Cancer-derived exosomal lncRNA SNHG3 promotes the metastasis of colorectal cancer through hnRNPC-mediating RNA stability of ß-catenin.

Metastasis is the leading cause of death in colorectal cancer (CRC) patients. By mediating intercellular communication, exosomes exhibit considerable value in regulating tumor metastasis. Long non-coding RNAs (lncRNAs) are abundant in exosomes and participate in regulating tumor progression. However, it is poorly understood how the cancer-secreted exosomal lncRNAs affect CRC proliferation and metastasis. Here, by analyzing the public databases we identified a lncRNA SNHG3 and demonstrated that SNHG3 was delivered through CRC cells-derived exosomes to promote metastasis in CRC. Mechanistically, exosomal SNHG3 was internalized by CRC cells and afterward upregulated the expression of ß-catenin by facilitating the intranuclear transport of hnRNPC. Consequently, the RNA stability of ß-catenin was enhanced which led to the activation of EMT and metastasis of CRC cells. Our findings expand the oncogenic mechanisms of exosomal SNHG3 and identify it as a diagnostic marker for CRC.

METTL14 decreases FTH1 mRNA stability via m6A methylation to promote sorafenib-induced ferroptosis of cervical cancer.

Cervical cancer (CC) is a prevalent malignancy among women worldwide. This study was designed to investigate the role of METTL14 in sorafenib-induced ferroptosis in CC. METTL14 expression and m6A methylation were determined in CC tissues, followed by analyzes correlating these factors with clinical features. Subsequently, METTL14 was knocked down in CC cell lines, and the effects on cell proliferation, mitochondrial morphology and ferroptosis were assessed using CCK-8, microscopy, and markers associated with ferroptosis, respectively. The regulatory relationship between METTL14 and FTH1 was verified using qRT-PCR and luciferase reporter assays. The functional significance of this interaction was further investigated both in vitro and in vivo by co-transfecting cells with overexpression vectors or shRNAs targeting METTL14 and FTH1 after sorafenib treatment. METTL14 expression and m6A methylation were significantly reduced in CC tissues, and lower METTL14 expression levels were associated with a poorer CC patients' prognosis. Notably, METTL14 expression increased during sorafenib-induced ferroptosis, and METTL14 knockdown attenuated the ferroptotic response induced by sorafenib in CC cells. FTH1 was identified as a direct target of METTL14, with METTL14 overexpression leading to increased m6A methylation of FTH1 mRNA, resulting in reduced stability and expression of FTH1 in CC. Furthermore, FTH1 overexpression or treatment with LY294002 partially counteracted the promotion of sorafenib-induced ferroptosis by METTL14. In vivo xenograft experiments demonstrated that inhibiting METTL14 reduced the anticancer effects of sorafenib, whereas suppression of FTH1 significantly enhanced sorafenib-induced ferroptosis and increased its anticancer efficacy. METTL14 reduces FTH1 mRNA stability through m6A methylation, thereby enhancing sorafenib-induced ferroptosis, which contributes to suppressing CC progression via the PI3K/Akt signaling pathway.

Condensation of RNase L promotes its rapid activation in response to viral infection in mammalian cells.

Oligoadenylate synthetase 3 (OAS3) and ribonuclease L (RNase L) are components of a pathway that combats viral infection in mammals. Upon detection of viral double-stranded RNA (dsRNA), OAS3 synthesizes 2'-5'-oligo(A), which activates the RNase domain of RNase L by promoting the homodimerization and oligomerization of RNase L monomers. Activated RNase L rapidly degrades all cellular mRNAs, shutting off several cellular processes. We sought to understand the molecular mechanisms underlying the rapid activation of RNase L in response to viral infection. Through superresolution microscopy and live-cell imaging, we showed that OAS3 and RNase L concentrated into higher-order cytoplasmic complexes known as dsRNA-induced foci (dRIF) in response to dsRNA or infection with dengue virus, Zika virus, or West Nile virus. The concentration of OAS3 and RNase L at dRIF corresponded with the activation of RNase L-mediated RNA decay. We showed that dimerized/oligomerized RNase L concentrated in a liquid-like shell surrounding a core OAS3-dRIF structure and dynamically exchanged with the cytosol. These data establish that the condensation of dsRNA, OAS3, and RNase L into dRIF is a molecular switch that promotes the rapid activation of RNase L upon detection of dsRNA in mammalian cells.

