RÉSUMÉ
A recently reported Schizophrenia-associated genetic variant in the 3'UTR of the human furin gene, a homolog of C. elegans kpc-1, highlights an important role of the furin 3'UTR in neuronal development. We isolate three kpc-1 mutants that display abnormal dendrite arborization in PVD neurons and defective male mating behaviors. We show that the kpc-1 3'UTR participates in dendrite branching and self-avoidance. The kpc-1 3'UTR facilitates mRNA localization to branching points and contact points between sibling dendrites and promotes translation efficiency. A predicted secondary structural motif in the kpc-1 3'UTR is required for dendrite self-avoidance. Animals with over-expression of DMA-1, a PVD dendrite receptor, exhibit similar dendrite branching and self-avoidance defects that are suppressed with kpc-1 over-expression. Our results support a model in which KPC-1 proteins are synthesized at branching points and contact points to locally down-regulate DMA-1 receptors to promote dendrite branching and self-avoidance of a mechanosensory neuron important for male courtship.
Sujet(s)
Régions 3' non traduites , Protéines de Caenorhabditis elegans , Caenorhabditis elegans , Parade nuptiale , Dendrites , ARN messager , Animaux , Protéines de Caenorhabditis elegans/génétique , Protéines de Caenorhabditis elegans/métabolisme , Mâle , Dendrites/métabolisme , Dendrites/génétique , Caenorhabditis elegans/génétique , Caenorhabditis elegans/métabolisme , Régions 3' non traduites/génétique , ARN messager/génétique , ARN messager/métabolisme , Biosynthèse des protéines , Comportement sexuel chez les animaux/physiologie , Humains , Mutation , Protéines membranairesRÉSUMÉ
Periodontitis, with its persistent nature, causes significant distress for most sufferers. Current treatments, such as mechanical cleaning and surgery, often fail to fully address the underlying overactivation of fibroblasts that drives this degradation. Targeting the post-transcriptional regulation of fibroblasts, particularly at the 3'-untranslated regions (3'UTR) of pathogenic genes, offers a therapeutic strategy for periodontitis. Herein, we developed a DNA nanorobot for this purpose. This system uses a dynamic DNA nanoframework to incorporate therapeutic microRNAs through molecular recognition and covalent bonds, facilitated by DNA monomers modified with disulfide bonds. The assembled-DNA nanoframework is encapsulated in a cell membrane embedded with a fibroblast-targeting peptide. By analyzing the 3'UTR regions of pathogenic fibroblast genes FOSB and JUND, we identified the therapeutic microRNA as miR-1-3p and integrated it into this system. As expected, the DNA nanorobot delivered the internal components to fibroblasts by the targeting peptide and outer membrane that responsively releases miR-1-3p under intracellular glutathione. It resulted in a precise reduction of mRNA and suppression of protein function in pathogenic genes, effectively reprogramming fibroblast behavior. Our results confirm that this approach not only mitigates the inflammation but also promotes tissue regeneration in periodontal models, offering a promising therapeutic avenue for periodontitis.
Sujet(s)
Régions 3' non traduites , ADN , Fibroblastes , microARN , Parodontite , Parodontite/génétique , Parodontite/anatomopathologie , Fibroblastes/métabolisme , Régions 3' non traduites/génétique , ADN/composition chimique , ADN/génétique , microARN/génétique , microARN/métabolisme , Humains , Animaux , SourisRÉSUMÉ
Atherosclerotic plaque formation is largely attributed to the impaired efferocytosis, which is known to be associated with the pathologic upregulation of cluster of differentiation 47 (CD47), a key antiphagocytic molecule. By gene expression omnibus (GEO) datasets analysis, we identified that four miRNAs are aberrantly downregulated in atherosclerosis, coronary artery disease, and obesity. Of them, hsa-miR-299-3p (miR-299-3p) was predicted to target the 3'UTR of human CD47 mRNA by bioinformatics analysis. Further, we demonstrated that miR-299-3p negatively regulates CD47 expression by binding to the target sequence "CCCACAU" in the 3'UTR of CD47 mRNA through luciferase reporter assay and site-directed mutagenesis. Additionally, we found that miR-299-3p was downregulated by ~32% in foam cells in response to oxidized low-density lipoprotein (ox-LDL) stimulation, thus upregulating CD47 and contributing to the impaired efferocytosis. Whereas, restoration of miR-299-3p reversed the ox-LDL-induced upregulation of CD47, thereby facilitating efferocytosis. In high-fat diet (HFD) fed ApoE-/- mice, we discovered that miR-299-3p was downregulated thus leading to upregulation of CD47 in abdominal aorta. Conversely, miR-299-3p restoration potently suppressed HFD-induced upregulation of CD47 and promoted phagocytosis of foam cells by macrophages in atherosclerotic plaques, thereby reducing necrotic core, increasing plaque stability, and mitigating atherosclerosis. Conclusively, we identify miR-299-3p as a negative regulator of CD47, and reveal a molecular mechanism whereby the ox-LDL-induced downregulation of miR-299-3p leads to the upregulation of CD47 in foam cells thus contributing to the impaired efferocytosis in atherosclerosis, and propose miR-299-3p can potentially serve as an inhibitor of CD47 to promote efferocytosis and ameliorate atherosclerosis.
