Inhibitory effect on the lipolysis-stimulated receptor of the 39-kDa receptor-associated protein.
Adenovirus vector-mediated transfer of the receptor-associated protein (RAP) gene into low density lipoprotein (LDL) receptor-deficient mice was shown to achieve plasma concentrations ranging between 20 and 200 micrograms/ml and to result in the accumulation of remnant lipoproteins (Willnow, T. E., Sheng, Z., Ishibashi, S., and Herz, J. (1994) Science 264, 1471-1474). Both this finding and the observation that in addition to various other members of the LDL receptor gene family, RAP binds to a yet unidentified protein of apparent molecular mass of 105 kDa prompted us to examine the effect of high concentrations of RAP on the lipolysis-stimulated receptor (LSR). LSR is a receptor distinct from the LDL receptor and the LDL receptor-related protein and is capable of binding apoB and apoE when activated by free fatty acids. Data reported here show that in fibroblasts isolated from a subject homozygous for familial hypercholesterolemia, RAP fusion protein inhibited LSR-mediated binding of 125I-LDL and the subsequent internalization and degradation of the particles. Studies on the interaction of RAP with LSR in isolated rat liver membranes revealed that at concentrations > or = 10 micrograms/ml, RAP inhibited in a dose-dependent manner the binding of LDL to LSR; half-maximum inhibition was obtained with 20 micrograms/ml RAP. Ligand blotting studies revealed that RAP bound directly to two rat liver membrane proteins of apparent molecular masses identical to those that bind 125I-LDL after preincubation with oleate. However, unlike LDL, binding of 125I-RAP to LSR did not require preincubation with oleate. Preincubation of nitrocellulose membranes with an excess of unlabeled RAP fusion protein decreased oleate-induced binding of 125I-LDL to LSR candidate proteins, whereas preincubation with excess unlabeled LDL was unable to prevent the subsequent binding of 125I-RAP to the LSR proteins. Both the latter data and analysis of the mechanism of inhibition were consistent with the RAP inhibitory effect on LSR being achieved by interference with a site distinct from the oleate-induced LDL binding site. In conclusion, this study shows that at concentrations reported to delay chylomicron remnant removal in LDL receptor-deficient mice, RAP exerted a significant inhibitory effect on LSR.
Troussard
A A
AA
INSERM U391, Faculté des Sciences Pharmaceutiques et Biologiques, Université de Rennes I, France.
Khallou
J
J
Mann
C J
CJ
André
P
P
Strickland
D K
DK
Bihain
B E
BE
Yen
F T
FT
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism / Q000494:pharmacology*; D002478:Cells, Cultured; D006023:Glycoproteins / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D026563:LDL-Receptor Related Protein-Associated Protein; D008077:Lipoproteins, LDL / Q000378:metabolism; D008970:Molecular Weight; D051381:Rats; D011973:Receptors, LDL / Q000037:antagonists & inhibitors*; D011994:Recombinant Proteins / Q000494:pharmacology
D002352:Carrier Proteins; D006023:Glycoproteins; D026563:LDL-Receptor Related Protein-Associated Protein; D008077:Lipoproteins, LDL; D011973:Receptors, LDL; D011994:Recombinant Proteins; C036053:VLDL receptor; C085992:lipolysis-stimulated receptor