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. 2013 Apr 22;288(22):16043–16054. doi: 10.1074/jbc.M113.473223

FIGURE 4.

FIGURE 4.

Dectin-1-dependent Syk activation is pH-independent. Panel A, RAW GFP-Dectin-1 or GFP-Dectin-1ΔY15F cells were incubated with β-1,3-glucan particles for the times indicated. Cells were lysed using Nonidet P-40 in the presence of protease and phosphatase inhibitors. Total cell lysates were resolved on SDS-PAGE and membranes blotted for phosphorylated Syk (P-Syk, arrowhead). Panel B, RAW GFP-Dectin-1 cells were pre-incubated with PP2, BafA1 or vehicle control for 30 min. Stimulation was performed using heat-killed C. albicans (yeast to macrophage ratio of 1:1), β-1,3-glucan beads (bead to cell ratio of 5:1) or resting conditions for 20 min. Following lysis, total cell lysate was resolved by SDS-PAGE and blotted for phosphorylated Syk (P-Syk, arrowhead). Representative β-actin blots are shown as loading controls.