RNA Biology最新文献

Solid tumours present major treatment obstacles because of their immunosuppressive microenvironment and poor response to traditional chimeric antigen receptor (CAR)-based immunotherapies. Recent advances in cellular engineering have introduced CAR-macrophages derived from induced pluripotent stem cells (CAR-iMacs) as a promising approach to get around these obstacles. CAR-iMacs are designed to attack tumours, but their phenotypic plasticity can cause them to transform into M2-like macrophages in the tumour environment (TME), where they may instead suppress immune responses and promote tumour progression and metastasis. Roquin-1 and Regnase-1 are RNA-binding proteins that act as negative regulators of inflammatory genes that contribute to the phenotypic plasticity of macrophages. This perspective highlights a novel approach to augmenting anti-tumour responses of CAR-iMacs by simultaneously knocking out Roquin-1 and Regnase-1 via CRISPR-Cas9 gene editing. This approach drives a shift from an immunosuppressive M2-like state to an M1 state, promoting sustained pro-inflammatory signalling, boosting phagocytic and cytotoxic capabilities within the tumour microenvironment. Addressing a serious constraint in conventional adoptive cell therapies, this dual-targeting platform could provide a potent and scalable immunotherapeutic treatment for solid malignancies.

We show that a small biotin-binding RNA aptamer that folds into a pseudoknot structure acts as a substrate for bacterial RNase P RNA (RPR) with and without the RNase P C5 protein. Cleavage in the single-stranded region in loop 1 was shown to depend on the presence of a RCCA-motif at the 3' end of the substrate. The nucleobase and the 2'hydroxyl at the position immediately 5' of the cleavage site contribute to both cleavage efficiency and site selection, where C at this position induces significant cleavage at an alternative site, one base upstream of the main cleavage site. The frequencies of cleavage at these two sites and Mg²⁺ binding change upon altering the structural topology in the vicinity of the cleavage site as well as by replacing Mg²⁺ with other divalent metal ions. Modelling studies of RPR in complex with the pseudoknot substrates suggest alternative structural topologies for cleavage at the main and the alternative site and a shift in positioning of Mg²⁺ that activates the H₂O nucleophile. Together, our data are consistent with a model where the organization of the active site structure and positioning of Mg²⁺ is influenced by the identities of residues at and in the vicinity of the site of cleavage.

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.

Liver diseases are a significant global health issue, characterized by elevated levels of disorder and death. The substantial impact of ageing on liver diseases and their prognosis is evident. Multiple processes are involved in the ageing process, which ultimately leads to functional deterioration of this organ. The process of liver ageing not only renders the liver more susceptible to diseases but also compromises the integrity of other organs due to the liver's critical function in metabolism regulation. A growing body of research suggests that long non-coding RNAs (lncRNAs) play a significant role in the majority of pathophysiological pathways. They regulate gene expression through a variety of interactions with microRNAs (miRNAs), messenger RNAs (mRNAs), DNA, or proteins. LncRNAs exert a major influence on the progression of age-related liver diseases through the regulation of cell proliferation, necrosis, apoptosis, senescence, and metabolic reprogramming. A concise overview of the current understanding of lncRNAs and their potential impact on the development of age-related liver diseases will be provided in this mini-review.

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.

We investigated the isomiR profiles of the parasitic worm Fasciola hepatica across three developmental stages: newly excysted juveniles (NEJ), juveniles (JUV), and adults. Our analysis revealed a distinct shift in isomiR distribution during maturation, with NEJs exhibiting a higher abundance and diversity of isomiRs compared to later stages. Notably, isomiRs were often the dominant miRNA form in NEJs, whereas a transition to canonical miRNAs occurred as the parasite matured. This temporal variation suggests that isomiR expression may be linked to the parasite's life cycle. We observed that truncated isomiRs were more prevalent, with uracil additions at the 3'end and adenosine at the 5' end being most common. At least 10% of the miRNA population consisted of 5' end isomiRs, which have the potential to redirect target interactions towards metabolic and developmental pathways. Furthermore, we show that the cleavage sites in F. hepatica primary miRNAs are similar to those found in mammalian cells, and Dicer-mediated cleavage appears to play a significant role in isomiR generation. We believe that the diversification of miRNA sequences through isomiR production is an evolutionary adaptation that enhances the parasite's ability to tune gene expression during infection and development. This regulatory plasticity may facilitate successful infection and long-term persistence within diverse mammalian hosts. Understanding the roles of isomiRs in parasitic worms could provide new insights into parasite biology and identify potential targets for controlling parasitic infections.

Piwi-interacting small RNAs (piRNA) play a key role in controlling the activity of transposable elements (TEs) in the animal germline. In diverse arthropod species, including the pathogen vectors mosquitoes, the piRNA pathway is also active in nongonadal somatic tissues, where its targets and functions are less clear. Here, we studied the features of small RNA production in head and thorax tissues of an uninfected laboratory strain of Anopheles coluzzii focusing on the 24-32-nt-long RNAs. Small RNAs derived from repetitive elements constitute a minor fraction while most small RNAs process from long noncoding RNAs (lncRNAs) and protein-coding gene mRNAs. The majority of small RNAs derived from repetitive elements and lncRNAs exhibited typical piRNAs features. By contrast, majority of protein-coding gene-derived 24-32 nt small RNAs lack the hallmarks of piRNAs and have signatures of nontemplated 3' end tailing. Most of the atypical small RNAs exhibit female-biased expression and originate from mitochondrial and nuclear genes involved in energy metabolism. We also identified atypical genic small RNAs in Anopheles gambiae somatic tissues, which further validates the noncanonical mechanism of their production. We discuss a novel mechanism of small RNA production in mosquito somatic tissues and the possible functional significance of genic small RNAs.

Super enhancers are important regulators of gene expression that often overlap with protein-coding genes. However, it is unclear whether the overlapping protein-coding genes and the RNA derived from them contribute to enhancer activity. Using an erythroid-specific super enhancer that overlaps the Cpox gene as a model, Cpox pre-mRNA is found to have a non-coding function in regulating neighbouring protein-coding genes, eRNA expression and TAD interactions. Depletion of Cpox pre-mRNA leads to accumulation of H3K27me3 and release of p300 from the Cpox locus, activating an intra-TAD enhancer and gene expression. Additionally, a head-to-tail interaction between the TAD boundary genes Cpox and Dcbld2 is identified, facilitated by a novel type of repressive loop anchored by p300 and PRC2/H3K27me3. These results uncover a regulatory role for pre-mRNA transcribed within a super enhancer context and provide insight into head-to-tail inter-gene interaction in the regulation of gene expression and oncogene activation.