Related toFigs. 3, 4, and 5: Chromatin accessibility, gene expression, VDJHrearrangement, and apoptosis in WT and Mki67−/−B cell progenitors. (A) Accessibility of the Igh locus (chr12:113,254,000-116,022,000) in the pro-B and small pre-B cells from WT and Mki67−/− mice. Data are representative of four mice from each group. (B) Accessibility at the Igκ locus (chr6:67,554,000-70,728,000) in the pro-B and small pre-B cells from WT and Mki67−/− mice. Data are representative of four mice from each group. (C) Accessibility at the Rag locus (chr2:101,550,000–111,655,000) in LT-HSC and DN3 thymocytes from WT mice (obtained from the ImmGen database) and in pre-pro-B cells from WT and Mki67−/− mice (representative of four mice from each group). The putative enhancer region annotated by ImmGen database for the Rag locus is highlighted. (D) mRNA expression of Pax5 in pre-pro-B, pro-B, and small pre-B cells from WT (n = 6) and Mki67−/− (n = 6) mice by RT-qPCR. Normalization was performed as described in Fig. 4. Data are merged from two independent experiments. Statistical differences were determined by multiple Mann–Whitney tests (corrected for multiple comparisons using the Holm–Sidak method). ns, P > 0.05. (E) Representative histograms showing intracellular μ chain expression in WT and Mki67−/− pre-pro-B, pro-B, and large pre-B cells. (F) Comparison of the median fluorescent intensity (MFI) of intracellular μ chain expression in WT (n = 3) and Mki67−/− (n = 3) pre-pro-B, pro-B, and large pre-B cells. Statistical differences were determined by one-way ANOVA using Dunnett’s multiple comparisons tests. (G) The relative abundance of recombined VHJ558 family members and each of the JH1–4 segments in WT (n = 9) or Mki67−/− (n = 6) pre-B cells measured by qPCR. ΔΔCT was calculated as described in Fig. 4 E. Statistical differences were determined by two-tailed unpaired t test with P values shown. (H) Gating strategy of apoptotic cells among small pre-B cells from WT- or Mki67−/−Rag1−/−SWHEL-HCKi/Ki mice. (I) Quantification of percentages of apoptotic cells among the small pre-B cells from WT- (n = 3) or Mki67−/−Rag1−/−SWHEL-HCKi/Ki (n = 3) mice. Statistical differences were determined by two-tailed unpaired t test with P values shown.
Figure S3.

Related to Figs. 3, 4, and 5: Chromatin accessibility, gene expression, VDJHrearrangement, and apoptosis in WT and Mki67−/−B cell progenitors. (A) Accessibility of the Igh locus (chr12:113,254,000-116,022,000) in the pro-B and small pre-B cells from WT and Mki67−/− mice. Data are representative of four mice from each group. (B) Accessibility at the Igκ locus (chr6:67,554,000-70,728,000) in the pro-B and small pre-B cells from WT and Mki67−/− mice. Data are representative of four mice from each group. (C) Accessibility at the Rag locus (chr2:101,550,000–111,655,000) in LT-HSC and DN3 thymocytes from WT mice (obtained from the ImmGen database) and in pre-pro-B cells from WT and Mki67−/− mice (representative of four mice from each group). The putative enhancer region annotated by ImmGen database for the Rag locus is highlighted. (D) mRNA expression of Pax5 in pre-pro-B, pro-B, and small pre-B cells from WT (n = 6) and Mki67−/− (n = 6) mice by RT-qPCR. Normalization was performed as described in Fig. 4. Data are merged from two independent experiments. Statistical differences were determined by multiple Mann–Whitney tests (corrected for multiple comparisons using the Holm–Sidak method). ns, P > 0.05. (E) Representative histograms showing intracellular μ chain expression in WT and Mki67−/− pre-pro-B, pro-B, and large pre-B cells. (F) Comparison of the median fluorescent intensity (MFI) of intracellular μ chain expression in WT (n = 3) and Mki67−/− (n = 3) pre-pro-B, pro-B, and large pre-B cells. Statistical differences were determined by one-way ANOVA using Dunnett’s multiple comparisons tests. (G) The relative abundance of recombined VHJ558 family members and each of the JH1–4 segments in WT (n = 9) or Mki67−/− (n = 6) pre-B cells measured by qPCR. ΔΔCT was calculated as described in Fig. 4 E. Statistical differences were determined by two-tailed unpaired t test with P values shown. (H) Gating strategy of apoptotic cells among small pre-B cells from WT- or Mki67−/−Rag1−/−SWHEL-HCKi/Ki mice. (I) Quantification of percentages of apoptotic cells among the small pre-B cells from WT- (n = 3) or Mki67−/−Rag1−/−SWHEL-HCKi/Ki (n = 3) mice. Statistical differences were determined by two-tailed unpaired t test with P values shown.

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