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Published on: May 31, 2018
A CD36 transmembrane domain peptide interrupts CD36 interactions with membrane partners on macrophages and inhibits
Wenxin Huang1, Renhao Li2, Jue Zhang1
1Laboratory of Vascular Pathobiology, Versiti Blood Center of Wisconsin, Blood Research Institute, Milwaukee, Wisconsin.
Insights
The CD36 N-terminal transmembrane domain (nTMD) peptide disrupts CD36 interactions with CD9, reducing oxidized low-density lipoprotein (oxLDL) uptake and foam cell formation in macrophages.
Area of Science:
- Cardiovascular Biology
- Molecular Cell Biology
- Immunology
Background:
- CD36 is a transmembrane glycoprotein receptor involved in athero-thrombotic processes by binding oxidized low-density lipoprotein (oxLDL) and danger signals.
- CD36 interacts with other transmembrane proteins, modulating its signaling pathways.
- The CD36 N-terminal transmembrane domain (nTMD) possesses a GXXXG motif, suggesting a role in protein-protein interactions.
Purpose of the Study:
- To investigate the role of the CD36 nTMD in protein-protein interactions and its impact on CD36-mediated cellular responses.
- To determine if disrupting the GXXXG motif in the CD36 nTMD affects its association with other membrane proteins and downstream signaling.
Main Methods:
- Utilized proximity ligation crosslinking assay (PLA) and immunoprecipitation/immunoblot to detect CD36 interactions with partner proteins on murine peritoneal macrophages.
- Employed synthetic peptides corresponding to the CD36 nTMD, including a control peptide with mutated glycine residues in the GXXXG motif.
- Assessed functional responses, including oxLDL uptake, foam cell formation, reactive oxygen species (ROS) generation, and cell migration, after peptide treatment and oxLDL stimulation.
Main Results:
- CD36 nTMD peptide treatment significantly decreased CD36 surface associations with tetraspanin CD9.
- Macrophages pretreated with the CD36 nTMD peptide exhibited ameliorated oxLDL uptake and reduced foam cell formation.
- The CD36 nTMD peptide also decreased ROS generation and partially inhibited oxLDL-induced migration.
Conclusions:
- The CD36 nTMD plays a crucial role in mediating interactions with other cell surface proteins like CD9.
- Disrupting the CD36 nTMD, specifically the GXXXG motif, can attenuate key athero-thrombotic CD36-mediated responses to oxLDL.
- Targeting the CD36 nTMD offers a potential therapeutic strategy for cardiovascular diseases.
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.
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