Low density lipoprotein-receptor plays a major role in the binding of verylow density lipoproteins and their remnants on HepG2 cells

The binding to HepG2 cells of very low density lipoproteins (VLDL) and thei r remnants (IDL) was alternatively, in the past, attributed to the low dens ity lipoprotein receptor (LDLr) or to an apoE-specific receptor. In order t o resolve this issue, we have compared the binding of those lipoproteins la belled with iodine-125 to normal and LDLr deficient HepG2 cells. Those defi cient cells were obtained by a constitutive antisense strategy and their LD Lr level is 14% the level of normal HepG2 cells. By saturation curve analys is, we show that VLDL and IDL bind to high and low affinity sites on cells. The low affinity binding was eliminated bl conducting the assay in presenc e of a 200-fold excess of HDL3 respective to the concentrations of I-125-la belled VLDL and IDL, For I-125-VLDL high affinity binding to normal HepG2 c ells. we found a dissociation constant (K-d) of 21.2 +/- 3.7 mu g prot./ml (S.E., N = 5) and a maximal binding capacity (B-max) of 0.0312 +/- 0.0063 m u g prot./mg cell prot, while we have measured a K-d of 5.3 +/- 0.8 and a B -max of 0.0081 +/- 0.0014 with LDLr deficient cells. This indicates that LD Lr is responsible for 74% of VLDL binding to HepG2 cells and that the non-L DLr high affinity receptor has a higher affinity for VLDL than LDLr. A 53% loss of I-125-IDL binding capacity was measured with LDLr deficient cells c ompared with normal cells (B-max:0.028 +/- 0.005 versus 0.059 +/- 0.006), w hile no significant statistical difference was found between affinities. Th e study shows that the LDLr is almost the only contributor in VLDL binding, while it shares IDL binding capacity with another high affinity receptor. The physiological importance of LDLr is confirmed by an almost equivalent l oss of IDL and VLDL degradation in LDLr deficient cells. (C) 1999 Elsevier Science Ltd. All rights reserved.