U6 snRNA m6A modification is required for accurate and efficient splicing of C. elegans and human pre-mRNAs.

pre-mRNA splicing is a critical feature of eukaryotic gene expression. Both cis- and trans-splicing rely on accurately recognising splice site sequences by spliceosomal U snRNAs and associated proteins. Spliceosomal snRNAs carry multiple RNA modifications with the potential to affect different stages of pre-mRNA splicing. Here, we show that the conserved U6 snRNA m6A methyltransferase METT-10 is required for accurate and efficient cis- and trans-splicing of C. elegans pre-mRNAs. The absence of METT-10 in C. elegans and METTL16 in humans primarily leads to alternative splicing at 5' splice sites with an adenosine at +4 position. In addition, METT-10 is required for splicing of weak 3' cis- and trans-splice sites. We identified a significant overlap between METT-10 and the conserved splicing factor SNRNP27K in regulating 5' splice sites with +4A. Finally, we show that editing endogenous 5' splice site +4A positions to +4U restores splicing to wild-type positions in a mett-10 mutant background, supporting a direct role for U6 snRNA m6A modification in 5' splice site recognition. We conclude that the U6 snRNA m6A modification is important for accurate and efficient pre-mRNA splicing.

U6 snRNA m6A modification is required for accurate and efficient cis- and trans-splicing of C. elegans mRNAs.

pre-mRNA splicing is a critical feature of eukaryotic gene expression. Many eukaryotes use cis-splicing to remove intronic sequences from pre-mRNAs. In addition to cis-splicing, many organisms use trans-splicing to replace the 5' ends of mRNAs with a non-coding spliced-leader RNA. Both cis- and trans-splicing rely on accurately recognising splice site sequences by spliceosomal U snRNAs and associated proteins. Spliceosomal snRNAs carry multiple RNA modifications with the potential to affect different stages of pre-mRNA splicing. Here, we show that m6A modification of U6 snRNA A43 by the RNA methyltransferase METT-10 is required for accurate and efficient cis- and trans-splicing of C. elegans pre-mRNAs. The absence of U6 snRNA m6A modification primarily leads to alternative splicing at 5' splice sites. Furthermore, weaker 5' splice site recognition by the unmodified U6 snRNA A43 affects splicing at 3' splice sites. U6 snRNA m6A43 and the splicing factor SNRNP27K function to recognise an overlapping set of 5' splice sites with an adenosine at +4 position. Finally, we show that U6 snRNA m6A43 is required for efficient SL trans-splicing at weak 3' trans-splice sites. We conclude that the U6 snRNA m6A modification is important for accurate and efficient cis- and trans-splicing in C. elegans.

Contrasting seasonal and spatial distribution of native and invasive Codium seaweed revealed by targeting species-specific eDNA.

AIM: Codium fragile, an invasive seaweed, has spread widely during the last century, impacting on local seaweed communities through competition and disturbance. Early detection of C. fragile can help on its control and management. Environmental DNA (eDNA) has proved successful for early detection of aquatic invasive species but its potential use for seaweed remains understudied. We used a species-specific eDNA qPCR approach to investigate the spatial distribution, abundance, and coexistence of the invasive C. fragile and three native Codium species (Codium vermilara, Codium tomentosum, and Codium decorticatum) in the Cantabrian Sea. LOCATION: Bay of Biscay, Northern Atlantic Coast of the Iberian Peninsula; two ports, a beach and a rocky cliff. METHODS: We designed species-specific primers in barcoding regions targeting short fragments of the rbcL gene for the invasive Codium species, and the elongation factor Tu (tufA) gene for the native species, to assess their spatial and seasonal distributions using quantitative real-time PCR in samples collected during summer, autumn, and winter. RESULTS: We found seasonal differences in the presence of the invasive Codium fragile and two of the native Codium species, but did not detect C. decorticatum at any point. Species distribution patterns produced with qPCR targeting species-specific eDNA coincided with the known distribution based on previous conventional sampling, with a seasonal alternance of C. fragile and C. vermilara, and a marked dominance of invasive C. fragile in ports, which are known hotspots for invasive species. MAIN CONCLUSIONS: Our results demonstrate the utility of using eDNA for early detection and monitoring of invasive seaweed. Native and invasive Codium spp. displayed significant seasonal and spatial differentiation that needs to be taken into account in risk management. Regular monitoring of ports and adjacent areas using eDNA should help to assess the potential expansion of invasive Codium and the need for management interventions to avoid the displacement of native seaweed.