FIGURE 5.

Tollip contributes to IL-1Rrp2 localization to lysosomes after agonist addition. A, the effects of Tollip in the colocalization to Rab11 endosomes. B, the effects of Tollip on the colocalization of IL-1Rrp2 to LAMP1 lysosomes. The results in A and B had NCI cells treated with siRNA to Tollip (T) or to a nonspecific control (C). Tollip and LAMP1 colocalization was determined by immunoconfocal microscopy. *, p < 0.05. All data shown were quantified from over 20 cells from three independently prepared samples. C, Western blot showing the accumulation of IL-1Rrp2, IL-1RAcp, and IL-1R1 in NCI or hDF cells 3 h after IL-36γ addition. All images are from Western blots performed with the same set of cell lysates. The abundance of β-actin was used as an internal control. These results for Tollip knockdown decreasing IL-1Rrp2 and IL-1RAcp accumulation were reproduced in more than three independent experiments. The decrease in the Tollip knockdown was ∼50% in these experiments. D, Western blot showing that IL-1β-treated NCI and hDF cells also exhibit an increase in IL-1R1 accumulation. Cell lysates used for the analysis were from 3 h after the addition of IL-1β. E, model for the role of Tollip in IL-36R trafficking in the absence or presence of IL-36R agonists.