Sujet(s)
Athérosclérose , Antigènes CD47 , , microARN , Animaux , Humains , Souris , Régions 3' non traduites , Athérosclérose/métabolisme , Athérosclérose/génétique , Athérosclérose/anatomopathologie , Antigènes CD47/métabolisme , Antigènes CD47/génétique , Alimentation riche en graisse/effets indésirables , Cellules spumeuses/métabolisme , Cellules spumeuses/anatomopathologie , Lipoprotéines LDL/métabolisme , Souris de lignée C57BL , microARN/génétique , microARN/métabolismeRÉSUMÉ
BACKGROUND: Colon cancer remains a major health concern worldwide, with genetic factors playing a crucial role in its development. Toll-like receptors (TLRs) has been implicated in various cancers, but their role in colon cancer is not well understood. This study aims to identify functional polymorphisms in the promoter and 3'UTR regions of TLRs and evaluate their association with colon cancer susceptibility. METHODS: We conducted a case-control study involving 410 colon cancer patients and 410 healthy controls from the Chinese population. Genotyping of polymorphisms in TLR3, TLR4, TLR5 and TLR7 was performed using PCR-RFLP and TaqMan MGB probes. Using logistic regression analysis, we evaluated the association of TLRs polymorphisms and the susceptibility to colon cancer. To understand the biological implications of the TLR4 rs1927914 polymorphism, we conducted functional assays, including luciferase reporter assay and electrophoretic mobility shift assay (EMSA). RESULTS: Our results demonstrated that the G-allele of the TLR4 rs1927914 polymorphism is significantly associated with a decreased risk of colon cancer (OR = 0.68, 95%CI = 0.50-0.91). Stratified analysis showed that TLR4 rs1927914 AG or GG genotype contributed to a decreased risk of colon cancer among younger individuals (OR = 0.52, 95%CI = 0.34-0.81), males (OR = 0.58, 95%CI = 0.38-0.87), non-smokers (OR = 0.58, 95%CI = 0.41-0.83) and non-drinker with OR (95%CI) of 0.66 (0.46-0.93). Functional assays demonstrated that in HCT116 and LOVO colon cancer cells, the luciferase activity driven by the TLR4 promoter with the rs1927914A allele was 5.43 and 2.07 times higher, respectively, compared to that driven by the promoter containing the rs1927914G allele. Electrophoretic mobility shift assay (EMSA) results indicated that the rs1927914G allele enhanced transcription factor binding. Using the transcription factor prediction tool, we found that the G allele facilitates binding of the repressive transcription factor Oct1, while the A allele does not. CONCLUSION: The TLR4 rs1927914 polymorphism influence the susceptibility to colon cancer, with the G allele offering a protective effect through modulation of gene expression. These insights enhance our understanding of the genetic determinants of colon cancer risk and highlight TLR4 as a promising target for cancer prevention strategies.
Sujet(s)
Tumeurs du côlon , Prédisposition génétique à une maladie , Polymorphisme de nucléotide simple , Récepteur de type Toll-4 , Humains , Récepteur de type Toll-4/génétique , Mâle , Femelle , Tumeurs du côlon/génétique , Adulte d'âge moyen , Études cas-témoins , Génotype , Sujet âgé , Régions promotrices (génétique) , Allèles , Études d'associations génétiques , Régions 3' non traduites/génétique , Adulte , Asiatiques/génétique , Facteurs de risqueRÉSUMÉ
BACKGROUND: JUB1, a NAC domain containing hydrogen peroxide-induced transcription factor, plays a critical role in plant immunity. Little is known about how JUB1 responds to leaf rust disease in wheat. Recent discoveries in genomics have also unveiled a multitude of sORFs often assumed to be non-functional, to argue for the necessity of including them as potential regulatory players of translation. However, whether methylation on sORFs spanning the 3'UTR of regulatory genes like JUB1 modulate gene expression, remains unclear. METHODS AND RESULTS: In this study, we identified the methylation states of two sORFs in 3'UTR of a homologous gene of JUB1 in wheat, TaJUB1-L, at cytosine residues in CpG, CHH and CHG sites at different time points of disease progression in two near-isogenic lines of wheat (HD2329), with and without Lr24 gene during leaf rust pathogenesis. Here, we report a significant demethylation of the CpG dinucleotides occurring in the sORFs of the 3'UTR in the resistant isolines after 24 h post-infection. Also, the up-regulated gene expression observed through RT-qPCR was directly proportional to the demethylation of the CpG sites in the sORFs. CONCLUSIONS: Our findings indicate that TaJUB1-L might be a positive regulator in providing tolerance during leaf rust pathogenesis and cytosine methylation at 3'UTR might act as a switch for its expression control. These results enrich the potential benefit of conventional methylation assay techniques for unraveling the unexplored enigma in epigenetics during plant-pathogen interaction in a cost-effective and confidentially conclusive manner.
