RNA Biology最新文献

mRNAs undergo a series of chemical modifications to become competent for nuclear export and translation. This is referred to as mRNA maturation or processing and includes capping, splicing, and 3'end formation. These steps can be hijacked in cancer to alter proteins' forms and levels in the absence of mutation or changes to transcript levels. Here, we focus on an emerging idea that some factors act in multiple processing events and that their dysregulation in both their canonical and noncanonical functions contributes to cancer with a focus on Acute Myeloid Leukaemia (AML). As examples, we discuss the eukaryotic translation initiation factor (eIF4E), splice factor 3 complex B subunit 1 (SF3B1), U2 small nuclear auxiliary factor (U2AF1), and associated factors. These physically interact with each other and play roles in splicing, export, and translation. Malignant dysregulation of this mRNA processing-export-translation axis diversifies the proteome to support cancer. Finally, we discuss the simultaneous dysregulation of mRNA processing in malignancy and related therapeutic development.

Fusion transcripts (FTs) are RNA molecules, also known as chimeric transcripts, formed through chromosomal rearrangements or transcriptional processes, contributing to tumorigenesis. This study systematically examined tumour-specific FTs in hepatocellular carcinoma (HCC) using high-throughput RNA sequencing data from independent datasets and The Cancer Genome Atlas (TCGA). Our meta cohort analysis included 328 HCC samples. Using STAR-Fusion, we identified 15 novel tumour-specific FTs, with SERPINA1-H19 as the most recurrent fusion event. Comparative expression analysis of fusion partner genes revealed significant downregulation in HCC tumours relative to normal adjacent liver tissues (NAT). We validated the expression levels of the key partner genes with 436 TCGA samples serving as an in silico validation cohort and in wet lab validation cohorts with 42 samples. ALB, APOA2, IGF2, MT2A, SERPINA1, and H19, which are key liver-associated genes, were frequently involved in tumour-specific fusion events suggesting either a loss of tumour suppressor property or gaining a novel function playing a role in hepatocarcinogenesis. Detailed characterization of SERPINA1-H19 identified 16 transcript variants with distinct structural modifications that may impact its functional output. Furthermore, low expression of SERPINA1 and H19 was associated with more aggressive HCC phenotypes. Overall, this study established a comprehensive repository of FTs for the first time, offering valuable insights into their role in HCC and their potential to serve as diagnostic and prognostic biomarkers for HCC.

R2 elements serve as a class of non-long terminal repeat (non-LTR) retrotransposons found in animal genomes that specifically insert into the ribosomal DNA (rDNA) sequences of host genomes. Each R2 element contains a single open reading frame (ORF) encoding a multifunctional protein with nucleic acid-binding, reverse transcriptase, and endonuclease activities, enabling specific genomic integration via a mechanism called target-primed reverse transcription (TPRT). As a classical model for studying retrotransposition mechanisms, R2 elements possess unique biological properties and precise integration capabilities, which have inspired novel genome engineering strategies. In this review, we summarize the components and integration mechanisms of R2 retrotransposons and highlight the recent advances in employing these mobile elements for targeted gene integration. Finally, we present future directions for the utilization of R2 retrotransposons as novel biotechnological tools.

Alternative splicing is a very important mechanism to diversify an organism's transcriptome with minimal increases in genome size. It can modify the function of the finished protein or affect its regulation, e.g. induce nonsense-mediated decay (NMD) to degrade the transcript. Mechanisms that affect alternative splicing are therefore of great interest. It has been shown that splicing can be affected by RNA secondary structures within pre-mRNAs. These structured regions of RNA (strucRNA) would affect their transcript in cis, but only a few such cases are known in plants. In this study, we interrogate plant genomes for cis-regulatory strucRNAs. By applying a comparative-genomics-based approach to 130 plant genomes, we identified 16 strucRNA candidates. Five candidates likely regulate in cis using alternative splicing and NMD. Other predictions might not regulate alternative splicing, including four putative small nucleolar RNAs (snoRNAs). Of our five cis-regulatory strucRNAs that are implicated in alternative splicing control, two are now experimentally validated in follow-up studies. These results stand in contrast to the few previously validated examples. Although we were able to predict some strucRNAs, all motifs had generally modest levels of covariation, which is a pattern of mutations that indicates a conserved secondary structure. With few mutations, comparative-genomics-based approaches to find strucRNAs are less effective. Other approaches of finding regulatory RNAs in plants might thus be needed, and more available genomic or transcriptomic data might improve the quality and quantity of promising candidates.

