TLR2 is mobilized into an apical lipid raft receptor complex to signal infection in airway epithelial cells
J. Clin. Invest. Grace Soong, et al. 113:1482 doi:10.1172/JCI20773 [
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Figure 4Lipid rafts are involved in clustering of receptors and signaling. (
A) Confocal z-section images demonstrate caveolin-1 labeled with Alexa Fluor 594 (red) and GM1, identified with cholera toxin β-subunit (CTB) conjugated to Alexa Fluor 488 (green), on the apical surfaces of polarized 16HBE cells permeabilized after stimulation with Pam
3Cys-Ser-Lys
4. (
B) CF nasal polyp cells were infected with
P. aeruginosa PAO1 (PA) and grown on semipermeable supports, and transmission electron micrograph were obtained after 3 hours of bacterial exposure. Arrow (Cav) indicates the flask-shaped electron-dense structures typical of caveolae (magnification, ∞30,000). (
C) Flow cytometry was used to detect superficial caveolin-1 on polarized 16HBE cells after exposure to
S. aureus. Unstim, unstimulated. (
D) Aliquots of Triton-insoluble lysates of 16HBE cells obtained before (–) and after (+) stimulation with
P. aeruginosa PAO1 were fractionated on discontinuous sucrose gradients (4–40%) and were immunoblotted with anti-flotillin, anti-caveolin, anti-TLR-2, anti-IRAK-1, or anti-asialoGM1. Downward arrow indicates raft fraction containing all the components after stimulation. (
E) Aliquots of sucrose gradient fractions from cells treated with filipin prior to stimulation with
P. aeruginosa PAO1 were immunoblotted with anti-flotillin, anti-TLR2, and anti-asialoGM1.
