Figure 6
Figure 6. Jdp2, a Myc-collaborating gene is a direct target of Hdac1/2 and is required for the survival of Hdac1Δ/Δ; Hdac2+/Δ lymphomas. (A) Western blot analysis of histone H3 and H4 acetylation in nuclear lysates of preleukemic thymocytes from 3 independent 1-week-old wild-type, LckCre+;Hdac1Δ/Δ and LckCre+;Hdac1Δ/Δ;Hdac2+/Δ mice (left). Acetylated H3 and H4 signals were quantified over total H3 and H4 signal, respectively. While AcH3/H4 levels were significantly higher in LckCre+;Hdac1Δ/Δ and LckCre+;Hdac1Δ/Δ;Hdac2+/Δ thymocytes compared with wild-type thymocytes, only AcH3 levels were increased in LckCre+;Hdac1Δ/Δ;Hdac2+/Δ thymocytes compared with LckCre+;Hdac1Δ/Δ counterparts. (B) Venn diagram demonstrating the overlap between sets of differentially expressed genes (adjusted P value < 0.05, fold change ≥1.5×, n ≥3 independent mice per group) in 1-week-old and 3-week-old Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and lymphomas. (C) Schematic representation of the overlap between CTGs identified in insertional mutagenesis screens and genes deregulated in Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and lymphomas (“HDAC”; Fisher exact test, P = 6.36e-10). (D) Fold changes in mRNA of Myc-collaborating genes Jdp2, Jazf1, Dyrk3, Lpar2, 2010107G12RIK, and Ephb2 in 1-week-old and 3-week-old Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and Hdac1Δ/Δ;Hdac2Δ/+ lymphomas, relative to age-matched wild-type thymi. (E) Western blot analysis of protein lysates of lymphomas of indicated genotypes for Hdac1, Jdp2 (lower band), and p53. Tubulin served as a loading control. (F) Western blot analysis (left) and chromatin immunoprecipitation analysis (right) of GFP, Hdac1-GFP, or Hdac2 GFP expressing p53−/− T-cell lymphoma cell lines for the Jdp2 intron 1-2 and 5′ UTR. (G) Cell viability assessed by cell titer blue assay (left) and representative images (right; magnification 100×) of Hdac1Δ/Δ;Hdac2Δ/+ and p53−/− lymphoma cell lines infected with independent lentiviral Jdp2 shRNA constructs (Jdp2_KD1 or Jdp2_KD2), a lentiviral nontargeting shRNA (NT) construct, or mock infection. WT, wild type.

Jdp2, a Myc-collaborating gene is a direct target of Hdac1/2 and is required for the survival of Hdac1Δ/Δ; Hdac2+/Δ lymphomas. (A) Western blot analysis of histone H3 and H4 acetylation in nuclear lysates of preleukemic thymocytes from 3 independent 1-week-old wild-type, LckCre+;Hdac1Δ/Δ and LckCre+;Hdac1Δ/Δ;Hdac2+/Δ mice (left). Acetylated H3 and H4 signals were quantified over total H3 and H4 signal, respectively. While AcH3/H4 levels were significantly higher in LckCre+;Hdac1Δ/Δ and LckCre+;Hdac1Δ/Δ;Hdac2+/Δ thymocytes compared with wild-type thymocytes, only AcH3 levels were increased in LckCre+;Hdac1Δ/Δ;Hdac2+/Δ thymocytes compared with LckCre+;Hdac1Δ/Δ counterparts. (B) Venn diagram demonstrating the overlap between sets of differentially expressed genes (adjusted P value < 0.05, fold change ≥1.5×, n ≥3 independent mice per group) in 1-week-old and 3-week-old Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and lymphomas. (C) Schematic representation of the overlap between CTGs identified in insertional mutagenesis screens and genes deregulated in Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and lymphomas (“HDAC”; Fisher exact test, P = 6.36e-10). (D) Fold changes in mRNA of Myc-collaborating genes Jdp2, Jazf1, Dyrk3, Lpar2, 2010107G12RIK, and Ephb2 in 1-week-old and 3-week-old Hdac1Δ/Δ;Hdac2Δ/+ thymocytes and Hdac1Δ/Δ;Hdac2Δ/+ lymphomas, relative to age-matched wild-type thymi. (E) Western blot analysis of protein lysates of lymphomas of indicated genotypes for Hdac1, Jdp2 (lower band), and p53. Tubulin served as a loading control. (F) Western blot analysis (left) and chromatin immunoprecipitation analysis (right) of GFP, Hdac1-GFP, or Hdac2 GFP expressing p53−/− T-cell lymphoma cell lines for the Jdp2 intron 1-2 and 5′ UTR. (G) Cell viability assessed by cell titer blue assay (left) and representative images (right; magnification 100×) of Hdac1Δ/Δ;Hdac2Δ/+ and p53−/− lymphoma cell lines infected with independent lentiviral Jdp2 shRNA constructs (Jdp2_KD1 or Jdp2_KD2), a lentiviral nontargeting shRNA (NT) construct, or mock infection. WT, wild type.

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