Histone-modifying enzymes can regulate DNA damage-induced apoptosis through modulation of p53 function. Here, we show that, in p53-deficient tumor cells, Set9 and LSD1 regulate DNA damage-induced cell death in a manner opposite to that observed in p53(+/+) cells, via modulation of E2F1 stabilization. Set9 methylates E2F1 at lysine-185, which prevents E2F1 accumulation during DNA damage and activation of its proapoptotic target gene p73. This methyl mark is removed by LSD1, which is required for E2F1 stabilization and apoptotic function. The molecular mechanism involves crosstalks between lysine methylation and other covalent modifications that affect E2F1 stability. Methylation at lysine-185 inhibits acetylation and phosphorylation at distant positions and, in parallel, stimulates ubiquitination and degradation of the protein. The findings illustrate that the function of methyltransferases can have opposing biological outcomes depending on the specificity of transcription factor targets.

SummaryTGF-β signaling regulates a variety of cellular processes, including proliferation, apoptosis, differentiation, immune responses, and fibrogenesis. Here, we describe a lysine methylation-mediated mechanism that controls the pro-fibrogenic activity of TGF-β. We find that the methyltransferase Set9 potentiates TGF-β signaling by targeting Smad7, an inhibitory downstream effector. Smad7 methylation promotes interaction with the E3 ligase Arkadia and, thus, ubiquitination-dependent degradation. Depletion or pharmacological inhibition of Set9 results in elevated Smad7 protein levels and inhibits TGF-β-dependent expression of genes encoding extracellular matrix components. The inhibitory effect of Set9 on TGF-β-mediated extracellular matrix production is further demonstrated in mouse models of pulmonary fibrosis. Lung fibrosis induced by bleomycin or Ad-TGF-β treatment was highly compromised in Set9-deficient mice. These results uncover a complex regulatory interplay among multiple Smad7 modifications and highlight the possibility that protein methyltransferases may represent promising therapeutic targets for treating lung fibrosis.

Summary The oncogenic function of suppressor of variegation, enhancer of zeste and MYeloid-Nervy-DEAF1-domain family methyltransferase Smyd3 has been implicated in various malignancies, including hepatocellular carcinoma (HCC). Here, we show that targeting Smyd3 by next-generation antisense oligonucleotides (Smyd3-ASO) is an efficient approach to modulate its mRNA levels in vivo and to halt the growth of already initiated liver tumors. Smyd3-ASO treatment dramatically decreased tumor burden in a mouse model of chemically induced HCC and negatively affected the growth rates, migration, oncosphere formation, and xenograft growth capacity of a panel of human hepatic cancer cell lines. Smyd3-ASOs prevented the activation of oncofetal genes and the development of cancer-specific gene expression program. The results point to a mechanism by which Smyd3-ASO treatment blocks cellular de-differentiation, a hallmark feature of HCC development, and, as a result, it inhibits the expansion of hepatic cancer stem cells, a population that has been presumed to resist chemotherapy.

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