Many of the recent exciting advances in the therapy and understanding of myeloid leukemia pathogenesis have centered on the process of DNA methylation. This includes the discovery of mutations in enzymes that methylate DNA (such as mutations in DNMT3A) and the machinery involved in DNA demethylation (including TET2 and IDH1/2 mutations). Additionally, several therapies used commonly in myeloid leukemias impact the process of DNA methylation such as DNA methyltransferase inhibitors (decitabine and 5-azacitidine), as well as mutant IDH1/2 inhibitors.
While these advances have focused on the idea that DNA residues can undergo reversible methylation, it has been known for some time that RNA residues can also be modified in a reversible manner (reviewed recently1,2). In fact, more than 100 different RNA modifications have been discovered, but the functional significance of modifications on RNA is largely not well understood. Now, two studies have highlighted that the enzyme METTL3, which places a methylation mark on RNA known as N6-methyladenosine (or “m6A”), may be a novel dependency in acute myeloid leukemia (AML). The m6A modification is placed on coding as well as noncoding RNAs and has been shown to play a key role in regulating mRNA translation to produce protein. Although they used distinct approaches, both studies found that METTL3 is required for the survival of AML cells, but not normal human or mouse hematopoietic progenitors, and that METTL3 is upregulated at the mRNA and protein level in AML compared to normal cells. In fact, in both reports, overexpression of wild-type METTL3, but not an enzymatic dead version, enhanced the growth of AML cells.
Given the results shown here, one key question is how an enzyme that methylates RNA would be required for AML cell survival and promote leukemogenesis. To address this question, Dr. Ly P. Vu and colleagues mapped the location of m6A in the RNA of AML cells at single-nucleotide resolution. Because the m6A modification on RNA has been previously shown to affect the translation of mRNAs by the ribosome, the authors also determined the mRNAs that were occupied by the ribosome in the presence and absence of m6A. This effort revealed that m6A-containing transcripts regulated by METTL3 conspicuously included c-MYC, BCL2, and PTEN — three factors that are well known to be proto-oncogenic. METTL3 knockdown influenced the translation of mRNAs encoding these transcripts and, consequently, regulated protein levels of MYC, BCL2, and PTEN.
Similar to the results by Dr. Vu and colleagues, Dr. Isaia Barbieri and team identified that METTL3 is also required for AML cell survival using a CRISPR/Cas9 screen focusing on a large number of genes encoding known or putative RNA-modifying enzymes. This revealed that Mettl3, Mettl14, Mettl1, and Mettl16 are all required in AML cells. Given that Mettl3 and Mettl14 form a complex (with METTL3 encoding the enzymatic component of the complex), they decided to focus on METTL3/METTL14 for further studies. Nonetheless, the results from their CRISPR screen suggest that numerous additional RNA-modifying enzymes may be distinctly required in AML over normal cells.
One key question related to the study of RNA m6A catalysis is how the METTL3/METTL14 enzymatic complex is recruited to its RNA substrates. Dr. Barbieri and coauthors therefore tested whether METTL3 might regulate RNA methylation of transcripts by binding to chromatin encoding these RNAs. They thus performed anti-METTL3 ChIP-seq (chromatin immunoprecipitation followed by next-generation sequencing) and found that 1) METTL3 binds chromatin, largely at transcription start sites, and 2) transcripts derived from METTL3-bound promoters harbor m6A within their coding sequence. These data suggest that METTL3 regulates mRNAs derived from their chromatin target genes.
Overall, these two studies identify an enzyme that appears to be overexpressed in AML and on which AML cells can become dependent. While the molecular basis for this unique requirement of METTL3 in AML over normal cells is not yet clear, these studies provide a strong rationale for the development of pharmacologic approaches to block m6A placement as a novel therapy for AML. It will also be interesting to study whether METTL3 plays a role in normal hematopoiesis at any point and if it is distinctly required in other myeloid malignancies such as myelodysplastic syndromes or myeloproliferative neoplasms. Finally, the “eraser” of the m6A mark (an enzyme known as ALKBH5) is an α-ketoglutarate dependent enzyme, the activity of which may be influenced by IDH1/2 mutations. Further studies to clarify the role of ALKBH5 in cancer pathogenesis will therefore be important as well.
Yue Y, Liu J, He C. RNA N6-methyladenosine methylation in post-transcriptional gene expression regulation. Genes Dev. 2015;29:1343-1355.
Jaffrey SR, Kharas MG. Emerging links between m6A and misregulated mRNA methylation in cancer. Genome Med. 2017;9:2.
Conflict of Interests
Dr. Abdel-Wahab indicated no relevant conflicts of interest.
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