Figure S3.
DCAF13 depletion exerted slighter effects on rRNA transcription rather than protein synthesis. (A) Nucleolar morphology of naïve or stimulated CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells detected by electron microscopy. Representative images (upper) and the quantities of areas (right) are shown. The white dotted line outlined the shape of the nucleus and the white tip marked obvious DFC regions (n = 13). (B) FSA of naïve CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells. (C) Confocal z-stack three-dimensional reconstruction of naïve CD4 T cells was isolated from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) to compare their ratios of cytoplasm and nucleus. Representative images (left) and corresponding quantitation from three independent experiments (n = 36) are shown. DAPI (blue) and CD4 (green). Scale bar, 2 μm. (D) Flow cytometry analysis of HPG signals in ERT2 Cre+Dcaf13fl/fl CD4+ and CD8+ T cells treated with tamoxifen (regarded as DCAF13 KO) or without tamoxifen (regarded as WT). T cells were stimulated with anti-CD3/CD28 for 24 h. (E) Polysome profiles analysis of the equal amounts of naïve T cells from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO). Representative pictures of A254 absorbance profile in three independent experiments(left) and quantification of the 40S peaks (right, n = 3) are shown. (F) IF of EU signals in CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells stimulated by anti-CD3/CD28 for 24 h. Actinomycin D (ActD) was added as an EU inhibitor. The gray value of the EU signal was measured by ImageJ (n = 5). (G) Ribosomal RNA processing products in naïve T cells from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) mice were detected by northern blotting with OTSI and OTSII oligos. Loaded RNA was purified from equal amounts of cells. The gray values of bands were quantified with ImageJ. WT cells were normalized to 1.0. The fold change of 18S-E was summarized from three independent experiments (n = 3). Data are presented as the mean ± SD. A two-tailed Student’s t test was used. ns is not significant. *P < 0.05, **P < 0.01, ***P < 0.001. Source data are available for this figure: SourceData FS3.

DCAF13 depletion exerted slighter effects on rRNA transcription rather than protein synthesis. (A) Nucleolar morphology of naïve or stimulated CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells detected by electron microscopy. Representative images (upper) and the quantities of areas (right) are shown. The white dotted line outlined the shape of the nucleus and the white tip marked obvious DFC regions (n = 13). (B) FSA of naïve CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells. (C) Confocal z-stack three-dimensional reconstruction of naïve CD4 T cells was isolated from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) to compare their ratios of cytoplasm and nucleus. Representative images (left) and corresponding quantitation from three independent experiments (n = 36) are shown. DAPI (blue) and CD4 (green). Scale bar, 2 μm. (D) Flow cytometry analysis of HPG signals in ERT2 Cre+Dcaf13fl/fl CD4+ and CD8+ T cells treated with tamoxifen (regarded as DCAF13 KO) or without tamoxifen (regarded as WT). T cells were stimulated with anti-CD3/CD28 for 24 h. (E) Polysome profiles analysis of the equal amounts of naïve T cells from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO). Representative pictures of A254 absorbance profile in three independent experiments(left) and quantification of the 40S peaks (right, n = 3) are shown. (F) IF of EU signals in CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) T cells stimulated by anti-CD3/CD28 for 24 h. Actinomycin D (ActD) was added as an EU inhibitor. The gray value of the EU signal was measured by ImageJ (n = 5). (G) Ribosomal RNA processing products in naïve T cells from CD4 Cre−Dcaf13fl/fl (WT) and CD4 Cre+Dcaf13fl/fl (cKO) mice were detected by northern blotting with OTSI and OTSII oligos. Loaded RNA was purified from equal amounts of cells. The gray values of bands were quantified with ImageJ. WT cells were normalized to 1.0. The fold change of 18S-E was summarized from three independent experiments (n = 3). Data are presented as the mean ± SD. A two-tailed Student’s t test was used. ns is not significant. *P < 0.05, **P < 0.01, ***P < 0.001. Source data are available for this figure: SourceData FS3.

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