Dominant negative mutations of the scavenger receptor. Native receptor inactivation by expression of truncated variants.
The bovine scavenger receptor was truncated at amino acid 266 or 310 to delete either all or part, respectively, of the collagen-like domain. The truncated receptors were inactive in the binding and internalization of acetyl (Ac) low density lipoprotein (LDL). Coexpression of truncated receptor with the native receptor dramatically reduced the percentage of cells internalizing fluorescently labeled Ac LDL, compared with cells expressing the native receptor alone. The mutant truncated at amino acid 266 was most effective in receptor inactivation, resulting in a 42% or 80% decrease in the percentage of cells expressing active receptor when transfected in a 1:1 or 1:2 molar ratio (native:mutant), respectively, with native receptor. Degradation of 125I-Ac LDL was reduced up to 90% when the native and truncated mutant receptors were coexpressed. Scavenger receptor inhibition was specific because the activity of the LDL receptor was not altered. Transient transfection of the mouse macrophage cell line P388D1 with truncated scavenger receptor resulted in a 65% decrease in the uptake and degradation of Ac LDL but did not decrease the degradation of beta-migrating very low density lipoprotein, which is LDL receptor-mediated. These results demonstrate that expression of truncated bovine scavenger receptor inactivates both the native bovine and murine scavenger receptors, producing a dominant negative phenotype in vitro.
Dejager
S
S
Gladstone Institute of Cardiovascular Disease, University of California, San Francisco 94141-9100.
Mietus-Snyder
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M
Friera
A
A
Pitas
R E
RE
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002791:Cholesterol, Dietary; D005434:Flow Cytometry; D014644:Genetic Variation; D006801:Humans; D006937:Hypercholesterolemia / Q000097:blood; D008074:Lipoproteins / Q000097:blood / Q000302:isolation & purification / Q000378:metabolism; D008264:Macrophages; D008565:Membrane Proteins; D051379:Mice; D016296:Mutagenesis; D011817:Rabbits; D011971:Receptors, Immunologic / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D051122:Scavenger Receptors, Class B; D017384:Sequence Deletion; D014162:Transfection
D002791:Cholesterol, Dietary; D008074:Lipoproteins; D008565:Membrane Proteins; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; D011994:Recombinant Proteins; C454739:Scarb1 protein, mouse; D051122:Scavenger Receptors, Class B