ABSTRACT
[This corrects the article DOI: 10.7150/thno.40144.].
Subject(s)
Kidney Calculi , Lasers, Solid-State , Lithotripsy, Laser , Lithotripsy , Humans , Kidney , Kidney Calculi/therapySubject(s)
COVID-19 , Humans , Broadly Neutralizing Antibodies , Antibodies, Viral , Antibodies, NeutralizingABSTRACT
The mechanistic action of histone deacetylase 8 (HDAC8) in cancer motility, including epithelial-mesenchymal transition (EMT), remains largely undefined. We found that the expression of HDAC8 was upregulated in breast cancer (BC) cells and tissues as compared to the controls. Further, BC tissues had the highest values of HDAC8 expression among 31 kinds of cancers. Cellular study indicated that HDAC8 can positively regulate the dissemination and EMT of BC cells. It increased the protein stability of Snail, an important regulator of EMT, by phosphorylation of its motif 2 in serine-rich regions. There are 21 factors that have been reported to regulate the protein stability of Snail. Among them, HDAC8 can decrease the expression of GSK-3ß through increasing its Ser9-phosphorylation. Mass spectrum analysis indicated that HDAC8 interact with AKT1 to decrease its acetylation while increase its phosphorylation, which further increased Ser9-phosphorylation of GSK-3ß. The C-terminal of AKT1 was responsible for the interaction between HDAC8 and AKT1. Further, Lys426 was the key residue for HDAC8-regulated deacetylation of AKT1. Moreover, HDAC8/Snail axis acted as adverse prognosis factors for in vivo progression and overall survival (OS) rate of BC patients. Collectively, we found that HDAC8 can trigger the dissemination of BC cells via AKT/GSK-3ß/Snail signals, which imposed that inhibition of HDAC8 is a potential approach for BC treatment.
Subject(s)
Breast Neoplasms/genetics , Glycogen Synthase Kinase 3 beta/genetics , Histone Deacetylases/genetics , Proto-Oncogene Proteins c-akt/genetics , Repressor Proteins/genetics , Signal Transduction/genetics , Snail Family Transcription Factors/genetics , Animals , Breast Neoplasms/metabolism , Breast Neoplasms/therapy , Cell Line, Tumor , Cell Movement/genetics , Female , Gene Expression Regulation, Neoplastic , Glycogen Synthase Kinase 3 beta/metabolism , Histone Deacetylases/metabolism , Humans , Kaplan-Meier Estimate , Mice, Nude , Proto-Oncogene Proteins c-akt/metabolism , Repressor Proteins/metabolism , Snail Family Transcription Factors/metabolism , Xenograft Model Antitumor Assays/methodsABSTRACT
Studies on biological functions of N6-methyladenosine (m6A) modification in mRNA have drawn significant attention in recent years. Here we describe the construction and characterization of a CRISPR-Cas13b-based tool for targeted demethylation of specific mRNA. A fusion protein, named dm6ACRISPR, was created by linking a catalytically inactive Type VI-B Cas13 enzyme from Prevotella sp. P5-125 (dPspCas13b) to m6A demethylase AlkB homolog 5 (ALKBH5). dm6ACRISPR specifically demethylates m6A of targeted mRNA such as cytochrome b5 form A (CYB5A) to increase its mRNA stability. It can also demethylate ß-catenin-encoding CTNNB1 mRNA that contains multiple m6A sites to trigger its translation. In addition, the dm6ACRISPR system incurs efficient demethylation of targeted epitranscriptome transcripts with limited off-target effects. Targeted demethylation of transcripts coding for oncoproteins such as epidermal growth factor receptor (EGFR) and MYC can suppress proliferation of cancer cells. Together, we provide a programmable and in vivo manipulation tool to study mRNA modification of specific genes and their related biological functions.