Sujet(s)
Régions 3' non traduites , Méthylation de l'ADN , Régulation de l'expression des gènes végétaux , Maladies des plantes , Protéines végétales , Facteurs de transcription , Triticum , Triticum/microbiologie , Triticum/génétique , Maladies des plantes/microbiologie , Maladies des plantes/génétique , Régions 3' non traduites/génétique , Méthylation de l'ADN/génétique , Régulation de l'expression des gènes végétaux/génétique , Protéines végétales/génétique , Protéines végétales/métabolisme , Facteurs de transcription/génétique , Facteurs de transcription/métabolisme , Basidiomycota/pathogénicité , Basidiomycota/génétique , Feuilles de plante/microbiologie , Feuilles de plante/génétique , Résistance à la maladie/génétique , 5-Méthyl-cytosine/métabolismeRÉSUMÉ
Exposure to blue light can lead to retinal degeneration, causing adverse effects on eye health. Although the loss of retinal cells due to blue light exposure has been observed, the precise molecular mechanisms underlying this process remain poorly understood. In this study, we investigate the role of alpha-crystallin A (CRYAA) in neuro-retinal degeneration and their regulation by blue light. We observed significant apoptotic cell death in both the retina of rats and the cultured neuro-retinal cells. The expressions of Cryaa mRNA and protein were significantly downregulated in the retina exposed to blue light. We identified that miR-325-3p reduces Cryaa mRNA and protein by binding to its 3'-untranslated region. Upregulation of miR-325-3p destabilized Cryaa mRNA and suppresses CRYAA, whereas downregulation of miR-325-3p increased both expressions. Blue light-induced neuro-retinal cell death was alleviated by CRYAA overexpression. These results highlight the critical role of Cryaa mRNA and miR-325-3p molecular axis in blue light-induced retinal degeneration. Consequently, targeting CRYAA and miR-325-3p presents a potential strategy for protecting against blue light-induced retinal degeneration.
Sujet(s)
Lumière , microARN , Rétine , Animaux , microARN/génétique , microARN/métabolisme , Lumière/effets indésirables , Rats , Rétine/métabolisme , Rétine/effets des radiations , Chaine A de la cristalline alpha/métabolisme , Chaine A de la cristalline alpha/génétique , Dégénérescence de la rétine/métabolisme , Dégénérescence de la rétine/génétique , Dégénérescence de la rétine/anatomopathologie , Dégénérescence de la rétine/étiologie , Apoptose/effets des radiations , Rat Sprague-Dawley , Mâle , Régulation négative , Régions 3' non traduites ,RÉSUMÉ
As an important mechanism in the post-transcriptional regulation of eukaryotic gene expression, alternative polyadenylation (APA) plays a key role in biological processes such as cell proliferation and differentiation. However, the role and dynamic pattern of APA during Litopenaeus vannamei metamorphosis are poorly understood. Here, RNA-seq data covering from the embryo to the maturation (16 time points) of L. vannamei were utilized. We identified 247 differentially expressed APA events between early and adult stages, and through fuzzy mean clustering analysis, we discovered five dynamic APA patterns. Among them, the gradual elongation of the 3'UTR is the major APA pattern that changes over time, and its genes are enriched in the pathways of protein and energy metabolism. Finally, we constructed mRNA-miRNA and PPI networks and detected several central miRNAs that may regulate L. vannamei development. Our results revealed the complex APA mechanisms in L. vannamei metamorphosis, shedding new light on post-transcriptional regulation of crustacean metamorphosis.
Sujet(s)
Régions 3' non traduites , Régulation de l'expression des gènes au cours du développement , Métamorphose biologique , Penaeidae , Polyadénylation , Animaux , Métamorphose biologique/génétique , Penaeidae/génétique , Penaeidae/croissance et développement , Polyadénylation/génétique , Régions 3' non traduites/génétique , microARN/génétique , ARN messager/génétique , ARN messager/métabolismeRÉSUMÉ
Extracellular vesicles (EVs) are garnering attention as a safe and efficient biomolecule delivery system. EVs intrinsically play a crucial role in intercellular communication and pathophysiology by transporting functionally active DNA molecules. The internalized DNA pleiotropically affects the recipient cells. Considering these salient features, an intentional incorporation of specific DNA gene cassettes into EVs and their subsequent delivery to the target cells has potential applications in genetic engineering. Moreover, efficient ways to insert the DNA into EVs during their biogenesis is valuable. Our current research is a step in the development of this technology. As such, cancer cells are known to secrete exosomes containing increased amounts of double-stranded DNA than normal cells. The clonal analysis in our previously published data revealed that exosomes released from various cancer cells contained a significantly larger population of NANOGP8 DNA with a 22-base pair insertion in the 3'-untranslated region (UTR) compared to those secreted by normal cells. This finding led us to hypothesize that the 22-base pair insertion may act as a signal to facilitate the incorporation of NANOGP8 DNA into the exosomes. To test this hypothesis, we compared the EV localization of an Enhanced Green Fluorescent Protein (EGFP) gene fused with the NANOGP8 3'-UTR, with and without the 22-base pair insertion. The quantitative PCR analysis showed a significantly higher EGFP DNA accumulation in exosomes released from cells transfected with the gene cassette containing the 3'-UTR with the 22-base pair insertion. The discovery of a DNA localization signal in exosomal DNA's 3'-UTR could pave the way for the development of an EV-based DNA delivery system. This technology will open new possibilities in genetic engineering and innovative therapies using nucleic acid medicine.
