Inhibition of α-KG-dependent histone and DNA demethylases by fumarate and succinate that are accumulated in mutations of FH and SDH tumor suppressors

Abstract

Two Krebs cycle genes, fumarate hydratase (FH) and succinate dehydrogenase (SDH), are mutated in a subset of human cancers, leading to accumulation of their substrates, fumarate and succinate, respectively. Here we demonstrate that fumarate and succinate are competitive inhibitors of multiple a-ketoglutarate (a-KG)-dependent dioxygenases, including histone demethylases, prolyl hydroxylases, collagen prolyl-4-hydroxylases, and the TET (ten-eleven translocation) family of 5-methlycytosine (5mC) hydroxylases. Knockdown of FH and SDH results in elevated intracellular levels of fumarate and succinate, respectively, which act as competitors of a-KG to broadly inhibit the activity of a-KG-dependent dioxygenases. In addition, ectopic expression of tumor-derived FH and SDH mutants inhibits histone demethylation and hydroxylation of 5mC. Our study suggests that tumor-derived FH and SDH mutations accumulate fumarate and succinate, leading to enzymatic inhibition of multiple a-KG-dependent dioxygenases and consequent alterations of genome-wide histone and DNA methylation. These epigenetic alterations associated with mutations of FH and SDH likely contribute to tumorigenesis.[Keywords: FH; SDH; metabolites; a-KG-dependent dioxygenases; DNA methylation; histone methylation] Supplemental material is available for this article. Received March 7, 2012; revised version accepted May 9, 2012. Several lines of evidence, including the recent identification of mutations affecting isocitrate dehydrogenase (IDH), fumarate hydratase (FH), and succinate dehydrogenase (SDH), have demonstrated that mutations in certain metabolic enzymes may play a causal role in tumorigenesis. The NADP + -dependent IDH genes IDH1 and IDH2 are frequently mutated in >75% of glioma (Parsons et al. 2008),

Succinate dehydrogenase deficiency is associated with decreased 5-hydroxymethylcytosine production in gastrointestinal stromal tumors: implications for mechanisms of tumorigenesis

“…Recent in vitro work has demonstrated that the accumulation of succinate in SDH-deficient tumors may lead to the inhibition of a-ketoglutarate-dependent enzymes, including TET family proteins, which convert 5-methylcytosine to 5-hmC. 26 In the current study, we examined 5-hmC levels by immunohistochemistry in a cohort of 30 genotyped GISTs and demonstrate that 5-hmC staining is absent in 90% of SDH-deficient GISTs, whereas loss of 5-hmC staining is seen in approximately 20% of KIT-or PDGFRA-mutant GISTs. The alteration of 5-hmC content through inhibition of TET activity seen in the SDH-deficient GISTs demonstrates the dysregulation of epigenetic modifications in these tumors.…”

“…24,25 Because of structural similarities between succinate and a-ketoglutarate, succinate is thought to inhibit the activity of a-ketoglutarate-dependent dioxygenase enzymes. 26 In particular, recent work has demonstrated that the TET family of DNA hydroxylases may be inhibited by elevated levels of succinate. 26 TET proteins have been shown to have a role in the regulation of DNA methylation.…”

“…26 In particular, recent work has demonstrated that the TET family of DNA hydroxylases may be inhibited by elevated levels of succinate. 26 TET proteins have been shown to have a role in the regulation of DNA methylation. Methylation of cytosine at the 5-position generates 5-methylcytosine, an important modification in the epigenetic regulation of gene expression.…”

“…Indeed, recent work has shown that siRNA-mediated knockdown of SDHA in cultured cells and in mice leads to an inhibition of TET activity and decreased levels of 5-hmC. 26 However, it remains unknown whether SDH deficiency leads to alterations in TET activity in human tumors. Therefore, in this study, we evaluated 5-hmC levels in a cohort of genotyped GISTs to determine whether loss of SDH is associated with inhibition of TET activity.…”

Mitochondrial metabolic remodeling in response to genetic and environmental perturbations

“…Deficiencies in FH lead to high steady‐state concentrations of fumarate in the cell and surrounding extracellular space. In parallel to succinate, fumarate has been shown to competitively inhibit αKG‐dependent dioxygenase enzymes, resulting in a pseudohypoxic transcriptome through HIFα stabilization76 and a hypermethylator phenotype 76, 77. However, contrary to initial thoughts, now there is evidence to suggest that stabilization of HIF is not the driver of tumorigenesis in FH‐deficient tumors, and that a different candidate transcription factor—the nuclear‐related factor 2 (NRF2) pathway—may instead be involved 78, 79…”