Overcoming thermostability challenges in mRNA-lipid nanoparticle systems with piperidine-based ionizable lipids.

Lipid nanoparticles (LNPs) have emerged as promising platforms for efficient in vivo mRNA delivery owing to advancements in ionizable lipids. However, maintaining the thermostability of mRNA/LNP systems remains challenging. While the importance of only a small amount of lipid impurities on mRNA inactivation is clear, a fundamental solution has not yet been proposed. In this study, we investigate an approach to limit the generation of aldehyde impurities that react with mRNA nucleosides through the chemical engineering of lipids. We demonstrated that piperidine-based lipids improve the long-term storage stability of mRNA/LNPs at refrigeration temperature as a liquid formulation. High-performance liquid chromatography analysis and additional lipid synthesis revealed that amine moieties of ionizable lipids play a vital role in limiting reactive aldehyde generation, mRNA-lipid adduct formation, and loss of mRNA function during mRNA/LNP storage. These findings highlight the importance of lipid design and help enhance the shelf-life of mRNA/LNP systems.

Collection and transportation of SARS-CoV-2 and influenza A virus diagnostic samples: Optimizing the usage of guanidine-based chaotropic salts for enhanced biosafety and viral genome preservation.

Many virus lysis/transport buffers used in molecular diagnostics, including the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, contain guanidine-based chaotropic salts, primarily guanidine hydrochloride (GuHCl) or guanidine isothiocyanate (GITC). Although the virucidal effects of GuHCl and GITC alone against some enveloped viruses have been established, standardized data on their optimum virucidal concentrations against SARS-CoV-2 and effects on viral RNA stability are scarce. Thus, we aimed to determine the optimum virucidal concentrations of GuHCl and GITC against SARS-CoV-2 compared to influenza A virus (IAV), another enveloped respiratory virus. We also evaluated the effectiveness of viral RNA stabilization at the determined optimum virucidal concentrations under high-temperature conditions (35°C) using virus-specific real-time reverse transcription polymerase chain reaction. Both viruses were potently inactivated by 1.0 M GITC and 2.5 M GuHCl, but the GuHCl concentration for efficient SARS-CoV-2 inactivation was slightly higher than that for IAV inactivation. GITC showed better viral RNA stability than GuHCl at the optimum virucidal concentrations. An increased concentration of GuHCl or GITC increased viral RNA degradation at 35°C. Our findings highlight the need to standardize GuHCl and GITC concentrations in virus lysis/transport buffers and the potential application of these guanidine-based salts alone as virus inactivation solutions in SARS-CoV-2 and IAV molecular diagnostics.

KAP1 stabilizes MYCN mRNA and promotes neuroblastoma tumorigenicity by protecting the RNA m6A reader YTHDC1 protein degradation.

BACKGROUND: Neuroblastoma (NB) patients with amplified MYCN often face a grim prognosis and are resistant to existing therapies, yet MYCN protein is considered undruggable. KAP1 (also named TRIM28) plays a crucial role in multiple biological activities. This study aimed to investigate the relationship between KAP1 and MYCN in NB. METHODS: Transcriptome analyses and luciferase reporter assay identified that KAP1 was a downstream target of MYCN. The effects of KAP1 on cancer cell proliferation and colony formation were explored using the loss-of-function assays in vitro and in vivo. RNA stability detection was used to examine the influence of KAP1 on MYCN expression. The mechanisms of KAP1 to maintain MYCN mRNA stabilization were mainly investigated by mass spectrum, immunoprecipitation, RIP-qPCR, and western blotting. In addition, a xenograft mouse model was used to reveal the antitumor effect of STM2457 on NB. RESULTS: Here we identified KAP1 as a critical regulator of MYCN mRNA stability by protecting the RNA N6-methyladenosine (m6A) reader YTHDC1 protein degradation. KAP1 was highly expressed in clinical MYCN-amplified NB and was upregulated by MYCN. Reciprocally, KAP1 knockdown reduced MYCN mRNA stability and inhibited MYCN-amplified NB progression. Mechanistically, KAP1 regulated the stability of MYCN mRNA in an m6A-dependent manner. KAP1 formed a complex with YTHDC1 and RNA m6A writer METTL3 to regulate m6A-modified MYCN mRNA stability. KAP1 depletion decreased YTHDC1 protein stability and promoted MYCN mRNA degradation. Inhibiting MYCN mRNA m6A modification synergized with chemotherapy to restrain tumor progression in MYCN-amplified NB. CONCLUSIONS: Our research demonstrates that KAP1, transcriptionally activated by MYCN, forms a complex with YTHDC1 and METTL3, which in turn maintain the stabilization of MYCN mRNA in an m6A-dependent manner. Targeting m6A modification by STM2457, a small-molecule inhibitor of METTL3, could downregulate MYCN expression and attenuate tumor proliferation. This finding provides a new alternative putative therapeutic strategy for MYCN-amplified NB.