RNA-focused therapy and diagnostics have been making waves in molecular biology due to the advantages RNA has over DNA; for instance, the ability of RNA to target nearly any genetic component in the cell is a big step in treating disorders. Moreover, RNA-based diagnosis of diseases is only becoming increasingly popular, especially after the COVID-19 pandemic, which brought up the need for cost-effective and efficient diagnosing kits for the vast majority. RNA-based techniques also have close to no risk of genotoxicity and can efficiently target undruggable regions of the cell. RNA treatments have effectively shown the future of the medical industry in the past couple of decades, and they will only be seen to improve. This review paper provides an overview on the different techniques that use RNA-based approaches in the field of diagnostics and therapeutics.

Mesenchymal Stromal/Stem Cells (MSCs) are among the most frequently studied cell types in clinical trials, and their small extracellular vesicles (sEVs) are now being extensively investigated for therapeutic applications. The RNA cargo of MSC-sEVs, particularly miRNAs and mRNAs, is widely believed to be a key therapeutic component of these vesicles. In this review, we critically examine using first principles and peer-reviewed literature, whether MSC- extracellular vesicles (MSC-EVs) can deliver sufficient quantity of functional miRNA or mRNA to target compartments within recipient cells to elicit a pharmacological response. Several RNA sequencing studies reveal that miRNAs are underrepresented in the small RNA population of MSC-sEVs compared to the parent MSCs. Additionally, the majority of miRNAs are mature forms that are not associated with Argonaute (AGO) proteins, essential for their function in RNA-induced silencing complexes (RISCs). Compounding this, cellular uptake of EVs is generally inefficient, with less than 1% being internalized, and only a fraction of these reaching the cytosol. This suggests that EVs may not deliver miRNAs in sufficient quantities to meaningfully interact with AGO proteins, either through canonical or non-canonical pathways, or with other proteins like Toll-like receptors (TLRs). Further, MSC-sEV RNAs are generally small, with sizes less than 500 nucleotides indicating that any mRNA present is likely fragmented as the average mammalian mRNA is approximately 2000 nucleotides, a fact confirmed by RNA sequencing data. Together, these findings challenge the notion that RNA, particularly miRNAs and mRNAs, are primary therapeutic attributes of MSC-sEVs.

Exosomes, nanosized extracellular vesicles (30-150 nm) secreted by various cell types, have emerged as crucial mediators of intercellular communication and promising therapeutic agents. This review highlights the diverse RNA cargo of exosomes derived from human umbilical cord mesenchymal stem cells (HucMSC-Exos), including mRNAs, miRNAs, long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), which regulate gene expression and cellular functions in target cells. The mechanisms of exosome biogenesis, release, and uptake are discussed, with emphasis on their ability to cross biological barriers such as the blood - brain barrier. HucMSC-derived exosomes exhibit therapeutic potential in wound healing, angiogenesis, neuroprotection, immunomodulation, and treatment of diseases like Parkinson's, preeclampsia, and renal or hepatic injury. Specific exosomal miRNAs, such as miR-136, miR-335-5p, and miR-1246, demonstrate targeted effects. Additionally, exosomal RNAs show promise as disease biomarkers. Future directions involve standardization, targeted engineering, RNA profiling, clinical trials, and integration into personalized medicine strategies for regenerative therapy.

