Extended Data Fig. 10: Lentiviral reconstitution of wild-type and mutant SR-B1 expression in human endothelial cells. | Nature

Extended Data Fig. 10: Lentiviral reconstitution of wild-type and mutant SR-B1 expression in human endothelial cells.

From: SR-B1 drives endothelial cell LDL transcytosis via DOCK4 to promote atherosclerosis

Extended Data Fig. 10

ac, Studies of reconstituted wild-type SR-B1, extracellular point mutants of SR-B1, or SR-B1(Q445A). The extracellular point mutants were: SR-B1(M159E), SR-B1(T165E), SR-B1(F171A), SR-B1(T175A), and SR-B1(E178A). Whole cell lysate abundance (a), cell surface abundance (b; except for Q445A, which was previously evaluated), and nLDL and oxLDL binding (c) were evaluated. df, Studies of reconstituted wild-type SR-B1 or C-terminal cytoplasmic tail deletion mutants of SR-B1. The mutants were: SR-B1(∆C15) (∆495–509), SR-B1(∆C23) (∆487–509) and SR-B1(∆C30) (∆480–509). Whole cell lysate abundance (d), cell surface abundance (e), and nLDL and oxLDL binding (f) were evaluated. g, Top, sequence alignment of amino acids in the C-terminal cytoplasmic tail of SR-B1 homologues (residues 487–494) from human (Homo sapiens, Hs, Q8WTVO, Swiss-Prot), mouse (Mus musculus, Mm, Q61009, Swiss-Prot), rat (Rattus norvegicus, Rr, P97943, Swiss-Prot), bovine (Bos taurus, Bt, O18824, Swiss-Prot), pig (Sus scrofa, Ss, Q8SQC1, Swiss-Prot), and Chinese hamster (Cricetulus griseus, Cg, Q60417, Swiss-Prot). Fully conserved residues are shown in bold. Bottom, comparison of human SR-B1 residues 487–494 and entire human CD36 C-terminal cytoplasmic tail. Residues of SR-B1 not shared with CD36 are shown in bold. hj, Studies of reconstituted wild-type or C-terminal cytoplasmic tail substitution mutants of SR-B1. The mutants were: SR-B1(IQAY), SR-B1(SESL), SR-B1(Y490A), SR-B1(Q488A), SR-B1(S491A), SR-B1(E492A), SR-B1(S493A), and SR-B1(L494A). Whole cell lysate abundance (h), cell surface abundance (i), and nLDL and oxLDL binding (j) were evaluated. km, Binding (k), uptake (l) and transcytosis (m) of nLDL were evaluated with the various mutants shown at an nLDL concentration of 100 μg ml–1. n, Whole cell lysate abundance of CD36 and LDLR following reconstitution with the SR-B1 mutants tested in km. Data are mean ± s.e.m. For cell surface abundance by flow cytometry, n = 3 or 4. For LDL binding, n = 4 or 8. For LDL transcytosis, n = 3. In c, km, P values for comparison with wild-type calculated by two-sided Student’s t-test.

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