Messenger RNA splicing and degradation are critical for gene expression regulation, the abnormality of which leads to diseases. Previous methods for estimating kinetic rates have limitations, assuming uniform rates across cells. DeepKINET is a deep generative model that estimates splicing and degradation rates at single-cell resolution from scRNA-seq data. DeepKINET outperforms existing methods on simulated and metabolic labeling datasets. Applied to forebrain and breast cancer data, it identifies RNA-binding proteins responsible for kinetic rate diversity. DeepKINET also analyzes the effects of splicing factor mutations on target genes in erythroid lineage cells. DeepKINET effectively reveals cellular heterogeneity in post-transcriptional regulation.

Cancer is a disease that poses a serious threat to human health, the occurrence and development of which involves complex molecular mechanisms. Long non‑coding RNAs (lncRNAs) and RNA‑binding proteins (RBPs) are important regulatory molecules within cells, which have garnered extensive attention in cancer research in recent years. The binding of lncRNAs and RBPs plays a crucial role in the post‑transcriptional regulation of mRNA, affecting the synthesis of proteins related to cancer by regulating the stability of mRNA. This, in turn, regulates the malignant biological behaviors of tumor cells, such as proliferation and metastasis, and serves an important role in therapeutic resistance. The present study reviewed the role of lncRNA‑RBP interactions in the regulation of mRNA stability in various malignant tumors, with a focus on the molecular mechanisms underlying this regulatory interaction. The aim of the present review was to gain a deeper understanding of these molecular mechanisms to provide new strategies and insights for the precise treatment of cancer.

RNase H1 has been acknowledged as an endoribonuclease specializing in the internal degradation of the RNA moiety within RNA-DNA hybrids, and its ribonuclease activity is indispensable in multifaceted aspects of nucleic acid metabolism. However, the molecular mechanism underlying RNase H1-mediated hybrid cleavage remains inadequately elucidated. Herein, using single-molecule approaches, we probe the dynamics of the hybrid cleavage by Saccharomyces cerevisiae RNase H1. Remarkably, a single RNase H1 enzyme displays 3\'-to-5\' exoribonuclease activity. The directional RNA degradation proceeds processively and yet discretely, wherein unwinding approximately 6-bp hybrids as a prerequisite for two consecutive 3-nt RNA excisions limits the overall rate within each catalytic cycle. Moreover, Replication Protein A (RPA) reinforces RNase H1\'s 3\'-to-5\' nucleolytic rate and processivity and stimulates its 5\'-to-3\' exoribonuclease activity. This stimulation is primarily realized through the pre-separation of the hybrids and consequently transfers RNase H1 to a bidirectional exoribonuclease, further potentiating its cleavage efficiency. These findings unveil unprecedented characteristics of an RNase and provide a dynamic view of RPA-enhanced processive hybrid cleavage by RNase H1.

Recent studies have suggested that mRNA internal m7G and its writer protein METTL1 are closely related to cell metabolism and cancer regulation. Here, we identify that IGF2BP family proteins IGF2BP1-3 can preferentially bind internal mRNA m7G. Such interactions, especially IGF2BP3 with m7G, could promote the degradation of m7G target transcripts in cancer cells. IGF2BP3 is more responsive to changes of m7G modification, while IGF2BP1 prefers m6A to stabilize the bound transcripts. We also demonstrate that p53 transcript, TP53, is m7G-modified at its 3\'UTR in cancer cells. In glioblastoma, the methylation level and the half lifetime of the modified transcript could be modulated by tuning IGF2BP3, or by site-specific targeting of m7G through a dCas13b-guided system, resulting in modulation of cancer progression and chemosensitivity.

Upregulation of MYC is a hallmark of cancer, wherein MYC drives oncogenic gene expression and elevates total RNA synthesis across cancer cell transcriptomes. Although this transcriptional anabolism fuels cancer growth and survival, the consequences and metabolic stresses induced by excess cellular RNA are poorly understood. Herein, we discover that RNA degradation and downstream ribonucleotide catabolism is a novel mechanism of MYC-induced cancer cell death. Combining genetics and metabolomics, we find that MYC increases RNA decay through the cytoplasmic exosome, resulting in the accumulation of cytotoxic RNA catabolites and reactive oxygen species. Notably, tumor-derived exosome mutations abrogate MYC-induced cell death, suggesting excess RNA decay may be toxic to human cancers. In agreement, purine salvage acts as a compensatory pathway that mitigates MYC-induced ribonucleotide catabolism, and inhibitors of purine salvage impair MYC+ tumor progression. Together, these data suggest that MYC-induced RNA decay is an oncogenic stress that can be exploited therapeutically. Significance: MYC is the most common oncogenic driver of poor-prognosis cancers but has been recalcitrant to therapeutic inhibition. We discovered a new vulnerability in MYC+ cancer where MYC induces cell death through excess RNA decay. Therapeutics that exacerbate downstream ribonucleotide catabolism provide a therapeutically tractable approach to TNBC (Triple-negative Breast Cancer) and other MYC-driven cancers.