Subject(s)
AlkB Homolog 5, RNA Demethylase/metabolism , CRISPR-Associated Proteins/genetics , CRISPR-Cas Systems , RNA, Messenger/metabolism , 5' Untranslated Regions , Adenosine/analogs & derivatives , Adenosine/metabolism , AlkB Homolog 5, RNA Demethylase/genetics , Cell Proliferation , Demethylation , HEK293 Cells , HeLa Cells , Humans , Oncogenes , Prevotella/enzymology , Protein Engineering , RNA, Messenger/chemistry , Recombinant Fusion Proteins/metabolismABSTRACT
Studies on biological functions of N6-methyladenosine (m6A) modification in mRNA have sprung up in recent years. We find m6A can positively regulate the glycolysis of cancer cells. Specifically, m6A-sequencing and functional studies confirm that pyruvate dehydrogenase kinase 4 (PDK4) is involved in m6A regulated glycolysis and ATP generation. The m6A modified 5'UTR of PDK4 positively regulates its translation elongation and mRNA stability via binding with YTHDF1/eEF-2 complex and IGF2BP3, respectively. Targeted specific demethylation of PDK4 m6A by dm6ACRISPR system can significantly decrease the expression of PDK4 and glycolysis of cancer cells. Further, TATA-binding protein (TBP) can transcriptionally increase the expression of Mettl3 in cervical cancer cells via binding to its promoter. In vivo and clinical data confirm the positive roles of m6A/PDK4 in tumor growth and progression of cervical and liver cancer. Our study reveals that m6A regulates glycolysis of cancer cells through PDK4.
Subject(s)
Adenosine/analogs & derivatives , Glycolysis/physiology , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/metabolism , Uterine Cervical Neoplasms/genetics , 5' Untranslated Regions , Adenosine/metabolism , Adenosine Triphosphate/metabolism , Animals , Clustered Regularly Interspaced Short Palindromic Repeats , Female , Glycolysis/genetics , HeLa Cells , Humans , Kaplan-Meier Estimate , Methyltransferases/genetics , Methyltransferases/metabolism , Mice, Inbred BALB C , Promoter Regions, Genetic , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/genetics , RNA Stability , RNA-Binding Proteins/metabolism , Uterine Cervical Neoplasms/mortality , Xenograft Model Antitumor AssaysABSTRACT
Background: Drug resistance severely reduces treatment efficiency of chemotherapy and leads to poor prognosis. However, regulatory factors of chemoresistant cancer cells are largely unknown. Methods: The expression of estrogen receptor related receptors (ERRs) in chemoresistant cancer cells are checked. The roles of ERRγ in chemoresistance are confirmed by in vitro and in vivo studies. The mechanisms responsible for ERRγ-regulated expression of ABCB1 and CPT1B are investigated. Results: The expression of ERRγ is upregulated in chemoresistant cancer cells. Targeted inhibition of ERRγ restores the chemosensitivity. ERRγ can directly bind to the promoter of ABCB1 to increase its transcription. An elevated interaction between ERRγ and p65 in chemoresistant cells further strengthens transcription of ABCB1. Further, ERRγ can increase the fatty acid oxidation (FAO) in chemoresistant cells via regulation of CPT1B, the rate-limiting enzyme of FAO. The upregulated ERRγ in chemoresistant cancer cells might be due to increased levels of N6-methyladenosine (m6A) can trigger the splicing of precursor ESRRG mRNA. Conclusions: m6A induced ERRγ confers chemoresistance of cancer cells through upregulation of ABCB1 and CPT1B.
