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A CD36 transmembrane domain peptide interrupts CD36 interactions with membrane partners on macrophages and inhibits atherogenic functions

  • Wenxin Huang
    Affiliations
    Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin
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  • Renhao Li
    Affiliations
    Department of Pediatrics, Division of Hematology/Oncology, Emory University School of Medicine, Atlanta, Georgia
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  • Jue Zhang
    Affiliations
    Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin
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  • Yiliang Cheng
    Affiliations
    Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin

    Department of Medicine, Medical Colleges of Wisconsin, Milwaukee, Wisconsin

    Department of Biochemistry, Medical Colleges of Wisconsin, Milwaukee, Wisconsin
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  • Devi P. Ramakrishnan
    Affiliations
    Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin
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  • Roy L. Silverstein
    Correspondence
    Reprint requests: Roy L. Silverstein, Medical College of Wisconsin, 8700 W Watertown Plank Rd, Milwaukee, Wisconsin, 53226.
    Affiliations
    Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin

    Department of Medicine, Medical Colleges of Wisconsin, Milwaukee, Wisconsin
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Published:October 26, 2022DOI:https://doi.org/10.1016/j.trsl.2022.10.005

      Abstract

      CD36 is a transmembrane glycoprotein receptor for oxidized low density lipoprotein (LDL) and other endogenous danger signals and promotes athero-thrombotic processes. CD36 has been shown to associate physically with other transmembrane proteins, including integrins, tetraspanins, and toll-like receptors, which modulate CD36-mediated cell signaling. The CD36 N-terminal transmembrane domain (nTMD) contains a GXXXG sequence motif that mediates protein-protein interactions in many membrane proteins. We thus hypothesized that the nTMD is involved in CD36 interactions with other membrane proteins. CD36 interactions with partner cell surface proteins on murine peritoneal macrophages were detected with an immunofluorescence-based proximity ligation cross linking assay (PLA) and confirmed by immunoprecipitation/immunoblot. Prior to performing these assays, cells were incubated with a synthetic 29 amino acid peptide containing the 22 amino acid of CD36 nTMD or a control peptide in which the glycine residues in GXXXG motif were replaced by valines. In functional experiments, macrophages were preincubated with peptides and then treated with oxLDL to assess LDL uptake, foam cell formation, ROS formation and cell migration. CD36 nTMD peptide treated cells compared to untreated or control peptide treated cells showed decreased CD36 surface associations with tetraspanin CD9 and ameliorated pathologically important CD36 mediated responses to oxLDL, including uptake of DiI-labeled oxLDL, foam cell formation, ROS generation, and inhibition of migration.

      Abbreviations:

      LDL (low density lipoprotein), oxLDL (oxidized low density lipoprotein), nTMD (N-terminal transmembrane domain), ROS (reactive oxygen species), SDS-PAGE (sodium dodecyl sulfate–polyacrylamide gel electrophoresis), DiI (1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate), MAPK (mitogen activated protein kinase), PLA (proximity ligation assay)
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