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. Author manuscript; available in PMC: 2021 Jul 13.
Published in final edited form as: Nat Aging. 2021 May 13;1(5):454–472. doi: 10.1038/s43587-021-00063-1

Extended Data Fig. 4 |. Enhanced SASP expression and decreased H3K9/H3K36 methylation are regulated by KDM4B.

Extended Data Fig. 4 |

a, Quantitative measurement of the SASP expression at transcription level. Stromal cells were transduced with a lentiviral construct encoding human KDM4B and/or exposed to BLEO treatment before analyzed. Signals normalized to CTRL cells (transduced with empty vector and untreated). b, Immunoblot assay of DNA damage repair (DDR) signaling, H3K9/H3K36 methylation, CXCL8 and senescence marker expression in stromal cells treated differently as described in (a). GAPDH, loading control. Chromatin fractionation was performed to evaluate KDM4A/B levels in nuclei, histone H3 as a nuclear lysate loading control. c, Immunoblot assay of DDR signaling, H3K9/H3K36 methylation, CXCL8 and senescence marker expression in cells treated with BLEO and/or Chaetocin. GPADH, loading control. SE, short exposure. LE, long exposure. d, Representative images of SA-β-Gal and BrdU staining of PSC27 treated in the ways described in (a). Scale bar, 20 μm. e, Comparative statistics of SA-β-Gal and BrdU staining results of stromal cells in the individual conditions of (a). Upper, SA-β-Gal staining. Lower, BrdU staining. f, Immunoblot analysis of key targets expressed in PSC27 sublines depleted of KDM4A or KDM4B via lentiviral transduction and/or subject to genotoxic stress by BLEO. g, Transcript expression of hallmark SASP factors in PSC27 sublines transduced with lentiviral constructs encoding shRNAs specific for KDM4B. Scrambled, transduction control. Cells subject to vehicle or BLEO treatment before analyzed. h, Comparative statistics of SA-β-Gal staining results of stromal cells transduced with a lentiviral construct encoding human KDM4A or 4B and/or exposed to BLEO treatment before processed. i, Comparative statistics of BrdU staining results of stromal cells transduced with a lentiviral construct encoding human KDM4A or 4B and/or exposed to BLEO treatment before processed. Data in all bar plots are shown as mean ± SD and representative of 3 biological replicates. a, e, g, P values were determined by two-sided unpaired t-test, and adjusted for multiple comparisons. Data in b-c, d, f are representative of 3 biological replicates. h-i, P values were determined by one-way ANOVA, and adjusted for multiple comparisons.