Sujet(s)
Régions 3' non traduites , Exosomes , Vésicules extracellulaires , Exosomes/métabolisme , Exosomes/génétique , Humains , Vésicules extracellulaires/métabolisme , Vésicules extracellulaires/génétique , ADN/génétique , ADN/métabolisme , Protéines à fluorescence verte/métabolisme , Protéines à fluorescence verte/génétique , Lignée cellulaire tumoraleRÉSUMÉ
miRNA has emerged as a crucial regulator in various of pathological and physiological processes, yet its precise mechanism of action the detailed mechanism of their action in Head and neck squamous cell carcinoma (HNSCC) remains incompletely understood. This study sheds light on the role of mi-151-5p, revealing its significantly elevated expression in tumor cells, which notably enhances the invasion and migration of HNSCC cells. This effect is achieved through directly targeting LY6/PLAUR Domain Containing 3 (LYPD3) by miR-151-5p, involving complementary binding to the 3'-untranslated regions (3'-UTR) in the mRNA of LYPD3. Consequently, this interaction accelerates the metastasis of HNSCC. Notably, clinical observations indicate a correlation between high expression of miR-151-5p and low levels of LYPD3 in clinical settings are correlated with poor prognosis of HNSCC patients. Furthermore, our investigation demonstrates that glycosylation of LYPD3 modulates its subcellular localization and reinforces its role in suppressing HNSCC metastasis. Additionally, we uncover a potential regulatory mechanism involving the facilitation of miR-151-5p maturation and accumulation through N6-methyladenosine (m6A) modification. This process is orchestrated by methyltransferase-like 3 (METTL3) and mediated by a newly identified reader, heterogeneous nuclear ribonucleoprotein U (hnRNP U). These findings collectively underscore the significance of the METTL3/miR-151-5p/LYPD3 axis serves as a prominent driver in the malignant progression of HNSCC.
Sujet(s)
Adénosine , Régulation de l'expression des gènes tumoraux , Tumeurs de la tête et du cou , microARN , Carcinome épidermoïde de la tête et du cou , Humains , Régions 3' non traduites/génétique , Adénosine/analogues et dérivés , Adénosine/métabolisme , Lignée cellulaire tumorale , Mouvement cellulaire/génétique , Tumeurs de la tête et du cou/génétique , Tumeurs de la tête et du cou/anatomopathologie , Tumeurs de la tête et du cou/métabolisme , Methyltransferases/génétique , Methyltransferases/métabolisme , microARN/génétique , microARN/métabolisme , Carcinome épidermoïde de la tête et du cou/génétique , Carcinome épidermoïde de la tête et du cou/anatomopathologie , Carcinome épidermoïde de la tête et du cou/métabolismeRÉSUMÉ
Hepatocellular carcinoma (HCC) is one of the most malignant tumors in the world. Inadequate efficacy of 5-fluorouracil (5-FU) on HCC could be related to low expression of human organic anion transporter 2 (OAT2). However, the knowledge of downregulation of OAT2 in HCC remains limited. We explored the underlying mechanism focusing on protein expression regulation and attempted to design a strategy to sensitize HCC cells to 5-FU. In this study, we revealed that the 1 bp to 300 bp region of OAT2 mRNA 3' untranslated region (UTR) reduced its protein expression and uptake activity in Li-7 and PLC/PRF/5 cells. Mechanistically, it was demonstrated that staphylococcal nuclease and Tudor domain containing 1 (SND1) bound at the 1 bp to 300 bp region of OAT2 mRNA 3' UTR, leading to a decrease in OAT2 protein expression. Enrichment analysis results indicated reduction of OAT2 might be mediated by translational inhibition. Furthermore, the knockdown of SND1 upregulated OAT2 protein expression and uptake activity. Based on this, decreasing SND1 expression enhanced 5-FU-caused G1/S phase arrest in Li-7 and PLC/PRF/5 cells, resulting in suppression of cell proliferation. Additionally, the knockdown of SND1 augmented the inhibitory effect of 5-FU on PLC/PRF/5 xenograft tumor growth in vivo by increasing OAT2 protein expression and accumulation of 5-FU in the tumor. Collectively, a combination of inhibition of SND1 with 5-FU might be a potential strategy to sensitize HCC cells to 5-FU from the perspective of restoring OAT2 protein level. SIGNIFICANCE STATEMENT: We investigated the regulatory mechanism of OAT2 protein expression in HCC cells and designed a strategy to sensitize them to 5-FU (OAT2 substrate) via restoring OAT2 protein level. It found that SND1, an RNA binding protein, regulated OAT2 protein expression by interacting with OAT2 mRNA 3' UTR 1-300 bp region. Through decreasing SND1, the antitumor effect of 5-FU on HCC was enhanced in vitro and in vivo, indicating that SND1 could be a potential target for sensitizing HCC cells to 5-FU.
