Fig. 5: Notch signaling is downregulated by CIC-ATXN1L complex deficiency. | Nature Communications

Fig. 5: Notch signaling is downregulated by CIC-ATXN1L complex deficiency.

From: The capicua-ataxin-1-like complex regulates Notch-driven marginal zone B cell development and sepsis progression

Fig. 5

a Volcano plots showing differentially expressed genes (DEGs, log2 fold-change > 0.5) in Cic-deficient MZB cells. Two samples of each genotype were subjected to RNA sequencing. Downregulated Notch signaling-related genes and upregulated CIC target genes are shown next to each corresponding dot. b Gene set enrichment analysis (GSEA) of DEGs in Cic-deficient MZB cells. The gene set database HALLMARK_NOTCH_SIGNALING (MM3870) was employed for GSEA. RT-qPCR analysis showing the downregulation of Notch signaling-related genes in Cic-null (c) and Atxn1l-null (d) MZB cells. For Notch1 and Asb2 levels, N = 5 per group; for Notch2, Dtx1, and Hes1 levels, N = 6 per group; and for Cr2 levels, N = 3 per group (c). N = 3 per group (d). e Surface expression levels of NOTCH1, NOTCH2, and CD21 in MZB and MZP cells from Cd19-Cre and Cicf/f;Cd19-Cre mice. N = 6 per group. f Surface expression levels of NOTCH1, NOTCH2, and CD21 in MZB and MZP cells from Cd19-Cre and Atxn1lf/f;Cd19-Cre mice. N = 6 per group. The MFIs for NOTCH1, NOTCH2, and CD21 were determined using flow cytometry. In vitro MZB cell differentiation assay to assess the differentiation potential of Cic-null (g) and Atxn1l-null (h) T1B cells into MZB cells. Sorted T1B cells were co-cultured with either control or DLL1-expressing OP9 cells for 72 h and subjected to flow cytometry to determine the frequency of MZB (CD21hiCD1dhi) cells. N = 6 per group. i In vitro MZB cell differentiation assay using control and Atxn1l-null T1B cells transfected with either control or Atxn1 siRNAs. T1B cells were co-cultured with DLL1-expressing OP9 cells for 72 h. N = 3 per group. Data represent 2–3 independent experiments. Statistics: Kolmogorov–Smirnov test (b) and two-tailed Student’s t-test (c–i). Bar graphs present the data as mean ± S.D. Ctrl: Cd19-Cre, Cic cKO: Cicf/f;Cd19-Cre, and A1L cKO: Atxn1lf/f;Cd19-Cre. MZB marginal zone B cells, CIC capicua, RT-qPCR quantitative real-time polymerase chain reaction, MZP marginal zone progenitor cells, MFI mean fluorescence intensity, and NES normalized enrichment score. Source data are provided as a Source Data file.

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