Cholesterol-Dependent and -Independent CD40 Internalization and Signaling Activation in Cardiovascular Endothelial Cells
- 1 September 2007
- journal article
- Published by Ovid Technologies (Wolters Kluwer Health) in Arteriosclerosis, Thrombosis, and Vascular Biology
- Vol. 27 (9), 2005-2013
- https://doi.org/10.1161/atvbaha.107.145961
Abstract
Objective— It remains elusive how CD40 endocytosis or clustering on the cell surface is induced by different forms of CD40 agonist. This study aims to investigate whether lipid rafts differentially regulate CD40 traffic and signaling in proinflammatory activation of cardiovascular endothelial cells (ECs). Methods and Results— Using fluorescent microscopy and flow cytometry, we demonstrated that soluble CD40L and agonistic antibody G28.5 induced CD40 internalization via clathrin-independent pathway. Furthermore, depletion of cholesterol by methyl-β-cyclodextrin (MCD) or siRNA knockdown of caveolin-1 efficiently blocked CD40 internalization, suggesting that caveolae-rafts pathway regulates CD40 internalization. In contrast, a membrane-bound CD40L mimic (megamer) triggered aggregation of CD40 rafts outside of the conventional cholera toxin B subunit-positive lipid rafts resistant to cholesterol depletion. Finally, both G28.5 and megamer induced CD40 translocation to Brij58-insoluble, low buoyant density rafts, a movement insensitive to cholesterol depletion. However, MCD effectively inhibited G28.5 but not megamer-induced CD40 activation, and such inhibition could be alleviated by cholesterol reconstitution, suggesting that 2 different raft structures of CD40 induced by G28.5 or megamer possess differential sensitivity to cellular cholesterol levels in downstream signaling. Conclusions— Depending on different forms of agonist, CD40 uses either a cholesterol-dependent or -independent mode for trafficking and signaling in ECs.Keywords
This publication has 40 references indexed in Scilit:
- Death-receptor activation halts clathrin-dependent endocytosisProceedings of the National Academy of Sciences of the United States of America, 2006
- Internalization of CD40 regulates its signal transduction in vascular endothelial cellsBiochemical and Biophysical Research Communications, 2006
- Lipid rafts: contentious only from simplistic standpointsNature Reviews Molecular Cell Biology, 2006
- The extracellular domains of FasL and Fas are sufficient for the formation of supramolecular FasL-Fas clusters of high stabilityThe Journal of cell biology, 2005
- Regulation of the Subcellular Localization of Tumor Necrosis Factor Receptor–associated Factor (TRAF)2 by TRAF1 Reveals Mechanisms of TRAF2 SignalingThe Journal of Experimental Medicine, 2002
- Intracellular Retention of Glycosylphosphatidyl Inositol-Linked Proteins in Caveolin-Deficient CellsMolecular and Cellular Biology, 2002
- Floating the Raft Hypothesis: Lipid Rafts Play a Role in Immune Cell ActivationImmunity, 2001
- CD40 AND CD154 IN CELL-MEDIATED IMMUNITYAnnual Review of Immunology, 1998
- The many roles of CD40 in cell-mediated inflammatory responsesImmunology Today, 1996
- A soluble form of TRAP (CD40 ligand) is rapidly released after T cell activationEuropean Journal of Immunology, 1995