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Supplementary Table from SMYD3 Impedes Small Cell Lung Cancer Sensitivity to Alkylation Damage through RNF113A Methylation–Phosphorylation Cross-talk

Small cell lung cancer (SCLC) is the most fatal form of lung cancer, with dismal survival, limited therapeutic options, and rapid development of chemoresistance. We identified the lysine methyltransferase SMYD3 as a major regulator of SCLC sensitivity to alkylation-based chemotherapy. RNF113A methylation by SMYD3 impairs its interaction with the phosphatase PP4, controlling its phosphorylation levels. This cross-talk between posttranslational modifications acts as a key switch in promoting and maintaining RNF113A E3 ligase activity, essential for its role in alkylation damage response. In turn, SMYD3 inhibition restores SCLC vulnerability to alkylating chemotherapy. Our study sheds light on a novel role of SMYD3 in cancer, uncovering this enzyme as a mediator of alkylation damage sensitivity and providing a rationale for small-molecule SMYD3 inhibition to improve responses to established chemotherapy.Significance:SCLC rapidly becomes resistant to conventional chemotherapy, leaving patients with no alternative treatment options. Our data demonstrate that SMYD3 upregulation and RNF113A methylation in SCLC are key mechanisms that control the alkylation damage response. Notably, SMYD3 inhibition sensitizes cells to alkylating agents and promotes sustained SCLC response to chemotherapy.This article is highlighted in the In This Issue feature, p. 2007

Supplementary Data from SMYD3 Impedes Small Cell Lung Cancer Sensitivity to Alkylation Damage through RNF113A Methylation–Phosphorylation Cross-talk

Small cell lung cancer (SCLC) is the most fatal form of lung cancer, with dismal survival, limited therapeutic options, and rapid development of chemoresistance. We identified the lysine methyltransferase SMYD3 as a major regulator of SCLC sensitivity to alkylation-based chemotherapy. RNF113A methylation by SMYD3 impairs its interaction with the phosphatase PP4, controlling its phosphorylation levels. This cross-talk between posttranslational modifications acts as a key switch in promoting and maintaining RNF113A E3 ligase activity, essential for its role in alkylation damage response. In turn, SMYD3 inhibition restores SCLC vulnerability to alkylating chemotherapy. Our study sheds light on a novel role of SMYD3 in cancer, uncovering this enzyme as a mediator of alkylation damage sensitivity and providing a rationale for small-molecule SMYD3 inhibition to improve responses to established chemotherapy. Significance: SCLC rapidly becomes resistant to conventional chemotherapy, leaving patients with no alternative treatment options. Our data demonstrate that SMYD3 upregulation and RNF113A methylation in SCLC are key mechanisms that control the alkylation damage response. Notably, SMYD3 inhibition sensitizes cells to alkylating agents and promotes sustained SCLC response to chemotherapy. This article is highlighted in the In This Issue feature, p. 2007

International audience; Summary Central to genotoxic responses is their ability to sense highly specific signals to activate the appropriate repair response. We previously reported that the activation of the ASCC-ALKBH3 repair pathway is exquisitely specific to alkylation damage in human cells. Yet the mechanistic basis for the selectivity of this pathway was not immediately obvious. Here, we demonstrate that RNA but not DNA alkylation is the initiating signal for this process. Aberrantly methylated RNA is sufficient to recruit ASCC, while an RNA dealkylase suppresses ASCC recruitment during chemical alkylation. In turn, recruitment of ASCC during alkylation damage, which is mediated by the E3 ubiquitin ligase RNF113A, suppresses transcription and R-loop formation. We further show that alkylated pre-mRNA is sufficient to activate RNF113A E3 ligase in vitro in a manner dependent on its RNA binding Zn-finger domain. Together, our work identifies an unexpected role for RNA damage in eliciting a specific response to genotoxins.

Protein lysine methylation is a functionally diverse post-translational modification involved in various major cellular processes. Lysine methylation can modulate proteins activity, stability, localization, and/or interaction, resulting in specific downstream signaling and biological outcomes. Lysine methylation is a dynamic and fine-tuned process, deregulation of which often leads to human pathologies. In particular, the lysine methylome and its associated signaling network can be linked to carcinogenesis and cancer progression. Histone modifications and chromatin regulation is a major aspect of lysine methylation importance, but increasing evidence suggests that a high relevance and impact of non-histone lysine methylation signaling has emerged in recent years. In this review, we draw an updated picture of the current scientific knowledge regarding non-histone lysine methylation signaling and its implication in physiological and pathological processes. We aim to demonstrate the significance of lysine methylation as a major and yet underestimated posttranslational modification, and to raise the importance of this modification in both epigenetic and cellular signaling by focusing on the observed activities of SET- and 7β-strandcontaining human lysine methyltransferases. Recent evidence suggests that what has been observed so far regarding lysine methylation’s implication in human pathologies is only the tip of the iceberg. Therefore, the exploration of the “methylome network” raises the possibility to use these enzymes and their substrates as promising new therapeutic targets for the development of future epigenetic and methyllysine signaling cancer treatments.

SummaryA critical question in genome stability is the nature of the chemical damage responsible for repair activation. We previously reported a novel pathway specifically activated during alkylation damage in human cells, where the E3 ubiquitin ligase RNF113A mediates the recruitment of the ASCC repair complex. Yet the mechanistic basis for the alkylation damage selectivity of this pathway remains unclear. Here, we demonstrate that RNA but not DNA alkylation is the initiating signal for this process. Aberrantly methylated RNA is sufficient to recruit ASCC, while an RNA dealkylase suppresses ASCC recruitment during chemical alkylation. This aberrant RNA methylation causes transcriptional repression in a manner dependent on the ASCC complex. We show that an alkylated pre-mRNA, or an RNA containing a single damaged base, is sufficient to activate RNF113A E3 activity in a phosphorylation-dependent manner. Together, our work identifies an unexpected role for RNA damage in eliciting a DNA repair response, and suggests that RNA may serve as the “canary in the coal mine” for sensing alkylation damage.

Despite better knowledge of its genetic basis, pancreatic cancer is still highly lethal with very few therapeutic options. In this review, we discuss the potential impact of epigenetic therapies, focusing on lysine methylation signaling and its implication in pancreatic cancer.Protein lysine methylation, a key mechanism of posttranslational modifications of histone proteins, has emerged as a major cell signaling mechanism regulating physiologic and pathologic processes including cancer. This finely tuned and dynamic signaling mechanism is regulated by lysine methyltransferases (KMT), lysine demethylases (KDM) and signal transducers harboring methyl-binding domains. Recent evidence demonstrates that overexpression of cytoplasmic KMT and resulting enhanced lysine methylation is a reversible event that enhances oncogenic signaling through the Ras and Mitogen-Activated Protein Kinases pathway in pancreatic cancer, opening perspectives for new anticancer chemotherapeutics aimed at controlling these activities.The development of potent and specific inhibitors of lysine methylation signaling may represent a hitherto largely unexplored avenue for new forms of targeted therapy in cancer, with great potential for yet hard-to-treat cancers such as pancreatic cancer.