CYP19A1 Expression Is Controlled by mRNA Stability of the Upstream Transcription Factor AP-2γ in Placental JEG3 Cells.

CYP19A1 encodes aromatase, which converts testosterone to estrogen, and is induced during placental maturation. To elucidate the molecular mechanism underlying this function, histone methylation was analyzed using the placental cytotrophoblast cell line, JEG3. Treatment of JEG3 cells with 3-deazaneplanocin A, an inhibitor of several methyltransferases, resulted in increased CYP19A1 expression, accompanied by removal of the repressive mark H3K27me3 from the CYP19A1 promoter. However, this increase was not observed in cells treated with GSK126, another specific inhibitor for H3K27me3 methylation. Expression of TFAP2C, which encodes AP-2γ, a transcription factor that regulates CYP19A1, was also elevated on 3-deazaneplanocin A treatment. Interestingly, TFAP2C messenger RNA (mRNA) was readily degraded in JEG3 cells but protected from degradation in the presence of 3-deazaneplanocin A. TFAP2C mRNA contained N6-methyladenosines, which were reduced on drug treatment. These observations indicate that the TFAP2C mRNA undergoes adenosine methylation and rapid degradation, whereas 3-deazaneplanocin A suppresses methylation, resulting in an increase in AP-2γ levels. We conclude that the increase in AP-2γ expression via stabilization of the TFAP2C mRNA is likely to underlie the increased CYP19A1 expression.

5'UTR G-quadruplex structure enhances translation in size dependent manner.

Translation initiation in bacteria is frequently regulated by various structures in the 5' untranslated region (5'UTR). Previously, we demonstrated that G-quadruplex (G4) formation in non-template DNA enhances transcription. In this study, we aim to explore how G4 formation in mRNA (RG4) at 5'UTR impacts translation using a T7-based in vitro translation system and in E. coli. We show that RG4 strongly promotes translation efficiency in a size-dependent manner. Additionally, inserting a hairpin upstream of the RG4 further enhances translation efficiency, reaching up to a 12-fold increase. We find that the RG4-dependent effect is not due to increased ribosome affinity, ribosome binding site accessibility, or mRNA stability. We propose a physical barrier model in which bulky structures in 5'UTR biases ribosome movement toward the downstream start codon, thereby increasing the translation output. This study provides biophysical insights into the regulatory role of 5'UTR structures in in vitro and bacterial translation, highlighting their potential applications in tuning gene expression.

IGF2BP3 promotes glutamine metabolism of endometriosis by interacting with UCA1 to enhances the mRNA stability of GLS1.

BACKGROUND: Insulin like growth factor II mRNA binding protein 3 (IGF2BP3) has been implicated in numerous inflammatory and cancerous conditions. However, its precise molecular mechanisms in endometriosis (EMs) remains unclear. The aim of this study is to examine the influence of IGF2BP3 on the occurrence and progression of EMs and to elucidate its underlying molecular mechanism. METHODS: Efects of IGF2BP3 on endometriosis were confrmed in vitro and in vivo. Based on bioinformatics analysis, RNA immunoprecipitation (RIP), RNA pull-down assays and Fluorescent in situ hybridization (FISH) were used to show the association between IGF2BP3 and UCA1. Single-cell spatial transcriptomics analysis shows the expression distribution of glutaminase 1 (GLS1) mRNA in EMs. Study the effect on glutamine metabolism after ectopic endometriotic stromal cells (eESCs) were transfected with Sh-IGF2BP3 and Sh-UCA1 lentivirus. RESULTS: Immunohistochemical staining have revealed that IGF2BP3 was upregulated in ectopic endometriotic lesions (EC) compared to normal endometrial tissues (EN). The proliferation and migration ability of eESCs were greatly reduced by downregulating IGF2BP3. Additionally, IGF2BP3 has been observed to interact with urothelial carcinoma associated 1 (UCA1), leading to increased stability of GLS1 mRNA and subsequently enhancing glutamine metabolism. Results also demonstrated that IGF2BP3 directly interacts with the 3' UTR region of GLS1 mRNA, influencing its expression and stability. Furthermore, UCA1 was able to bind with c-MYC protein, stabilizing c-MYC mRNA and consequently enhancing GLS1 expression through transcriptional promotion. CONCLUSION: These discoveries underscored the critical involvement of IGF2BP3 in the elevation and stability of GLS1 mRNA in the context of glutamine metabolism by interacting with UCA1 in EMs. The implications of our study extended to the identification of possible therapeutic targets for individuals with EMs.

