Identification and characterization of LRP8 (apoER2) in human blood platelets.
Recently, we reported that apoE inhibits platelet reactivity by stimulating NO release and postulated apoE-receptor activation of intracellular NO synthase (eNOS). Here, we implicate a low density lipoprotein receptor (LDL-R) family member by studying ligand requirements using purified apoE isoforms, synthetic peptides, and the receptor antagonist, receptor-associated protein (RAP). Then, using a homology cloning approach and degenerate PCR primers to amplify the conserved Cys-rich binding domain of the LDL-R family, this receptor was identified as LRP8 (formerly termed, apoER2), a newly described brain protein with several splice variants. Immunoprecipitation of platelet membranes with anti-peptide antisera confirmed protein expression, while analysis of RNA from platelets and two megakaryocytic cell lines (Meg-01 and HEL) disclosed that the major LRP8 transcript lacked binding repeats 4-6 (LRP8delta4-6) but contained the full-length cytoplasmic tail. Sequence analysis of cytoplasmic LRP8 revealed several peptide motifs with potential for cellular signaling and we propose this as a rational mechanism through which apoE inhibits platelet aggregation.
Riddell
D R
DR
Department of Medicine, Royal Free and University College Medical School, University College London, Royal Free Campus, London NW3 2PF, United Kingdom.
Vinogradov
D V
DV
Stannard
A K
AK
Chadwick
N
N
Owen
J S
JS
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001057:Apolipoproteins E / Q000737:chemistry / Q000494:pharmacology; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D017124:Conserved Sequence; D003545:Cysteine; D006801:Humans; D006949:Hyperlipidemias / Q000097:blood / Q000235:genetics; D007700:Kinetics; D026502:LDL-Receptor Related Proteins; D008079:Lipoproteins, VLDL / Q000097:blood; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008533:Megakaryocytes / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D010974:Platelet Aggregation; D012333:RNA, Messenger / Q000235:genetics; D011971:Receptors, Immunologic / Q000378:metabolism; D018110:Receptors, Lipoprotein / Q000097:blood* / Q000737:chemistry / Q000235:genetics*; D014158:Transcription, Genetic
D001057:Apolipoproteins E; D026502:LDL-Receptor Related Proteins; D008079:Lipoproteins, VLDL; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments; D012333:RNA, Messenger; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; C098773:low density lipoprotein receptor-related protein 8; D003545:Cysteine