Dataset Information

SAM homeostasis is regulated by CFI_m-mediated splicing of MAT2A.

ABSTRACT: S-adenosylmethionine (SAM) is the methyl donor for nearly all cellular methylation events. Cells regulate intracellular SAM levels through intron detention of MAT2A, the only SAM synthetase expressed in most cells. The N⁶-adenosine methyltransferase METTL16 promotes splicing of the MAT2A detained intron by an unknown mechanism. Using an unbiased CRISPR knock-out screen, we identified CFI_m25 (NUDT21) as a regulator of MAT2A intron detention and intracellular SAM levels. CFI_m25 is a component of the cleavage factor Im (CFI_m) complex that regulates poly(A) site selection, but we show it promotes MAT2A splicing independent of poly(A) site selection. CFI_m25-mediated MAT2A splicing induction requires the RS domains of its binding partners, CFI_m68 and CFI_m59 as well as binding sites in the detained intron and 3´ UTR. These studies uncover mechanisms that regulate MAT2A intron detention and reveal a previously undescribed role for CFI_m in splicing and SAM metabolism.

SUBMITTER: Scarborough AM

PROVIDER: S-EPMC8139829 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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SAM homeostasis is regulated by CFIm-mediated splicing of MAT2A.

Scarborough Anna M AM Flaherty Juliana N JN Hunter Olga V OV Liu Kuanqing K Kumar Ashwani A Xing Chao C Tu Benjamin P BP Conrad Nicholas K NK

eLife 20210505

S-adenosylmethionine (SAM) is the methyl donor for nearly all cellular methylation events. Cells regulate intracellular SAM levels through intron detention of MAT2A, the only SAM synthetase expressed in most cells. The N6-adenosine methyltransferase METTL16 promotes splicing of the MAT2A detained intron by an unknown mechanism. Using an unbiased CRISPR knock-out screen, we identified CFIm25 (NUDT21) as a regulator of MAT2A intron detention and intracellular SAM levels. CFI< ...[more]

PMID: 33949310

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Project description:BackgroundHepatic stellate cell (HSC) activation induced by transforming growth factor β1 (TGF-β1) plays a pivotal role in fibrogenesis, while the complex downstream mediators of TGF-β1 in such process are largely unknown.MethodsWe performed pharmacoproteomic profiling of the mice liver tissues from control, carbon tetrachloride (CCl4)-induced fibrosis and NPLC0393 administrated groups. The target gene MAT2A was overexpressed or knocked down in vivo by tail vein injection of AAV vectors. We examined NF-κB transcriptional activity on MAT2A promoter via luciferase assay. Intracellular SAM contents were analyzed by LC-MS method.FindingsWe found that methionine adenosyltransferase 2A (MAT2A) is significantly upregulated in the CCl4-induced fibrosis mice, and application of NPLC0393, a known small molecule inhibitor of TGF-β1 signaling pathway, inhibits the upregulation of MAT2A. Mechanistically, TGF-β1 induces phosphorylation of p65, i.e., activation of NF-κB, thereby promoting mRNA transcription and protein expression of MAT2A and reduces S-adenosylmethionine (SAM) concentration in HSCs. Consistently, in vivo and in vitro knockdown of MAT2A alleviates CCl4- and TGF-β1-induced HSC activation, whereas in vivo overexpression of MAT2A facilitates hepatic fibrosis and abolishes therapeutic effect of NPLC0393.InterpretationThis study identifies TGF-β1/p65/MAT2A pathway that is involved in the regulation of intracellular SAM concentration and liver fibrogenesis, suggesting that this pathway is a potential therapeutic target for hepatic fibrosis. FUND: This work was supported by National Natural Science Foundation of China (No. 81500469, 81573873, 81774196 and 31800693), Zhejiang Provincial Natural Science Foundation of China (No. Y15H030004), the National Key Research and Development Program from the Ministry of Science and Technology of China (No. 2017YFC1700200) and the Key Program of National Natural Science Foundation of China (No. 8153000502).

Dataset Information

SAM homeostasis is regulated by CFI_m-mediated splicing of MAT2A.

Publications

SAM homeostasis is regulated by CFI<sub>m</sub>-mediated splicing of MAT2A.

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