Sujet(s)
Carcinome hépatocellulaire , Endonucleases , Fluorouracil , Tumeurs du foie , Humains , Fluorouracil/pharmacologie , Carcinome hépatocellulaire/traitement médicamenteux , Carcinome hépatocellulaire/métabolisme , Carcinome hépatocellulaire/génétique , Carcinome hépatocellulaire/anatomopathologie , Tumeurs du foie/traitement médicamenteux , Tumeurs du foie/génétique , Tumeurs du foie/métabolisme , Tumeurs du foie/anatomopathologie , Animaux , Endonucleases/génétique , Endonucleases/métabolisme , Lignée cellulaire tumorale , Souris , Souris nude , Régulation de l'expression des gènes tumoraux/effets des médicaments et des substances chimiques , Tests d'activité antitumorale sur modèle de xénogreffe/méthodes , Antimétabolites antinéoplasiques/pharmacologie , Souris de lignée BALB C , Transporteurs d'anions organiques sodium-indépendants/métabolisme , Transporteurs d'anions organiques sodium-indépendants/génétique , Prolifération cellulaire/effets des médicaments et des substances chimiques , Régions 3' non traduites/génétique , MâleRÉSUMÉ
Many viruses have evolved structured RNA elements that can influence transcript abundance and translational efficiency, and help evade host immune factors by hijacking cellular machinery during replication. Here, we evaluated the functional impact of sub-genomic flaviviral RNAs (sfRNAs) known to stall exoribonuclease activity, by incorporating these elements into recombinant adeno-associated viral (AAV) genome cassettes. Specifically, sfRNAs from Dengue, Zika, Japanese Encephalitis, Yellow Fever, Murray Valley Encephalitis, and West Nile viruses increased transcript stability and transgene expression compared to a conventional woodchuck hepatitis virus element (WPRE). Further dissection of engineered transcripts revealed that sfRNA elements (i) require incorporation in cis within the 3' untranslated region (UTR) of AAV genomes, (ii) require minimal dumbbell structures to exert the observed effects, and (iii) can stabilize AAV transcripts independent of 5'-3' exoribonuclease 1 (XRN1)-mediated decay. Additionally, preliminary in vivo assessment of AAV vectors bearing sfRNA elements in mice achieved increased transcript abundance and expression in cardiac tissue. Leveraging the functional versatility of engineered viral RNA elements may help improve the potency of AAV vector-based gene therapies. IMPORTANCE: Viral RNA elements can hijack host cell machinery to control stability of transcripts and consequently, infection. Studies that help better understand such viral elements can provide insights into antiviral strategies and also potentially leverage these features for therapeutic applications. In this study, by incorporating structured flaviviral RNA elements into recombinant adeno-associated viral (AAV) vector genomes, we show improved AAV transcript stability and transgene expression can be achieved, with implications for gene transfer.
Sujet(s)
Dependovirus , Vecteurs génétiques , ARN viral , Dependovirus/génétique , Animaux , ARN viral/génétique , ARN viral/métabolisme , Vecteurs génétiques/génétique , Souris , Humains , Stabilité de l'ARN , Flaviviridae/génétique , Transgènes , Cellules HEK293 , Génome viral , Régions 3' non traduites/génétique , Exoribonucleases/métabolisme , Exoribonucleases/génétiqueRÉSUMÉ
The F11 receptor (F11R) gene encoding junctional adhesion molecule A has been associated with gastric cancer (GC) and colorectal cancer (CRC), in which its role and regulation remain to be further elucidated. Recently F11R was also identified as a potential target of adenosine-to-inosine (A-to-I) mediated by the adenosine deaminases acting on RNA (ADARs). Herein, using RNA-Seq and experimental validation, our current study revealed an F11R RNA trinucleotide over-edited by ADAR, with its regulation of gene expression and clinical significance in four GC and three CRC cohorts. Our results found an over-edited AAA trinucleotide in an AluSg located in the F11R 3'-untranslated region (3'-UTR), which showed editing levels correlated with elevated ADAR expression across all GC and CRC cohorts in our study. Overexpression and knockdown of ADAR in GC and CRC cells, followed by RNA-Seq and Sanger sequencing, confirmed the ADAR-mediated F11R 3'-UTR trinucleotide editing, which potentially disrupted an RBM45 binding site identified by crosslinking immunoprecipitation sequencing (CLIP-seq) and regulated F11R expression in luciferase reporter assays. Moreover, the F11R trinucleotide editing showed promising predictive performance for diagnosing GC and CRC across GC and CRC cohorts. Our findings thus highlight both the potential biological and clinical significance of an ADAR-edited F11R trinucleotide in GC and CRC, providing new insights into its application as a novel diagnostic biomarker for both cancers.