tRNA nucleotidyltransferase represents a ubiquitous and essential activity that adds the indispensable CCA triplet to the 3'-end of tRNAs. To fulfill this function, the enzyme contains a set of highly conserved motifs whose coordinated interplay is crucial for the sequence-specific CCA polymerization. In the human enzyme, alterations within these regions have been shown to lead to the manifestation of disease. Recently, we developed an in vivo screening system that allows for the selection and analysis of tRNA nucleotidyltransferase variants by challenging terminal AMP incorporation into tRNA during induced RNase T-catalyzed CCA-decay. Here, we extend this method for screening of full CCA-end repair by utilizing the CCA-trimming activity of exonuclease LCCR4. To demonstrate the combined potential of these two in vivo selection systems, we applied a semi-rational library design to investigate the mode of operation of catalytically important motifs in the human CCA-adding enzyme. This approach revealed unexpected requirements for amino acid composition in two motifs and gives new insights into the mechanism of CCA addition. The data show the potential of these RNase-based screening systems, as they allow the detection of enzyme variations that would not have been identified by a conventional rational approach. Furthermore, the combination of both RNase T and LCCR4 systems can be used to investigate and dissect the effects of pathogenic mutations on C- and A-addition.

The ELAV/Hu family represents a crucial group of RNA-binding proteins predominantly expressed in neurons, playing significant roles in mRNA transcription and translation. These proteins bind to AU-rich elements in transcripts to regulate the expression of cytokines, growth factors, and the development and maintenance of neurons. Elav-like RNA-binding proteins exhibit remarkable molecular weight conservation across different species, highlighting their evolutionary conservation. Although these proteins are widely expressed in the nervous system and other cell types, variations in the DNA sequences of the four Elav proteins contribute to their distinct roles in neurological disorders, cancer, and other Diseases . Elavl1, a ubiquitously expressed family member, is integral to processes such as cell growth, ageing, tumorigenesis, and inflammatory diseases. Elavl2, primarily expressed in the nervous and reproductive systems, is critical for central nervous system and retinal development; its dysregulation has been implicated in neurodevelopmental disorders such as autism. Both Elavl3 and Elavl4 are restricted to the nervous system and are involved in neuronal differentiation and excitability. Elavl3 is essential for cerebellar function and has been associated with epilepsy, while Elavl4 is linked to neurodegenerative diseases, including Parkinson's and Alzheimer's diseases. This paper provides a comprehensive review of the ELAV/Hu family's role in nervous system development, neurological disorders, cancer, and other diseases.

Packaging signals (PSs) of coronaviruses (CoVs) are specific RNA elements recognized by nucleocapsid (N) proteins that direct the selective packaging of genomic RNAs (gRNAs). These signals have been identified in the coding regions of the nonstructural protein 15 (Nsp 15) in CoVs classified under Embecovirus, a subgenus of betacoronaviruses (beta-CoVs). The PSs in other alpha- and beta-CoVs have been proposed to reside in the 5'-proximal regions of gRNAs, supported by comprehensive phylogenetic evidence. However, experimental data remain limited. In this study, we investigated the interactions between Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) 5'-proximal gRNA transcripts and N proteins using electrophoretic mobility shift assays (EMSAs). Our findings revealed that the in vitro synthesized 5'-proximal gRNA transcripts of CoVs can shift from a major conformation to alternative conformations. We also observed that the conformer comprising multiple stem-loops (SLs) is preferentially bound by N proteins. Deletions of the 5'-proximal structural elements of CoV gRNA transcripts, SL1 and SL5a/b/c in particular, were found to promote the formation of alternative conformations. Furthermore, we identified RNA-binding peptides from a pool derived from SARS-CoV N protein. These RNA-interacting peptides were shown to preferentially bind to wild-type SL5a RNA. In addition, our observations of N protein condensate formation in vitro demonstrated that liquid-liquid phase separation (LLPS) of N proteins with CoV-5'-UTR transcripts was influenced by the presence of SL5a/b/c. In conclusion, these results collectively reveal previously uncharacterized binding features between the 5'-proximal transcripts of CoV gRNAs and N proteins.