Extensive pre-mRNA back-splicing generates numerous circular RNAs (circRNAs) in human transcriptome. However, the biological functions of these circRNAs remain largely unclear. Here we report that N⁶-methyladenosine (m⁶A), the most abundant base modification of RNA, promotes efficient initiation of protein translation from circRNAs in human cells. We discover that consensus m⁶A motifs are enriched in circRNAs and a single m⁶A site is sufficient to drive translation initiation. This m⁶A-driven translation requires initiation factor eIF4G2 and m⁶A reader YTHDF3, and is enhanced by methyltransferase METTL3/14, inhibited by demethylase FTO, and upregulated upon heat shock. Further analyses through polysome profiling, computational prediction and mass spectrometry reveal that m⁶A-driven translation of circRNAs is widespread, with hundreds of endogenous circRNAs having translation potential. Our study expands the coding landscape of human transcriptome, and suggests a role of circRNA-derived proteins in cellular responses to environmental stress.

Depression and the Habenula: The lateral habenula (LHb) appears to have a role in depression. However, the underlying mechanisms are poorly understood, and by using multiple rodent models of depression, Li et al. (p. 1016 ) identified a signaling pathway and associated neuronal adaptations in which the enzyme βCaMKII was selectively up-regulated in the LHb. Manipulations that enhanced βCaMKII levels increased depression-related phenotypes, and RNA interference of CaMKIIb blunted depression. Enhanced βCaMKII levels in the habenula promoted excitatory synaptic transmission on these neurons and increased action potential firing mediated by an up-regulation of a specific subtype of glutamate receptors.

Structure and function of the spliceosome: When RNA is transcribed from DNA in the eukaryotic cell nucleus, the initial transcript includes noncoding introns that must be spliced out. This splicing is done by a complex macromolecular machine, the spliceosome, which comprises five small nuclear RNAs and more than 100 associated proteins. Now, two papers reveal insights into the structure and function of the yeast spliceosome. Yan et al. describe a high-resolution structure determined by electron microscopy of a spliceosome complex comprising four RNAs and 37 proteins. Hang et al. focus on the catalytic site and show how protein components anchor the transcribed RNA and allow sufficient flexibility to deliver RNA components involved in catalyzing the splicing reaction. Science , this issue pp. 1182 and 1191

The precise and large-scale identification of intact glycopeptides is a critical step in glycoproteomics. Owing to the complexity of glycosylation, the current overall throughput, data quality and accessibility of intact glycopeptide identification lack behind those in routine proteomic analyses. Here, we propose a workflow for the precise high-throughput identification of intact N-glycopeptides at the proteome scale using stepped-energy fragmentation and a dedicated search engine. pGlyco 2.0 conducts comprehensive quality control including false discovery rate evaluation at all three levels of matches to glycans, peptides and glycopeptides, improving the current level of accuracy of intact glycopeptide identification. The N-glycoproteome of samples metabolically labeled with ¹⁵N/¹³C were analyzed quantitatively and utilized to validate the glycopeptide identification, which could be used as a novel benchmark pipeline to compare different search engines. Finally, we report a large-scale glycoproteome dataset consisting of 10,009 distinct site-specific N-glycans on 1988 glycosylation sites from 955 glycoproteins in five mouse tissues.