This paper provides further characterization of a receptor that, in cells lacking the LDL receptor (FH fibroblasts), mediates lipoprotein binding, uptake, and degradation when incubated with oleate at concentrations not exceeding albumin binding capacity. This oleate-activated receptor is genetically distinct from the LDL receptor and is hereafter referred to as the lipolysis-stimulated receptor (LSR). Its apparent affinity was higher for triglyceride-rich lipoproteins (chylomicrons, VLDL) and for lipid emulsions supplemented with recombinant apoE, than for LDL which contains solely apoB. In contrast, VLDL isolated from a Type III hyperlipidemic patient (apoE2/2 phenotype) failed to bind to the LSR. Five lines of evidence indicated that the LSR is distinct from the LDL receptor-related protein (LRP): (1) the LRP ligand, alpha 2-macroglobulin-methylamine (alpha 2-MG*), did not bind to the oleate-induced LDL binding site; (2) oleate had no effect on the binding of alpha 2-MG* to LRP; (3) the LRP-associated protein, RAP, which inhibits LRP, had no effect on the LSR; (4) binding of lipoproteins to LSR was independent of Ca2+; and (5) LSR activity resolved as two proteins smaller than LRP (apparent molecular masses as determined by ligand blots: 115 and 85 kDa). That LSR provides a new candidate receptor contributing to the clearance of chylomicron remnants (CMR) is supported by the observation that LSR was inhibited by lactoferrin, a milk protein that delays CMR clearance when injected in vivo. Furthermore, in primary cultures of rat hepatocytes, oleate stimulated binding, uptake, and degradation of LDL with kinetic characteristics similar to that of LSR expressed in FH fibroblasts.(ABSTRACT TRUNCATED AT 250 WORDS)

In cultured human and rat cells, the lipolysis-stimulated receptor (LSR), when activated by free fatty acids (FFA), mediates the binding of apoprotein B- and apoprotein E-containing lipoproteins and their subsequent internalization and degradation. To better understand the physiological role of LSR, we developed a biochemical assay that optimizes both the activation and binding steps and, thus, allows the estimation of the number of LSR binding sites expressed in the livers of living animals. With this technique, a strong inverse correlation was found in rats between the apparent number of LSR binding sites in liver and the postprandial plasma triglyceride concentration (r = -0.828, p < 0.001, n = 12). No correlation existed between the number of LSR and plasma triglycerides measured in the same animals after 24 h of fasting. The same membrane binding assay was used to elucidate the mechanism by which FFA induce lipoprotein binding to LSR. The LSR activation step was mediated by direct interaction of FFA with LSR candidate proteins of apparent molecular masses of 115 and 90 kDa and occurred independently of the membrane lipid environment. The FFA-induced conformational shift that revealed the lipoprotein binding site remained fully reversible upon removal of the FFA. However, occupancy of the site by the apoprotein ligand stabilized the active form of LSR. Comparison of the effect of different FFA alone or in combination indicated that the same binding site is revealed by different FFA and that the length and saturation of the FFA monomeric carbon chain are critical in determining the potency of the FFA activating effect. We propose that the LSR pathway represents a limiting step for the cellular uptake of intestinally derived triglyceride-rich lipoproteins and speculate that FFA liberated by lipolysis initiate this process by altering the conformation of LSR to reveal the lipoprotein binding site.

No abstract

No abstract