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Technological advances in the past decade or so have revolutionized our understanding of the co-transcriptional and post-transcriptional control of gene expression. It has become clear that the different steps of pre-mRNA processing, from 5' capping to nuclear export, are tightly co-regulated and coordinated with transcription, translation and mRNA decay. Furthermore, recent discoveries and the functional characterization of several chemical modifications in mRNA such as N6-methyladenosine (m6A) and 5-methylcytosine (m5C), which together form the epitranscriptome, have revealed a new layer of regulation of mRNA processing and maturation. The articles in this series discuss different aspects of the biogenesis of mRNAs and other RNA species (including microRNAs, rRNAs, tRNAs, small RNAs and other non-coding RNAs), the regulation of RNA processing and chemical modification, and how these processes affect human physiology and disease.
Nucleobase modifications are prevalent in eukaryotic mRNA and are broadly required for post-transcriptional gene regulation. The most studied mRNA modification is N6-methyladenosine (m6A), yet various other modifications are now being identified and studied. This Review discusses the emerging mechanisms and roles of these non-m6A modifications.
Alternative cleavage and polyadenylation (APA) generates mRNA isoforms with alternative 3′ untranslated regions; these isoforms modulate protein abundance and functionality, including through subcellular localization of mRNA and translation. APA is modulated by signalling pathways that control co-transcriptional and post-transcriptional processes, and its dysregulation affects cell responses to environmental changes.
Recent studies have revolutionized our understanding of the interplay between mRNA poly(A) tails and the processes of translation and mRNA decay in the cytoplasm. Poly(A) tails interact with dedicated RNA-binding proteins and deadenylases, which together determine the impact of poly(A) tails on gene expression.
The non-canonical addition of non-templated nucleotides to RNA 3′ ends (tailing) by terminal nucleotidyltransferases includes uridylation, mixed-nucleotide tailing and post-transcriptional polyadenylation. Recent studies of human terminal nucleotidyltransferases have revealed their distinct specificities for substrates, including mRNAs, microRNAs and other non-coding RNAs, and how they control RNA stability and activity.
Circular RNAs, which are produced through back-splicing of exons, are emerging as key regulators of immune responses and cell proliferation. Recent studies have shed new light on the biogenesis and functions of circular RNAs, which include the modulation of transcription and splicing, and interference with microRNAs and other cellular signalling pathways.
The cleavage of microRNA (miRNA) precursors by Drosha and Dicer and their loading with Argonaute proteins into RNA-induced silencing complexes are key steps in miRNA biogenesis. Recent studies have clarified the mechanisms of action of these molecular machines and discovered non-canonical miRNA biogenesis pathways.
tRNAs exist as diverse species, including sequence isoforms and nuclease-generated fragments, which are further functionally diversified by base modifications and various protein interactions. Perhaps unsurprisingly, tRNAs are now being implicated in various cellular processes beyond protein synthesis per se, including in stress responses, proliferation, cell fate determination and tumorigenesis.
Atomic-resolution structures have recently been obtained for the intact spliceosome at different stages of the splicing cycle. These structural data have proved that the spliceosome is a protein-directed metalloribozyme and have increased our understanding of pre-mRNA splicing mechanisms, explaining a large body of existing genetic and biochemical data.
Pre-mRNA splicing occurs on nascent RNA, which is attached to chromatin by RNA polymerase II. Much splicing occurs co-transcriptionally, and the spatial and temporal coordination of the two processes is tightly coordinated with other mRNA-processing events.
Alternative splicing expands the complexity of the proteome by generating multiple transcript isoforms from a single gene. Numerous alternative splicing events occur during cell differentiation and tissue maturation, suggesting that alternative splicing supports proper development. Recent studies shed light on how alternative splicing and its coordination contribute to organ development and tissue homeostasis.
The chemical modifications and structural features of mRNAs are highly dynamic. Together, they regulate the composition and function of the transcriptome by shaping RNA–protein interactions at different stages of the gene expression process.
Genetic variants can produce phenotypic traits through effects on RNA processing, including effects on pre-mRNA splicing, 3′ end formation, and RNA stability, localization, structure and translation efficiency.
Reversible mRNA methylation is an emerging mode of eukaryotic post-transcriptional gene regulation.N6-methyladenosine (m6A) affects mRNA processing, translation and decay during cell differentiation, embryonic development and stress responses. Other mRNA modifications — N1-methyladenosine (m1A), 5-methylcytosine (m5C) and pseudouridine — together with m6A code a new layer of information that controls protein synthesis.
Alternative polyadenylation (APA) generates mRNAs with varying 3′ termini. It is regulated by variation in the concentration of cleavage and polyadenylation factors and by RNA-binding proteins, as well as by splicing and transcription. APA is important for cell proliferation and differentiation owing to its roles in mRNA metabolism and protein diversification.
RNA helicases can either remodel or lock the composition of messenger ribonucleoprotein complexes, and thus they have pleiotropic functions in the regulation of gene expression. RNA helicases can drive the progression of mRNAs between various RNA-processing factories, leading to protein synthesis or to mRNA storage or decay.
Circular RNAs (circRNAs) are produced from precursor RNA back-splicing. Recent findings reveal the complexity of the biogenesis of circRNAs and their cell type-specific expression. They also show that circRNAs can shape eukaryotic transcriptomes by sequestering microRNAs and by regulating transcription and interfering with splicing.
The versatile RNA-degradation functions of the RNA exosome complex make it crucial for RNA biogenesis. It is now emerging that the nuclear exosome is a specific regulator of gene expression in different physiological processes, and that it has a role in transcription regulation and in maintaining genome stability.