Subject(s)
Adenosine/analogs & derivatives , Drug Resistance, Neoplasm , Receptors, Estrogen/metabolism , ATP Binding Cassette Transporter, Subfamily B/genetics , ATP Binding Cassette Transporter, Subfamily B/metabolism , Adenosine/pharmacology , Animals , Cell Line, Tumor , Drug Delivery Systems , Drug Resistance, Multiple , Drug Resistance, Neoplasm/drug effects , Fatty Acids/metabolism , Gene Expression Regulation, Neoplastic , Hep G2 Cells , Humans , MCF-7 Cells , Mice , Up-RegulationABSTRACT
N6-methyladenosine (m6A) is the most abundant modification on eukaryotic mRNA, which regulates all steps of the mRNA life cycle. An increasing number of studies have shown that m6A methylation plays essential roles in tumor development. However, the relationship between m6A and the progression of cancers remains to be explored. Here, we reported that transforming growth factor-ß (TGFß1)-induced epithelial-mesenchymal transition (EMT) was inhibited in methyltransferase-like 3 (METTL3) knockdown (Mettl3Mut/-) cells. The expression of TGFß1 was up-regulated, while self-stimulated expression of TGFß1 was suppressed in Mettl3Mut/- cells. We further revealed that m6A promoted TGFB1 mRNA decay, but impaired TGFB1 translation progress. Besides this, the autocrine of TGFß1 was disrupted in Mettl3Mut/- cells via interrupting TGFß1 dimer formation. Lastly, we found that Snail, which was down-regulated in Mettl3Mut/- cells, was a key factor responding to TGFß1-induced EMT. Together, our research demonstrated that m6A performed multi-functional roles in TGFß1 expression and EMT modulation, suggesting the critical roles of m6A in cancer progression regulation.
Subject(s)
Adenosine/analogs & derivatives , Lung Neoplasms/metabolism , Methyltransferases/metabolism , Transforming Growth Factor beta1/genetics , 5' Untranslated Regions , Adenosine/metabolism , Animals , Cell Movement/drug effects , Cell Movement/genetics , Down-Regulation , Epithelial-Mesenchymal Transition/genetics , Gene Knockdown Techniques , HeLa Cells , Humans , Lung Neoplasms/genetics , Methyltransferases/genetics , Mice , Peptide Chain Elongation, Translational/drug effects , Protein Biosynthesis/drug effects , Protein Biosynthesis/genetics , Protein Stability/drug effects , Snail Family Transcription Factors/genetics , Snail Family Transcription Factors/metabolism , Transforming Growth Factor beta1/metabolism , Transforming Growth Factor beta1/pharmacologyABSTRACT
BACKGROUND: Brain metastasis (BM) is one of the principal causes of mortality for lung cancer patients. While the molecular events that govern BM of lung cancer remain frustrating cloudy. METHODS: The miRNA expression profiles are checked in the paired human BM and primary lung cancer tissues. The effect of miR-143-3p on BM of lung cancer cells and its related mechanisms are investigated. RESULTS: miR-143-3p is upregulated in the paired BM tissues as compared with that in primary cancer tissues. It can increase the invasion capability of in vitro blood brain barrier (BBB) model and angiogenesis of lung cancer by targeting the three binding sites of 3'UTR of vasohibin-1 (VASH1) to inhibit its expression. Mechanistically, VASH1 can increase the ubiquitylation of VEGFA to trigger the proteasome mediated degradation, further, it can endow the tubulin depolymerization through detyrosination to increase the cell motility. m6A methyltransferase Mettl3 can increase the splicing of precursor miR-143-3p to facilitate its biogenesis. Moreover, miR-143-3p/VASH1 axis acts as adverse prognosis factors for in vivo progression and overall survival (OS) rate of lung cancer. CONCLUSIONS: Our work implicates a causal role of the miR-143-3p/VASH1 axis in BM of lung cancers and suggests their critical roles in lung cancer pathogenesis.
