Alternative splicing of pre-mRNAs can regulate gene expression levels by coupling with nonsense-mediated mRNA decay (NMD). In order to elucidate a repertoire of mRNAs regulated by alternative splicing coupled with NMD (AS-NMD) in an organism, we performed long-read RNA sequencing of poly(A) + RNAs from an NMD-deficient mutant strain of Caenorhabditis elegans, and obtained full-length sequences for mRNA isoforms from 259 highconfidence AS-NMD genes. Among them are the S-adenosyl-L-methionine (SAM) synthetase (sams) genes sams-3 and sams-4. SAM synthetase activity autoregulates sams gene expression through AS-NMD in a negative feedback loop. We furthermore find that METT-10, the orthologue of human U6 snRNA methyltransferase METTL16, is required for the splicing regulation in vivo, and specifically methylates the invariant AG dinucleotide at the distal 3 0 splice site (3 0 SS) in vitro. Direct RNA sequencing coupled with machine learning confirms m 6 A modification of endogenous sams mRNAs. Overall, these results indicate that homeostasis of SAM synthetase in C. elegans is maintained by alternative splicing regulation through m 6 A modification at the 3 0 SS of the sams genes.

The nematode Caenorhabditis elegans has a single gene, lev-11, encoding multiple tropomyosin isoforms. Two seventh exons are alternatively selected in the head and body regions of muscle cells and differentially affect muscle contractility in the respective regions.

Tropomyosin isoforms contribute to generation of functionally divergent actin filaments. In the nematode Caenorhabditis elegans, multiple isoforms are produced from lev-11, the single tropomyosin gene, by combination of two separate promoters and alternative pre-mRNA splicing. In this study, we report that alternative splicing of lev-11 is regulated in a tissue-specific manner so that a particular tropomyosin isoform is expressed in each tissue. Reverse-transcription polymerase chain reaction analysis of lev-11 mRNAs confirms five previously reported isoforms (LEV-11A, LEV-11C, LEV-11D, LEV-11E and LEV-11O) and identifies a new sixth isoform LEV-11T. Using transgenic alternative-splicing reporter minigenes, we find distinct patterns of preferential exon selections in the pharynx, body wall muscles, intestine and neurons. The body wall muscles preferentially process splicing to produce high-molecular-weight isoforms, LEV-11A, LEV-11D and LEV-11O. The pharynx specifically processes splicing to express a low-molecular-weight isoform LEV-11E, whereas the intestine and neurons process splicing to express another low-molecular-weight isoform LEV-11C. The splicing pattern of LEV-11T was not predominant in any of these tissues, suggesting that this is a minor isoform. Our results suggest that regulation of alternative splicing is an important mechanism to express proper tropomyosin isoforms in particular tissue and/or cell types in C. elegans.

The front cover image is based on the Research Article Alternative splicing of the Caenorhabditis elegans lev‐11 tropomyosin gene is regulated in a tissue‐specific manner by Eichi Watabe et al., DOI: https://doi.org/10.1002/cm.21489.