The DEAD-box RNA helicase PfDOZI imposes opposing actions on RNA metabolism in Plasmodium falciparum.

In malaria parasites, the regulation of mRNA translation, storage and degradation during development and life-stage transitions remains largely unknown. Here, we functionally characterized the DEAD-box RNA helicase PfDOZI in P. falciparum. Disruption of pfdozi enhanced asexual proliferation but reduced sexual commitment and impaired gametocyte development. By quantitative transcriptomics, we show that PfDOZI is involved in the regulation of invasion-related genes and sexual stage-specific genes during different developmental stages. PfDOZI predominantly participates in processing body-like mRNPs in schizonts but germ cell granule-like mRNPs in gametocytes to impose opposing actions of degradation and protection on different mRNA targets. We further show the formation of stress granule-like mRNPs during nutritional deprivation, highlighting an essential role of PfDOZI-associated mRNPs in stress response. We demonstrate that PfDOZI participates in distinct mRNPs to maintain mRNA homeostasis in response to life-stage transition and environmental changes by differentially executing post-transcriptional regulation on the target mRNAs.

VHL governs m6A modification and PIK3R3 mRNA stability in clear cell renal cell carcinomas.

N6-Methyladenosine (m6A), a prevalent posttranscriptional modification, plays an important role in cancer progression. Clear cell renal cell carcinoma (ccRCC) is chiefly associated with the loss of the von Hippel-Lindau (VHL) gene, encoding a component of the E3 ubiquitin ligase complex. In this issue of the JCI, Zhang and colleagues unveiled a function of VHL beyond its canonical role as an E3 ubiquitin ligase in regulating hypoxia-inducible factors (HIFs). It also governed m6A modification by orchestrating the assembly of m6A writer proteins METTL3 and METTL14, thereby stabilizing PIK3R3 mRNA. Mechanistically, PIK3R3 contributed to p85 ubiquitination, which restrained PI3K/AKT signaling and consequently impeded ccRCC growth in cell and mouse models. This discovery provides potential treatment targets in VHL-deficient ccRCCs.

RNA m6A modification, signals for degradation or stabilisation?

The RNA modification N6-methyladenosine (m6A) is conserved across eukaryotes, and profoundly influences RNA metabolism, including regulating RNA stability. METTL3 and METTL14, together with several accessory components, form a 'writer' complex catalysing m6A modification. Conversely, FTO and ALKBH5 function as demethylases, rendering m6A dynamic. Key to understanding the functional significance of m6A is its 'reader' proteins, exemplified by YTH-domain-containing proteins (YTHDFs) canonical reader and insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) non-canonical reader. These proteins play a crucial role in determining RNA stability: YTHDFs mainly promote mRNA degradation through different cytoplasmic pathways, whereas IGF2BPs function to maintain mRNA stability. Additionally, YTHDC1 functions within the nucleus to degrade or protect certain m6A-containing RNAs, and other non-canonical readers also contribute to RNA stability regulation. Notably, m6A regulates retrotransposon LINE1 RNA stability and/or transcription via multiple mechanisms. However, conflicting observations underscore the complexities underlying m6A's regulation of RNA stability depending upon the RNA sequence/structure context, developmental stage, and/or cellular environment. Understanding the interplay between m6A and other RNA regulatory elements is pivotal in deciphering the multifaceted roles m6A plays in RNA stability regulation and broader cellular biology.

UPF1 regulates mRNA stability by sensing poorly translated coding sequences.