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To study the degradation of lncRNAs in EPMI in rat brain tissue, this study provides a new direction for the estimation of EPMI. LncRNA high-throughput sequencing was performed on the brain tissues of hemorrhagic shock model rats at 0 h and 24 h, and the target lncRNAs were screened. Samples at 0, 1, 3, 6, 12, 18 and 24 h after death were collected, and miRNA-9 and miRNA-125b were used as reference genes. The relative expression levels of lncRNAs at each PMI were detected by RT-qPCR, and a functional model involving lncRNAs and EPMI was established. Samples were collected at 6, 9, 15, and 21 h after death for functional model verification. The expression of several lncRNAs decreased with the prolongation of EPMI, and the mathematical model established by several lncRNA indices exhibited good fit. The verification results of the multi-index joint function model are significantly better than those of the single-index function model, and the established model is more practical. There is a linear relationship between lncRNAs and EPMI, and the multi-index function model is significantly better than the single-index function model, which is important for EPMI inference in forensic pathology practice.

In eukaryotic cells, N6-methyladenosine (m6A) is the most prevalent RNA epigenetic modification that plays crucial roles in multiple biological processes. Nevertheless, the functions and regulatory mechanisms of m6A in phytopathogenic fungi are poorly understood. Here, we showed that CpMTA1, an m6A methyltransferase in Cryphonectria parasitica, plays a crucial role in fungal phenotypic traits, virulence, and stress tolerance. Furthermore, the acid phosphatase gene CpAphA was implicated to be a target of CpMTA1 by integrated analysis of m6A-seq and RNA-seq, as in vivo RIP assay data confirmed that CpMTA1 directly interacts with CpAphA mRNA. Deletion of CpMTA1 drastically lowered the m6A level of CpAphA and reduced its mRNA expression. Moreover, we found that an m6A reader protein CpYTHDF1 recognizes CpAphA mRNA and increases its stability. Typically, the levels of CpAphA mRNA and protein exhibited a positive correlation with CpMTA1 and CpYTHDF1. Importantly, site-specific mutagenesis demonstrated that the m6A sites, A1306 and A1341, of CpAphA mRNA are important for fungal phenotypic traits and virulence in C. parasitica. Together, our findings demonstrate the essential role of the m6A methyltransferase CpMTA1 in C. parasitica, thereby advancing our understanding of fungal gene regulation through m6A modification.

RNA\'s diversity of structures and functions impacts all life forms since primordia. We use calorimetric force spectroscopy to investigate RNA folding landscapes in previously unexplored low-temperature conditions. We find that Watson-Crick RNA hairpins, the most basic secondary structure elements, undergo a glass-like transition below [Formula: see text]C where the heat capacity abruptly changes and the RNA folds into a diversity of misfolded structures. We hypothesize that an altered RNA biochemistry, determined by sequence-independent ribose-water interactions, outweighs sequence-dependent base pairing. The ubiquitous ribose-water interactions lead to universal RNA phase transitions below TG, such as maximum stability at [Formula: see text]C where water density is maximum, and cold denaturation at [Formula: see text]C. RNA cold biochemistry may have a profound impact on RNA function and evolution.

BACKGROUND: In recent years, covalent modifications on RNA nucleotides have emerged as pivotal moieties influencing the structure, function, and regulatory processes of RNA Polymerase II transcripts such as mRNAs and lncRNAs. However, our understanding of their biological roles and whether these roles are conserved across eukaryotes remains limited.
RESULTS: In this study, we leveraged standard polyadenylation-enriched RNA-sequencing data to identify and characterize RNA modifications that introduce base-pairing errors into cDNA reads. Our investigation incorporated data from three Poaceae (Zea mays, Sorghum bicolor, and Setaria italica), as well as publicly available data from a range of stress and genetic contexts in Sorghum and Arabidopsis thaliana. We uncovered a strong enrichment of RNA covalent modifications (RCMs) deposited on a conserved core set of nuclear mRNAs involved in photosynthesis and translation across these species. However, the cohort of modified transcripts changed based on environmental context and developmental program, a pattern that was also conserved across flowering plants. We determined that RCMs can partly explain accession-level differences in drought tolerance in Sorghum, with stress-associated genes receiving a higher level of RCMs in a drought tolerant accession. To address function, we determined that RCMs are significantly enriched near exon junctions within coding regions, suggesting an association with splicing. Intriguingly, we found that these base-pair disrupting RCMs are associated with stable mRNAs, are highly correlated with protein abundance, and thus likely associated with facilitating translation.
CONCLUSIONS: Our data point to a conserved role for RCMs in mRNA stability and translation across the flowering plant lineage.