Sujet(s)
Adenosine deaminase , Tumeurs colorectales , Régulation de l'expression des gènes tumoraux , Édition des ARN , Protéines de liaison à l'ARN , Tumeurs de l'estomac , Humains , Adenosine deaminase/génétique , Adenosine deaminase/métabolisme , Tumeurs colorectales/génétique , Tumeurs colorectales/diagnostic , Tumeurs colorectales/métabolisme , Tumeurs de l'estomac/génétique , Tumeurs de l'estomac/diagnostic , Tumeurs de l'estomac/métabolisme , Protéines de liaison à l'ARN/métabolisme , Protéines de liaison à l'ARN/génétique , Études de cohortes , Régions 3' non traduites/génétique , Lignée cellulaire tumorale , Marqueurs biologiques tumoraux/génétique , Marqueurs biologiques tumoraux/métabolisme , Mâle , FemelleRÉSUMÉ
The kinetoplastid parasite, Trypanosoma brucei, undergoes a complex life cycle entailing slender and stumpy bloodstream forms in mammals and procyclic and metacyclic forms (MFs) in tsetse fly hosts. The numerous gene regulatory events that underlie T. brucei differentiation between hosts, as well as between active and quiescent stages within each host, take place in the near absence of transcriptional control. Rather, differentiation is controlled by RNA-binding proteins (RBPs) that associate with mRNA 3' untranslated regions (3'UTRs) to impact RNA stability and translational efficiency. DRBD18 is a multifunctional T. brucei RBP, shown to impact mRNA stability, translation, export, and processing. Here, we use single-cell RNAseq to characterize transcriptomic changes in cell populations that arise upon DRBD18 depletion, as well as to visualize transcriptome-wide alterations to 3'UTR length. We show that in procyclic insect stages, DRBD18 represses expression of stumpy bloodstream form and MF transcripts. Additionally, DRBD18 regulates the 3'UTR lengths of over 1,500 transcripts, typically promoting the use of distal polyadenylation sites, and thus the inclusion of 3'UTR regulatory elements. Remarkably, comparison of polyadenylation patterns in DRBD18 knockdowns with polyadenylation patterns in stumpy bloodstream forms shows numerous similarities, revealing a role for poly(A) site selection in developmental gene regulation, and indicating that DRBD18 controls this process for a set of transcripts. RNA immunoprecipitation supports a direct role for DRBD18 in poly(A) site selection. This report highlights the importance of alternative polyadenylation in T. brucei developmental control and identifies a critical RBP in this process.
Sujet(s)
Régions 3' non traduites , Étapes du cycle de vie , Protéines de protozoaire , Protéines de liaison à l'ARN , Trypanosoma brucei brucei , Trypanosoma brucei brucei/génétique , Trypanosoma brucei brucei/métabolisme , Protéines de protozoaire/métabolisme , Protéines de protozoaire/génétique , Protéines de liaison à l'ARN/métabolisme , Protéines de liaison à l'ARN/génétique , Étapes du cycle de vie/génétique , Régions 3' non traduites/génétique , Animaux , Transcriptome , ARN messager/génétique , ARN messager/métabolisme , Poly A/métabolisme , Poly A/génétique , PolyadénylationRÉSUMÉ
Obesity is characterized by dysregulated adipogenesis that leads to increased number and/or size of adipocytes. Understanding the molecular mechanisms governing adipogenesis is therefore key to designing therapeutic interventions against obesity. In our study, we analyzed 3'-end sequencing data that we generated from human preadipocytes and adipocytes, as well as previously published RNA-seq datasets, to elucidate mechanisms of regulation via long non-coding RNA (lncRNA), alternative splicing (AS) and alternative polyadenylation (APA). We discovered lncRNAs that have not been previously characterized but may be key regulators of white adipogenesis. We also detected 100 AS events and, using motif enrichment analysis, identified RNA binding proteins (RBPs) that could mediate exon skipping-the most prevalent AS event. In addition, we show that usage of alternative poly(A) sites in introns or 3'-UTRs of key adipogenesis genes leads to isoform diversity, which can have significant biological consequences on differentiation efficiency. We also identified RBPs that may modulate APA and defined how 3'-UTR APA can regulate gene expression through gain or loss of specific microRNA binding sites. Taken together, our bioinformatics-based analysis reveals potential therapeutic avenues for obesity through manipulation of lncRNA levels and the profile of mRNA isoforms via alternative splicing and polyadenylation.
Sujet(s)
Adipogenèse , Épissage alternatif , Analyse de profil d'expression de gènes , Polyadénylation , ARN long non codant , ARN long non codant/génétique , ARN long non codant/métabolisme , Adipogenèse/génétique , Humains , Adipocytes/métabolisme , Adipocytes/cytologie , Régions 3' non traduites , microARN/génétique , microARN/métabolisme , Transcriptome , Régulation de l'expression des gènes , Protéines de liaison à l'ARN/métabolisme , Protéines de liaison à l'ARN/génétique , Biologie informatique/méthodesRÉSUMÉ
Biomolecular condensates organize biochemical processes at the subcellular level and can provide spatiotemporal regulation within a cell. Among these, ribonucleoprotein (RNP) granules are storage hubs for translationally repressed mRNA. Whether RNP granules can also activate translation and how this could be achieved remains unclear. Here, using single-molecule imaging, we demonstrate that the germ cell-determining RNP granules in Drosophila embryos are sites for active translation of nanos mRNA. Nanos translation occurs preferentially at the germ granule surface with the 3' UTR buried within the granule. Smaug, a cytosolic RNA-binding protein, represses nanos translation, which is relieved when Smaug is sequestered to the germ granule by the scaffold protein Oskar. Together, our findings uncover a molecular process by which RNP granules achieve localized protein synthesis through the compartmentalized loss of translational repression.