Subject(s)
Adenosine/analogs & derivatives , Brain Neoplasms/secondary , Cell Cycle Proteins/genetics , Gene Expression Regulation, Neoplastic , Lung Neoplasms/genetics , Lung Neoplasms/pathology , MicroRNAs/genetics , Animals , Blood-Brain Barrier/metabolism , Cell Line, Tumor , Disease Models, Animal , Disease Progression , Epithelial-Mesenchymal Transition/genetics , Gene Expression Profiling , Humans , Lung Neoplasms/metabolism , Mice , Models, Biological , Neovascularization, Pathologic/genetics , Neovascularization, Pathologic/metabolism , RNA Interference , Vascular Endothelial Growth Factor A/genetics , Vascular Endothelial Growth Factor A/metabolism , Xenograft Model Antitumor AssaysABSTRACT
Brain-derived neurotrophic factor (BDNF) is a master regulator of synaptic plasticity in various neural circuits of the mammalian central nervous system. Neuron activity-induced BDNF gene expression is regulated through the Ca2+/CREB pathway, but other regulatory factors may also be involved in controlling BDNF levels. We report here that Wnt/ß-catenin signaling plays a key role in controlling neuron activity-regulated BDNF expression. Using primary cortical cultures, we show that blockade of Wnt/ß-catenin signaling inhibits the BDNF up-regulation that is induced by activation of the N-methyl-d-aspartic acid (NMDA) receptor and that activation of the Wnt/ß-catenin signaling pathway stimulates BDNF expression. In vivo, Wnt/ß-catenin signaling activated BDNF expression and was required for peripheral pain-induced up-regulation of BDNF in the mouse spine. We also found that conditional deletion of one copy of either Wntless (Wls) or ß-catenin by Nestin-Cre-mediated recombination is sufficient to inhibit the pain-induced up-regulation of BDNF. We further show that the Wnt/ß-catenin/BDNF axis in the spinal neural circuit plays an important role in regulating capsaicin-induced pain. These results indicate that neuron activity-induced Wnt signaling stimulates BDNF expression in the pain neural circuits. We propose that pain-induced Wnt secretion may provide an additional mechanism for intercellular coordination of BDNF expression in the neural circuit.
Subject(s)
Brain-Derived Neurotrophic Factor/genetics , Cerebral Cortex/metabolism , Neurons/metabolism , Pain/genetics , Wnt3A Protein/genetics , beta Catenin/genetics , Animals , Anti-Anxiety Agents/pharmacology , Azepines/pharmacology , Benzamides/pharmacology , Brain-Derived Neurotrophic Factor/agonists , Brain-Derived Neurotrophic Factor/metabolism , Capsaicin/administration & dosage , Cerebral Cortex/drug effects , Embryo, Mammalian , Gene Expression Regulation , Hindlimb , Intracellular Signaling Peptides and Proteins/genetics , Intracellular Signaling Peptides and Proteins/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Neuronal Plasticity/physiology , Neurons/drug effects , Pain/chemically induced , Pain/drug therapy , Pain/physiopathology , Primary Cell Culture , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/metabolism , Receptors, N-Methyl-D-Aspartate/genetics , Receptors, N-Methyl-D-Aspartate/metabolism , Spinal Cord/drug effects , Spinal Cord/metabolism , Spinal Cord/physiopathology , Transcription, Genetic , Wnt Signaling Pathway , Wnt3A Protein/metabolism , beta Catenin/metabolismABSTRACT
Mammalian target of rapamycin (mTOR) signaling plays a critical role in the regulation of activity-dependent protein synthesis in neurons. It is well established that the GTPase-activating protein tuberous sclerosis complex proteins (2TSC2) is an upstream inhibitor of mTOR. In this study, we show that glutamate stimulation down-regulates TSC2 protein in cortical cultures via NMDA receptor (NMDAR) activation. Interestingly, the mTOR-specific inhibitor rapamycin blocks the glutamate-induced TSC2 down-regulation. This finding suggests that NMDAR activation evokes an mTOR-mediated negative regulation of TSC2. In addition, we also show that the glutamate-induced down-regulation of TSC2 protein is blocked by proteasome inhibitor MG132, indicating the involvement of proteasome-mediated protein degradation. We propose that the NMDAR activation stimulates an mTOR-proteasome pathway to degrade TSC2 protein.