Post-transcriptional mRNA regulation shapes gene expression, yet how cis-elements and mRNA translation interface to regulate mRNA stability is poorly understood. We find that the strength of translation initiation, upstream open reading frame (uORF) content, codon optimality, AU-rich elements, microRNA binding sites, and open reading frame (ORF) length function combinatorially to regulate mRNA stability. Machine-learning analysis identifies ORF length as the most important conserved feature regulating mRNA decay. We find that Upf1 binds poorly translated and untranslated ORFs, which are associated with a higher decay rate, including mRNAs with uORFs and those with exposed ORFs after stop codons. Our study emphasizes Upf1's converging role in surveilling mRNAs with exposed ORFs that are poorly translated, such as mRNAs with long ORFs, ORF-like 3' UTRs, and mRNAs containing uORFs. We propose that Upf1 regulation of poorly/untranslated ORFs provides a unifying mechanism of surveillance in regulating mRNA stability and homeostasis in an exon-junction complex (EJC)-independent nonsense-mediated decay (NMD) pathway that we term ORF-mediated decay (OMD).

mRNA Decapping Activator Pat1 Is Required for Efficient Yeast Adaptive Transcriptional Responses via the Cell Wall Integrity MAPK Pathway.

Cellular mRNA levels, particularly under stress conditions, can be finely regulated by the coordinated action of transcription and degradation processes. Elements of the 5'-3' mRNA degradation pathway, functionally associated with the exonuclease Xrn1, can bind to nuclear chromatin and modulate gene transcription. Within this group are the so-called decapping activators, including Pat1, Dhh1, and Lsm1. In this work, we have investigated the role of Pat1 in the yeast adaptive transcriptional response to cell wall stress. Thus, we demonstrated that in the absence of Pat1, the transcriptional induction of genes regulated by the Cell Wall Integrity MAPK pathway was significantly affected, with no effect on the stability of these transcripts. Furthermore, under cell wall stress conditions, Pat1 is recruited to Cell Wall Integrity-responsive genes in parallel with the RNA Pol II complex, participating both in pre-initiation complex assembly and transcriptional elongation. Indeed, strains lacking Pat1 showed lower recruitment of the transcription factor Rlm1, less histone H3 displacement at Cell Wall Integrity gene promoters, and impaired recruitment and progression of RNA Pol II. Moreover, Pat1 and the MAPK Slt2 occupied the coding regions interdependently. Our results support the idea that Pat1 and presumably other decay factors behave as transcriptional regulators of Cell Wall Integrity-responsive genes under cell wall stress conditions.

Hfq-Antisense RNA I Binding Regulates RNase E-Dependent RNA Stability and ColE1 Plasmid Copy Number.

The mechanisms and consequences of gene regulation by Hfq on trans-encoded small RNAs (sRNAs) have been well studied and documented. Recent employment of Genomic SELEX to search for Hfq-binding motifs has indicated that Hfq might frequently regulate gene expression controlled by cis-antisense RNAs. Here, we use the classic ColE1 plasmid antisense RNA-based regulation model (i.e., RNA I) to study the role of Hfq in controlling antisense regulatory functions. We show that Hfq exhibits a high binding affinity for RNA I and that binding limits RNase E cleavage, thereby stabilizing RNA I and reducing the plasmid copy number. Full-length RNA I displays a binding affinity for Hfq in the sub-micromolar range. In vivo overexpression of Hfq prolongs RNA I stability and reduces the ColE1 plasmid copy number, whereas deletion of hfq reduces RNA I stability and increases the plasmid copy number. RNA I predominantly binds to the proximal face of Hfq and exhibits competitive ability against a chromosome-borne proximal face-bound sRNA (DsrA) for Hfq binding. Through its strong promoter and high gene dosage features, plasmid-encoded antisense RNA I results in high RNA I expression, so it may antagonize the effects of trans-encoded RNAs in controlling target gene expression.

Turnover of PPP1R15A mRNA encoding GADD34 controls responsiveness and adaptation to cellular stress.

The integrated stress response (ISR) is a key cellular signaling pathway activated by environmental alterations that represses protein synthesis to restore homeostasis. To prevent sustained damage, the ISR is counteracted by the upregulation of growth arrest and DNA damage-inducible 34 (GADD34), a stress-induced regulatory subunit of protein phosphatase 1 that mediates translation reactivation and stress recovery. Here, we uncover a novel ISR regulatory mechanism that post-transcriptionally controls the stability of PPP1R15A mRNA encoding GADD34. We establish that the 3' untranslated region of PPP1R15A mRNA contains an active AU-rich element (ARE) recognized by proteins of the ZFP36 family, promoting its rapid decay under normal conditions and stabilization for efficient expression of GADD34 in response to stress. We identify the tight temporal control of PPP1R15A mRNA turnover as a component of the transient ISR memory, which sets the threshold for cellular responsiveness and mediates adaptation to repeated stress conditions.

