RNA Biology最新文献

Gene expression involves a series of consequential processes, beginning with mRNA synthesis and culminating in translation. Traditionally studied as a linear sequence of events, recent findings challenge this perspective, revealing coupling mechanisms that coordinate key steps of gene expression, even when spatially and temporally distant. In this review, we focus on translation, the final stage of gene expression, and examine its coupling with key stages of mRNA metabolism: synthesis, processing, export, and decay. For each of these processes, we provide an overview of known instances of coupling with translation. Furthermore, we discuss the role of high-throughput technologies in uncovering these intricate interactions on a genome-wide scale. Finally, we highlight key challenges and propose future directions to advance our understanding of how coupling mechanisms orchestrate robust and adaptable gene expression programs.

Neural stem cells (NSCs) are multipotent stem cells with self-renewal capacity, able to differentiate into all neural lineages of the central nervous system, including neurons, oligodendrocytes, and astrocytes; thus, their proliferation and differentiation are essential for embryonic neurodevelopment and adult brain homoeostasis. Dysregulation in these processes is implicated in neurological disorders, highlighting the need to elucidate how NSCs proliferate and differentiate to clarify the mechanisms of neurogenesis and uncover potential therapeutic targets. MicroRNAs (miRNAs) are small, post-transcriptional regulators of gene expression involved in many aspects of nervous system development and function. Multiple studies have shown that miRNAs control the balance between self-renewal and differentiation during development through transcriptional networks and fine-tuned signalling pathways. They also regulate key biological processes, including cell fate determination, developmental timing, neurogenesis, gliogenesis, and apoptosis. Transcriptomic analyses and high-resolution profiling have revealed temporally and spatially restricted miRNA expression patterns in NSCs and their progeny, suggesting highly context-dependent regulatory functions. Here, we provide an integrated overview of recent advances in miRNA biology relevant to NSC maintenance and lineage specification, with a focus on the mechanistic understanding of miRNA roles in neuronal differentiation, glial development, and programmed cell death across neural development.

Small Cajal body-specific RNAs (scaRNAs) are noncoding RNAs involved in the maturation of U-rich small nuclear RNAs. Except for a few that have their own transcription units, most scaRNA genes are embedded in introns and are predicted to be transcribed with host genes. Herein, we report that scaRNA28 is the first scaRNA with a dual synthesis pathway, and that this RNA is transcribed in an independent transcription unit (ITU) by RNA polymerase II while located in intron 2 of the transformation/transcription domain-associated protein (TRRAP) gene. We evaluated the scaRNA28 synthesis pathway using minigenes containing exon 2, intron 2, and exon 3 of TRRAP. A minigene with a mutation preventing 5' splicing recognition of the exon 2/intron 2 junction generated scaRNA28, suggesting a pathway processing unspliced transcripts into scaRNA28. Even promoterless minigenes and DNA fragments with regions from exons 2 to 3 of TRRAP showed RNA polymerase II-dependent synthesis of scaRNA28, indicating a novel synthesis pathway involving an ITU. Linker-scanning mutational analysis revealed that the promoter region required for scaRNA28 expression in the ITU is located within 60 bases including exon 2/intron 2 junction of TRRAP, and especially the first two bases of intron 2 region, a putative part of the MYC-binding (E-box) motif, are essential for scaRNA28 expression in the ITU. MYC promotes scaRNA28 expression by binding to the promoter region in the ITU. Our findings demonstrated a novel transcriptional pathway for the synthesis of scaRNA28, the first scaRNA with a dual synthesis pathway.

