Down the Drain? 1MNA Acts as a Potent ‘Methylation Sink’ in Cancer Cells
The cancer field consistently proves to be one of the most critical areas for investigations into mechanisms governing tight epigenetic regulation, as nearly all types of cancers demonstrate misregulated epigenetic patterns. Most studies focus on the final epigenomic landscape in terms of histone and DNA modifications and methylomes; relatively few, however, have placed as much focus on the molecular “paints and brushes” responsible for creating the dynamic epigenomic “canvas”. In a recent study by a team at Scripps, new light has been shed on the upstream processes that determine the availability of substrates essential for epigenetic regulation.
Ulanovskaya and colleagues investigated the close relationship between nicotinamide N-methyltransferase (NNMT) and the methylation potential of cancer cells using an unbiased metabolomics approach. NNMT is responsible for catalyzing the transfer of a methyl group from S-adenosyl methionine (SAM) to nicotinamide to yield 1-methylnicotinamide (1MNA) and S-adenosyl homocysteine (SAH) as products. Importantly, NNMT is typically overexpressed in cancer cells, and the researchers found that the levels of this enzyme correlated directly with aggressiveness or invasiveness of the cancer. The team created a carcinoma-derived cell line to overexpress NNMT and utilized mass spectrometry techniques to survey the levels of metabolites in these cells and several other cancer lines. The presence and levels of metabolites within each sample were compared, and in each comparison, SAH and 1MNA were the only metabolites found to be deregulated in the cells with increased NNMT activity.
In order to confirm that these differences could be attributed to the NNMT activity, the team created cells expressing a catalytically inactive form of NNMT and also performed siRNA-mediated knockdowns of NNMT. These cells did not demonstrate the elevated 1MNA or SAH levels seen in the aggressive cancer cells, providing further evidence for a role for NNMT in epigenetic regulation of cancer phenotypes. Further, while the cells overexpressing NNMT and the aggressive cancer cells demonstrated both elevated SAH and depleted SAM, the catalytic mutant and NNMT knockdown cells demonstrated the opposite trend – that is, their SAM content remained the same or increased, while the SAH levels decreased. As the ratio of SAM to SAH in the cells represents the methylation potential, the aggressive cancer cells with increased NNMT activity can then be defined as having a substantially lower methylation potential with a lower availability of methylation units for posttranslational modifications of proteins, nucleic acids, and fatty acids. Via utilization of deuterated 1MNA supplemented to the cells, the authors confirmed that methylation stored in the molecule is not further involved in cellular metabolism, as no other metabolites are detected with deuterated positions, confirming that these methyl groups are no longer available to modify other substrates.
After pinpointing 1MNA as a ‘methylation sink’, the authors queried which downstream substrates demonstrated the most pronounced methylation deficiencies. While the team found that there were no significant NNMT-dependent changes in global cytosine methylation, protein methylation was found to be markedly lower in cells with increased NNMT levels (and correspondingly lower methylation potentials). Specifically, histone methylation was shown to be widely reduced across several genomic locations. Interestingly, expression of many of the genes with dysregulated histone methylation were previously shown to be inappropriately regulated in the aggressive cancer types, which the authors also confirmed with their own gene expression analyses. The researchers demonstrated that the deficiencies in protein methylation were not limited to histones, but were also apparent in several non-histone proteins, including tumor suppressors.
This study represents an important step towards a more comprehensive understanding of cancer epigenomics through the lens of complete metabolomics. As a result of the mechanisms elucidated in this work, 1MNA and NNMT could serve as viable therapeutic targets for a sort of ‘delicate tuning’ of the metabolic events associated with some of the most aggressive cancers.
Ulanovskaya OA, Zuhl AM, & Cravatt BF (2013). NNMT promotes epigenetic remodeling in cancer by creating a metabolic methylation sink. Nature chemical biology, 9 (5), 300-6 PMID: 23455543