PAT mRNA decapping factors are required for proper development in Arabidopsis.

Evolutionarily conserved protein associated with topoisomerase II (PAT1) proteins activate mRNA decay through binding mRNA and recruiting decapping factors to optimize posttranscriptional reprogramming. Here, we generated multiple mutants of pat1, pat1 homolog 1 (path1), and pat1 homolog 2 (path2) and discovered that pat triple mutants exhibit extremely stunted growth and all mutants with pat1 exhibit leaf serration while mutants with pat1 and path1 display short petioles. All three PATs can be found localized to processing bodies and all PATs can target ASYMMETRIC LEAVES 2-LIKE 9 transcripts for decay to finely regulate apical hook and lateral root development. In conclusion, PATs exhibit both specific and redundant functions during different plant growth stages and our observations underpin the selective regulation of the mRNA decay machinery for proper development.

Understanding YTHDF2-mediated mRNA degradation by m6A-BERT-Deg.

N6-methyladenosine (m6A) is the most abundant mRNA modification within mammalian cells, holding pivotal significance in the regulation of mRNA stability, translation and splicing. Furthermore, it plays a critical role in the regulation of RNA degradation by primarily recruiting the YTHDF2 reader protein. However, the selective regulation of mRNA decay of the m6A-methylated mRNA through YTHDF2 binding is poorly understood. To improve our understanding, we developed m6A-BERT-Deg, a BERT model adapted for predicting YTHDF2-mediated degradation of m6A-methylated mRNAs. We meticulously assembled a high-quality training dataset by integrating multiple data sources for the HeLa cell line. To overcome the limitation of small training samples, we employed a pre-training-fine-tuning strategy by first performing a self-supervised pre-training of the model on 427 760 unlabeled m6A site sequences. The test results demonstrated the importance of this pre-training strategy in enabling m6A-BERT-Deg to outperform other benchmark models. We further conducted a comprehensive model interpretation and revealed a surprising finding that the presence of co-factors in proximity to m6A sites may disrupt YTHDF2-mediated mRNA degradation, subsequently enhancing mRNA stability. We also extended our analyses to the HEK293 cell line, shedding light on the context-dependent YTHDF2-mediated mRNA degradation.

Stability indicating ion-pair reversed-phase liquid chromatography method for modified mRNA.

Modified messenger RNA (mRNA) represents a rapidly emerging class of therapeutic drug product. Development of robust stability indicating methods for control of product quality are therefore critical to support successful pharmaceutical development. This paper presents an ion-pair reversed-phase liquid chromatography (IP-RPLC) method to characterise modified mRNA exposed to a wide set of stress-inducing conditions, relevant for pharmaceutical development of an mRNA drug product. The optimised method could be used for separation and analysis of large RNA, sized up to 1000 nucleotides. Column temperature, mobile phase flow rate and ion-pair selection were each studied and optimised. Baseline separations of the model RNA ladder sample were achieved using all examined ion-pairing agents. We established that the optimised method, using 100 mM Triethylamine, enabled the highest resolution separation for the largest fragments in the RNA ladder (750/1000 nucleotides), in addition to the highest overall resolution for the selected modified mRNA compound (eGFP mRNA, 996 nucleotides). The stability indicating power of the method was demonstrated by analysing the modified eGFP mRNA, upon direct exposure to heat, hydrolytic conditions and treatment with ribonucleases. Our results showed that the formed degradation products, which appeared as shorter RNA fragments in front of the main peak, could be well monitored, using the optimised method, and the relative stability of the mRNA under the various stressed conditions could be assessed.

LAGE3 promotes angiogenesis on hepatocellular carcinoma by stabilizing VEGFA mRNA.

RNA modification plays important roles in various physiological and pathological process. LAGE3 is a component of EKC/KEOPS complex, which is probably involved in the formation of a threonylcarbamoyl group on adenosine at position 37 (t(6)A37) in tRNAs, but its exact role in HCC is less studied. Our study reveals that LAGE3 exhibits upregulated expression in HCC compared with normal hepatocellular tissue. High expression of LAGE3 promotes hepatocellular cell proliferation and migration. Further investigations suggest that the increased expression of LAGE3 cloud lead to upregulated VEGFA secretion and angiogenesis in HCC. The mechanistic study reveals LAGE3 is required for the VEGFA mRNA stability. This research may open new avenues for diagnosis and targeted therapy in HCC.