Dysregulated translation is a hallmark of cancer, and recent genome-wide studies in tumour cells have uncovered widespread translation of non-canonical reading frames that often initiate at non-AUG codons. If an upstream non-canonical start site is located within a frame with an annotated coding sequence (CDS), such translation events can lead to the production of proteoforms with altered N-termini (PANTs). Certain examples of PANTs from oncogenes (e.g. c-MYC) and tumour suppressors (e.g. PTEN) have been previously linked to cancer. We have performed a systematic computational analysis on recently identified non-AUG initiation-derived N-terminal extensions of cancer-associated proteins, and we discuss how these extended proteoforms may acquire new oncogenic properties. We identified a loss of stability for the N-terminally extended proteoforms of oncogenes TCF-4 and SOX2. Furthermore, we discovered likely functional short linear motifs within the N-terminal extensions of oncogenes and tumour suppressors (SOX2, SUFU, SFPQ, TOP1 and SPEN/SHARP) that could provide an explanation for previously described functionalities or interactions of the proteins. In all, we identify novel cases where PANTs likely show different localization, functions, partner binding or turnover rates compared to the annotated proteoforms. Therefore, we propose that alterations in the stringency of translation initiation, often seen under conditions of cellular stress, may result in reprogramming of translation to generate novel PANTs that influence cancer progression.

In all domains of life, Archaea, Eukarya and Bacteria, the unusual amino acid selenocysteine (Sec) is co-translationally incorporated into proteins by recoding a UGA stop codon to a sense codon. A secondary structure on the mRNA, the selenocysteine insertion sequence (SECIS), is required, but its position, secondary structure and binding partner(s) are not conserved across the tree of life. Thus far, the nature of archaeal SECIS elements has been derived mainly from sequence analyses. A recently developed in vivo reporter system was used to study the structure-function relationships of SECIS elements in Methanococcus maripaludis. Through targeted mutagenesis, we defined the minimal functional SECIS element, the parts of the SECIS where structure and not the identity of the bases are relevant for function, and identified two conserved -and invariant- adenines that are most likely to interact with the other factor(s) of the Sec recoding machinery. Finally, we demonstrated the functionality of SECIS elements in the 5`-untranslated region of the mRNA and identified a potential mechanism of SECIS repositioning in the vicinity of the UGA for efficient selenocysteine insertion.

Iron (Fe) plays critical roles as enzyme cofactor involved in key biological processes but can also lead to toxicity by catalysing the formation of highly damaging reactive oxygen species. To stabilize Fe and perform catalysis, most organisms rely on Fe-S clusters, which are fundamental and evolutionary ancient cofactors. In E. coli, two distinct pathways for the biosynthesis of Fe-S cluster exist: the three-part iscR-SUA-hscBA-fdx-iscX (ISC-HSC) operon and the sufABCDSE (SUF) operon. The iscR-SUA section of the ISC-HSC operon is regulated at the promoter level by the IscR transcription factor and post-transcriptionally by the small RNA (sRNA) RyhB. The SUF operon is regulated by a combination of transcription factors, including the Fe-sensing Fur, the Fe-S using IscR, and the oxidative stress responsive OxyR. Here, we show evidence that the sRNA RyhB regulates the hscBA-fdx-iscX part of the ISC-HSC operon as well as part of the SUF operon. RyhB orchestrates a complex pattern of expression of the iscR-SUA-hscBA-fdx-iscX operon during Fe starvation. This results in increased level of iscR and constant expression of iscSUA, encoding the scaffold for Fe-S cluster formation. However, the third part of the operon, hscBA-fdx-iscX, encoding a chaperone that facilitates Fe-S cluster transfer, is repressed by RyhB during Fe starvation. Furthermore, RyhB represses part of the sufABCDSE transcript, which counteracts Fur derepression. Overall, RyhB represses both ISC and SUF systems under iron starvation, to reduce Fe-S biogenesis under such limiting conditions.