Sujet(s)
Régions 3' non traduites , Granulations cytoplasmiques , Protéines de Drosophila , Drosophila melanogaster , Biosynthèse des protéines , ARN messager , Protéines de liaison à l'ARN , Ribonucléoprotéines , Animaux , Protéines de Drosophila/métabolisme , Protéines de Drosophila/génétique , Protéines de liaison à l'ARN/métabolisme , Protéines de liaison à l'ARN/génétique , Ribonucléoprotéines/métabolisme , Ribonucléoprotéines/génétique , Drosophila melanogaster/métabolisme , Drosophila melanogaster/génétique , Granulations cytoplasmiques/métabolisme , Régions 3' non traduites/génétique , ARN messager/métabolisme , ARN messager/génétique , Imagerie de molécules uniques , Embryon non mammalien/métabolisme , Régulation de l'expression des gènes au cours du développement , Cellules germinales/métabolismeRÉSUMÉ
Deregulation or loss of the human leukocyte antigen class I (HLA-I) molecules on tumor cells leading to inhibition of CD8+ T cell recognition is an important tumor immune escape strategy, which could be caused by a posttranscriptional control of molecules in the HLA-I pathway mediated by RNA-binding proteins (RBPs). So far, there exists only limited information about the interaction of RBPs with HLA-I-associated molecules, but own work demonstrated a binding of the heterogeneous ribonucleoprotein C (hnRNP C) to the 3' untranslated region (UTR) of the TAP-associated glycoprotein tapasin (tpn). In this study, in silico analysis of pan-cancer TCGA datasets revealed that hnRNP C is higher expressed in tumor specimens compared to corresponding normal tissues, which is negatively correlated to tpn expression, T cell infiltration and the overall survival of tumor patients. Functional analysis demonstrated an upregulation of tpn expression upon siRNA-mediated downregulation of hnRNP C, which is accompanied by an increased HLA-I surface expression. Thus, hnRNP C has been identified to target tpn and its inhibition could improve the HLA-I surface expression on melanoma cells suggesting its use as a possible biomarker for T-cell-based tumor immunotherapies.
Sujet(s)
Régions 3' non traduites , Ribonucléoprotéine nucléaire hétérogène du groupe C , Mélanome , Protéines de transport membranaire , Humains , Mélanome/génétique , Mélanome/anatomopathologie , Mélanome/métabolisme , Mélanome/immunologie , Ribonucléoprotéine nucléaire hétérogène du groupe C/métabolisme , Ribonucléoprotéine nucléaire hétérogène du groupe C/génétique , Régions 3' non traduites/génétique , Protéines de transport membranaire/génétique , Protéines de transport membranaire/métabolisme , Lignée cellulaire tumorale , Régulation de l'expression des gènes tumorauxRÉSUMÉ
OBJECTIVE: To examine the influences and mechanisms of MicroRNA-19a-3p (miR-19a-3p) on endothelial dysfunction in atherosclerosis. METHODS: An analysis of miR-19a expression was carried out using the Gene Expression Omnibus (GEO) database. The effect of miR-19a-3p on endothelial function in HUVECs was evaluated by miR-19a-3p overexpression under TNF-α treatment. Luciferase assays were performed to explore the potential target genes. Overexpression of junctional protein associated with coronary artery disease (JCAD) was used to examine the effects of miR-19a-3p on cell adhesion, and proliferation. RESULTS: MiR-19a-3p expression in endothelial cells decreased after exposure to TNF-α and/or oscillatory flow, consistent with the expression change of miR-19a-3p found in atherosclerotic plaques. Additionally, endothelial cell dysfunction and inflammation were significantly diminished by miR-19a-3p overexpression but markedly exacerbated by miR-19a-3p inhibition. MiR-19a-3p transfection significantly decreased the expression of JCAD by binding to the 3'-UTR of JCAD mRNA. Furthermore, the protective effect of miR-19a-3p against endothelial cell dysfunction and inflammation was achieved by regulating JCAD and was closely linked to the Hippo/YAP signaling pathway. CONCLUSION: MiR-19a-3p expression is a crucial molecular switch in the onset of atherosclerosis and miR-19a-3p overexpression is a possible pharmacological therapeutic strategy for reversing the development of atherosclerosis.
Sujet(s)
Athérosclérose , Prolifération cellulaire , Cellules endothéliales de la veine ombilicale humaine , microARN , Transduction du signal , Protéines de signalisation YAP , microARN/génétique , microARN/métabolisme , Humains , Cellules endothéliales de la veine ombilicale humaine/métabolisme , Protéines de signalisation YAP/génétique , Protéines de signalisation YAP/métabolisme , Athérosclérose/génétique , Athérosclérose/métabolisme , Athérosclérose/anatomopathologie , Athérosclérose/physiopathologie , Adhérence cellulaire/effets des médicaments et des substances chimiques , Régulation de l'expression des gènes , Protéines adaptatrices de la transduction du signal/génétique , Protéines adaptatrices de la transduction du signal/métabolisme , Cellules cultivées , Bases de données génétiques , Facteur de nécrose tumorale alpha/métabolisme , Facteur de nécrose tumorale alpha/génétique , Régions 3' non traduites , Maladie des artères coronaires/génétique , Maladie des artères coronaires/métabolisme , Maladie des artères coronaires/physiopathologie , Voie de signalisation Hippo , Facteurs de transcription/génétique , Facteurs de transcription/métabolisme , Protein-Serine-Threonine Kinases/génétique , Protein-Serine-Threonine Kinases/métabolisme , Sites de fixationRÉSUMÉ
The RNA-binding protein LIN28B represses the biogenesis of the tumor suppressor let-7. The LIN28B/let-7 axis regulates cell differentiation and is associated with various cancers. The RNA-binding protein Q (hnRNP Q) or SYNCRIP (Synaptotagmin Binding Cytoplasmic RNA Interacting Protein) has been implicated in mRNA splicing, mRNA transport, translation, and miRNAs biogenesis as well as metabolism in cancer. To determine whether hnRNP Q plays a role in the LIN28B/let-7 axis, we tested for interactions between hnRNP Q and LIN28B. We demonstrated that hnRNP Q interacts with LIN28B in an RNA-dependent manner. Knockdown of hnRNP Q caused reduced expression of a well-known let-7 target TRIM71, an E3 ubiquitin ligase that belongs to the RBCC/TRIM family, and also LIN28B, whose mRNA itself is down-regulated by let-7. In addition, hnRNP Q knockdown increased let-7 family miRNA levels and reduced the activity of luciferase reporters fused with the TRIM71 3'UTR or a synthetic 3'UTR carrying 8X let-7 complementary sites. Finally, depletion of hnRNP Q inhibited the proliferation of a hepatocellular carcinoma cell line, Huh7. This observation is consistent with the survival curve for liver cancer patients from the TCGA database, which indicates that high expression of hnRNP Q is a prognostic marker for a poor outcome in individuals afflicted with hepatocellular carcinoma. Together, our findings suggest that hnRNP Q interacts with LIN28B and modulates the LIN28B/let-7 axis in hepatocellular carcinoma.
