive binding assays in cell culture were performed using [125I]rabbit 13-VLDL instead of human VLDL.…”

Nascent very-low-density lipoprotein triacylglycerol hydrolysis by lipoprotein lipase is inhibited by apolipoprotein E in a dose-dependent manner

“…Its most well-defined function is to mediate the high-affinity binding of chylomicron and VLDL remnants to the low-density-lipoprotein (LDL) receptor [1], the LDL-receptor-related protein (LRP) [2] and VLDL receptor [3]. In addition, apoE is postulated to play a role in the secretionrecapture process [4,5].…”