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

Anna M Scarborough,Ashwani Kumar,Chao Xing,Olga V Hunter,Nicholas K Conrad,Kuanqing Liu,Benjamin P Tu,Juliana N Flaherty

doi:10.7554/elife.64930

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 CFIm25 (NUDT21) as a regulator of MAT2A intron detention and intracellular SAM levels. CFIm25 is a component of the cleavage factor Im (CFIm) complex that regulates poly(A) site selection, but we show it promotes MAT2A splicing independent of poly(A) site selection. CFIm25-mediated MAT2A splicing induction requires the RS domains of its binding partners, CFIm68 and CFIm59 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 CFIm in splicing and SAM metabolism.

Highlights

S-adenosylmethionine (SAM) is the universal methyl donor, essential for the methylation of DNA, RNA, and proteins that regulates most cellular functions
Our reporter construct consists of GFP, a β-globin intron with flanking exonic regions, the MAT2A detained intron (DI) with flanking exons, and full-length MAT2A 3 ́ untranslated region (3 ́UTR) driven by a CMV promoter (Figure 1A)
In SAM-replete conditions, the MAT2A intron should be inefficiently spliced and produce little GFP protein, while SAM depletion should lead to efficient splicing, mRNA stabilization, and robust GFP signal

Summary

Introduction

S-adenosylmethionine (SAM) is the universal methyl donor, essential for the methylation of DNA, RNA, and proteins that regulates most cellular functions. Humans express a single SAM synthetase, encoded by the gene MAT2A, in nearly every tissue except the liver (Murray et al, 2019). Upon donating a methyl group, SAM converts to S-adenosylhomocysteine (SAH), which inhibits some methyltransferases (Ferreira de Freitas et al, 2019). Intracellular SAM levels are tightly controlled to maintain homeostasis, with loss of regulation resulting in cellular dysfunction (Gao et al, 2019; Lio and Huang, 2020; Ouyang et al, 2020; Parkhitko et al, 2019)

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