Sujet(s)
Carcinome hépatocellulaire , Ribonucléoprotéines nucléaires hétérogènes , Tumeurs du foie , microARN , Protéines de liaison à l'ARN , Humains , Protéines de liaison à l'ARN/métabolisme , Protéines de liaison à l'ARN/génétique , microARN/génétique , microARN/métabolisme , Carcinome hépatocellulaire/génétique , Carcinome hépatocellulaire/métabolisme , Carcinome hépatocellulaire/anatomopathologie , Tumeurs du foie/métabolisme , Tumeurs du foie/génétique , Tumeurs du foie/anatomopathologie , Ribonucléoprotéines nucléaires hétérogènes/métabolisme , Ribonucléoprotéines nucléaires hétérogènes/génétique , Lignée cellulaire tumorale , Régulation de l'expression des gènes tumoraux , Prolifération cellulaire , Liaison aux protéines , Régions 3' non traduitesRÉSUMÉ
N6-methyladenosine (m6A) plays critical roles in regulating mRNA metabolism. However, comprehensive m6A methylomes in different plant tissues with single-base precision have yet to be reported. Here, we present transcriptome-wide m6A maps at single-base resolution in different tissues of rice and Arabidopsis using m6A-SAC-seq. Our analysis uncovers a total of 205,691 m6A sites distributed across 22,574 genes in rice, and 188,282 m6A sites across 19,984 genes in Arabidopsis. The evolutionarily conserved m6A sites in rice and Arabidopsis ortholog gene pairs are involved in controlling tissue development, photosynthesis and stress response. We observe an overall mRNA stabilization effect by 3' UTR m6A sites in certain plant tissues. Like in mammals, a positive correlation between the m6A level and the length of internal exons is also observed in plant mRNA, except for the last exon. Our data suggest an active m6A deposition process occurring near the stop codon in plant mRNA. In addition, the MTA-installed plant mRNA m6A sites correlate with both translation promotion and translation suppression, depicting a more complicated regulatory picture. Our results therefore provide in-depth resources for relating single-base resolution m6A sites with functions in plants and uncover a suppression-activation model controlling m6A biogenesis across species.
Sujet(s)
Adénosine , Arabidopsis , Régulation de l'expression des gènes végétaux , Oryza , ARN messager , Oryza/génétique , Oryza/croissance et développement , Oryza/métabolisme , Arabidopsis/génétique , Arabidopsis/métabolisme , Arabidopsis/croissance et développement , Adénosine/analogues et dérivés , Adénosine/métabolisme , ARN messager/métabolisme , ARN messager/génétique , Transcriptome/génétique , ARN des plantes/génétique , ARN des plantes/métabolisme , Régions 3' non traduites/génétique , Analyse de profil d'expression de gènes/méthodes , Stabilité de l'ARN/génétique , Exons/génétique , Codon stop/génétiqueRÉSUMÉ
2'-O-methylation (Nm) is a prominent RNA modification well known in noncoding RNAs and more recently also found at many mRNA internal sites. However, their function and base-resolution stoichiometry remain underexplored. Here, we investigate the transcriptome-wide effect of internal site Nm on mRNA stability. Combining nanopore sequencing with our developed machine learning method, NanoNm, we identify thousands of Nm sites on mRNAs with a single-base resolution. We observe a positive effect of FBL-mediated Nm modification on mRNA stability and expression level. Elevated FBL expression in cancer cells is associated with increased expression levels for 2'-O-methylated mRNAs of cancer pathways, implying the role of FBL in post-transcriptional regulation. Lastly, we find that FBL-mediated 2'-O-methylation connects to widespread 3' UTR shortening, a mechanism that globally increases RNA stability. Collectively, we demonstrate that FBL-mediated Nm modifications at mRNA internal sites regulate gene expression by enhancing mRNA stability.