Sorafenib (Sfb) is a multikinase inhibitor regularly used for the management of patients with advanced hepatocellular carcinoma (HCC) that has been shown to increase very modestly life expectancy. We have shown that Sfb inhibits protein synthesis at the level of initiation in cancer cells. However, the global snapshot of mRNA translation following Sorafenib-treatment has not been explored so far. In this study, we performed a genome-wide polysome profiling analysis in Sfb-treated HCC cells and demonstrated that, despite global translation repression, a set of different genes remain efficiently translated or are even translationally induced. We reveal that, in response to Sfb inhibition, translation is tuned, which strongly correlates with the presence of established mRNA cis-acting elements and the corresponding protein factors that recognize them, including DAP5 and ARE-binding proteins. At the level of biological processes, Sfb leads to the translational down-regulation of key cellular activities, such as those related to the mitochondrial metabolism and the collagen synthesis, and the translational up-regulation of pathways associated with the adaptation and survival of cells in response to the Sfb-induced stress. Our findings indicate that Sfb induces an adaptive reprogramming of translation and provides valuable information that can facilitate the analysis of other drugs for the development of novel combined treatment strategies based on Sfb therapy.

Cellular senescence is a stable cell cycle arrest associated with upregulated inflammatory responses. Senescent cells contribute to various pathological and physiological processes including organismal ageing and cancer. Cellular senescence can be induced by various cellular stresses including DNA damage, telomere shortening, oncogene activation, and epigenetic alterations. We have shown that plasma membrane damage can also induce cellular senescence. However, common and specific molecular mechanisms among different senescent cell subtypes remain unknown. MicroRNAs (miRNAs) regulate mRNA and rewire gene expression profiles, contributing to multiple processes including cellular senescence. Here, we performed time-resolved miRNA sequencing and compared the results with mRNA sequencing results using cells experiencing plasma membrane damage-dependent senescence (PMD-Sen) and cells undergoing DNA damage response-dependent senescence (DDR-Sen). We found 65 miRNAs that are differentially regulated in PMD-Sen, contributing to 2,495 miRNA-mRNA pairs. Moreover, PMD-Sen and DDR-Sen shared 41 miRNAs across their sets of miRNA-mRNA pairs. Notably, miR-155-5p emerged as the miRNA with the largest number of shared miRNA-mRNA pairs that exhibit a highly negative correlation. These results highlight miR-155-5p as the potential key regulator of PMD-Sen and DDR-Sen.

The lack of a sufficient number of validated miRNA targets severely hampers the understanding of their biological function. Even for the well-studied miR-155-5p, there are only 239 experimentally validated targets out of 42,554 predicted targets. For a more complete assessment of the immune-related miR-155 targetome, we used an inverse correlation of time-resolved mRNA profiles and miR-155-5p expression of early CD4+ T cell activation to predict immune-related target genes. Using a high-throughput miRNA interaction reporter (HiTmIR) assay we examined 90 target genes and confirmed 80 genes as direct targets of miR-155-5p. Our study increases the current number of verified miR-155-5p targets approximately threefold and exemplifies a method for verifying miRNA targetomes as a prerequisite for the analysis of miRNA-regulated cellular networks.

The reprogramming of alternative splicing networks during development is a hallmark of tissue maturation and identity. Alternative splicing of microexons (small, genomic regions ≤ 51 nucleotides) functionally regulate protein-protein interactions in the brain and is altered in several neuronal diseases. However, little is known about the regulation and function of alternatively spliced microexons in striated muscle. Here, we investigated alternative splicing of a microexon in the synaptosome-associated protein 23 (Snap23) encoded gene. We found that inclusion of this microexon is developmentally regulated and tissue-specific, as it occurs exclusively in adult heart and skeletal muscle. The alternative region is highly conserved in mammalian species and encodes an in-frame sequence of 11 amino acids. Furthermore, we showed that alternative splicing of this microexon is mis-regulated in mouse models of heart and skeletal muscle diseases. We identified the RNA-binding proteins (RBPs) quaking (QKI) and RNA binding fox-1 homolog 2 (RBFOX2) as the primary splicing regulators of the Snap23 microexon. We found that QKI and RBFOX2 bind downstream of the Snap23 microexon to promote its inclusion, and this regulation can be escaped when the weak splice donor is mutated to the consensus 5' splice site. Finally, we uncovered the interplay between QKI and muscleblind-like splicing regulator (MBNL) as an additional, but minor layer of Snap23 microexon splicing control. Our results are one of the few reports detailing microexon alternative splicing regulation during mammalian striated muscle development.