Figure S3.

TAB2/3-mediated signal transduction in USP8-depleted cells. (A) Total RNA from HeLa cells transfected with the indicated siRNAs was analyzed by qRT-PCR. CCL5 expression levels were normalized to actin mRNA levels, and expression levels in cells treated with control siRNA were set to 1. Individual values, mean, and SE of the mean of relative mRNA levels are shown. Means ± SE were calculated from four biological replicates. **P < 0.01, ****P < 0.0001 (one-way ANOVA with Dunnett’s test). (B) Parental HeLa cells and HeLa cells stably expressing siRNA-resistant Myc-USP8 WT and C786A were transfected with the indicated siRNAs. Total RNA from cells was analyzed by qRT-PCR. CCL5 expression levels were normalized to actin mRNA levels, and expression levels in cells treated with control siRNA were set to 1. Individual values, mean, and SE of the mean of the relative mRNA levels are shown. Means ± SE were calculated from four biological replicates. **P < 0.01 (one-way ANOVA with Dunnett’s test). (C) Total cell lysates from parental HeLa cells and HeLa cells stably expressing FLAG-TAB2 WT, dNZF, and E685A were immunoblotted with the indicated antibodies. (D and E) HeLa cells stably expressing FLAG-TAB3 WT were transfected with the indicated siRNAs. Cells were immunostained with the indicated antibodies and DAPI. Scale bar, 20 μm. (F) HeLa cells stably expressing FLAG-TAB2 WT, dNZF, and E685A were transfected with the indicated siRNAs. Cells were immunostained with the indicated antibodies and DAPI. Scale bar, 20 μm. (G) Total cell lysates from HeLa cells transfected with the indicated siRNAs were immunoblotted with the indicated antibodies. (H) HeLa cells were stimulated with culture media from HeLa cells transfected with the indicated siRNAs. Total cell lysates were immunoblotted with the indicated antibodies. Source data are available for this figure: SourceData FS3.