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. 2014 Nov 6;10(11):e1004477. doi: 10.1371/journal.ppat.1004477

Figure 9. Lipid rafts serve as platforms for the αvβ3-integrin-dependent re-localization of TLR2, MAL and MYD88.

Figure 9

293T or 293T sh-β3 cells were transfected with plasmids encoding TLR2-Flag, MYD88-HA and MAL-Flag plasmids. When indicated, CD14 plasmid was included (E–H). 48 h after transfection, cells were exposed for 30 min to R7910 (60 PFU/cell) (C, D, G, H) or mock-infected (A, B, E, F). Cells were suspended in 1 ml of TNE buffer (10 mM Tris-HCl, pH 7.5, 150 mM NaCl, 5 mM EDTA) containing 1% Triton X-100 (Sigma Aldrich, Milan, Italy) and 0.3 mM protease inhibitors, and incubated on ice for 1 h. Membrane fractions were prepared as described [26]. The samples obtained from fractionation of the sucrose gradients (# 1–13) were subjected to SDS-PAGE and WB with MAb to HA for detection of MYD88, and MAb to Flag for detection of TLR2 and MAL.