bims-rimeca 2023-05-07 papers

bims-rimeca

Biomed News

on RNA methylation in cancer

Issue of 2023–05–07
twenty papers selected by
Sk Ramiz Islam, Saha Institute of Nuclear Physics

Advances in targeting RNA modifications for anticancer therapy.
Research Progress of m6A RNA Methylation in Skin Diseases.
The increase of Nrf2 m6A modification induced by FTO downregulation promotes hippocampal neuron injury and aggravates the progression of epilepsy in a rat model.
Fat mass and obesity-associated factor (FTO)-mediated N6-methyladenosine regulates spermatogenesis in an age-dependent manner.
RNA methylation pattern and immune microenvironment characteristics mediated by m6A regulator in ischemic stroke.
ZBTB7C m6A modification incurred by METTL3 aberration promotes osteosarcoma progression.
RNA demethylation-driven functional supramolecular structure for label-free detection of m6A modification eraser FTO in human breast tissues.
Comprehensive analysis of a novel RNA modifications-related model in the prognostic characterization, immune landscape and drug therapy of bladder cancer.
Peritoneal high-fat environment promotes peritoneal metastasis of gastric cancer cells through activation of NSUN2-mediated ORAI2 m5C modification.
KRT17 promotes T-lymphocyte infiltration through the YTHDF2-CXCL10 axis in colorectal cancer.
Mettl14 sustains FOXP3 expression to promote the differentiation and functions of induced-regulatory T cells via the mTOR signaling pathway.
LncRNA GAS5 regulated by FTO-mediated m6A demethylation promotes autophagic cell death in NSCLC by targeting UPF1/BRD4 axis.
m6A readers ECT2/ECT3/ECT4 enhance mRNA stability through direct recruitment of the poly(A) binding proteins in Arabidopsis.
LncRNA FEZF1-AS1 accelerates multiple myeloma progression by regulating IGF2BP1/BZW2 signaling.
KLF2 inhibits colorectal cancer progression and metastasis by inducing ferroptosis via the PI3K/AKT signaling pathway.
m6A-mediated upregulation of lncRNA-AC026356.1 promotes cancer stem cell maintenance in lung adenocarcinoma via activating Wnt signaling pathway.
NUF2 is correlated with a poor prognosis and immune infiltration in clear cell renal cell carcinoma.
Low RNA stability signifies increased post-transcriptional regulation of cell identity genes.
Myocardial IGF2R is a critical mediator of inflammation and fibrosis after ischemia-reperfusion injury.
SH3BGRL Suppresses Liver Tumor Progression through Enhanced ATG5-Dependent Autophagy.

Trends Cancer. 2023 May 04. pii: S2405-8033(23)00059-6. [Epub ahead of print]

Advances in targeting RNA modifications for anticancer therapy.

Monica M Pomaville, Chuan He.

  Numerous strategies are employed by cancer cells to control gene expression and facilitate tumorigenesis. In the study of epitranscriptomics, a diverse set of modifications to RNA represent a new player of gene regulation in disease and in development. N6-methyladenosine (m6A) is the most common modification on mammalian messenger RNA and tends to be aberrantly placed in cancer. Recognized by a series of reader proteins that dictate the fate of the RNA, m6A-modified RNA could promote tumorigenesis by driving protumor gene expression signatures and altering the immunologic response to tumors. Preclinical evidence suggests m6A writer, reader, and eraser proteins are attractive therapeutic targets. First-in-human studies are currently testing small molecule inhibition against the methyltransferase-like 3 (METTL3)/methyltransferase-like 14 (METTL14) methyltransferase complex. Additional modifications to RNA are adopted by cancers to drive tumor development and are under investigation.

Keywords:  METTL3; N(6)-methyladenosine; RNA modifications; cancer; epitranscriptomics

DOI:  https://doi.org/10.1016/j.trecan.2023.04.003
Biomed Res Int. 2023 ;2023 3091204

Research Progress of m6A RNA Methylation in Skin Diseases.

Chang Liu, Xin Wang, Shengju Yang, Shuanglin Cao.

N6-Methyladenosine (m6A) is the most common mRNA modification in eukaryotes and is a dynamically reversible posttranscriptional modification. The enzymes involved in m6A modification mainly include methyltransferases (writers), demethylases (erasers), and methylated readers (Readers). m6A modification is mainly catalyzed by m6A methyltransferase and removed by m6A demethylase. The modified RNA can be specifically recognized and bound by m6A recognition protein. This protein complex then mediates RNA splicing, maturation, nucleation, degradation, and translation. m6A also alters gene expression and regulates cellular processes such as self-renewal, differentiation, invasion, and apoptosis. An increasing body of evidence indicates that the m6A methylation modification process is closely related to the occurrence of various skin diseases. In this review, we discuss the role of m6A methylation in skin development and skin diseases including psoriasis, melanoma, and cutaneous squamous cell carcinoma.

DOI: https://doi.org/10.1155/2023/3091204
Synapse. 2023 Apr 30. e22270

The increase of Nrf2 m6A modification induced by FTO downregulation promotes hippocampal neuron injury and aggravates the progression of epilepsy in a rat model.

Mao-Qiang Tian, Juan Li, Xiao-Mei Shu, Chang-Hui Lang, Jing Chen, Long-Ying Peng, Wen-Ting Lei, Chang-Jian Yang.

  Epilepsy is a common chronic neurological disorder characterized by widespread neuronal death. The purpose of this study was to investigate the role of nuclear factor erythroid 2-related factor 2 (Nrf2) m6A methylation in epilepsy. To create epileptic models, the rats were given Lithium chloride and pilocarpine, and isolated primary rat hippocampal neurons were cultured in an Mg2+ -free medium. The frequency of seizures was recorded in the epilepsy group of rats. The functional tests included TUNEL, MTT, and flow cytometry. Mechanistically, RNA degradation assay, RNA immunoprecipitation, and methylated RNA immunoprecipitation were performed. In epileptic models, Nrf2 and fat mass and obesity-associated (FTO) levels were downregulated, whereas YT521-B homology (YTH) domain family protein 2 (YTHDF2) was upregulated. Additionally, in epileptic models, there was a rise in the m6A methylation level of Nrf2 mRNA. Overexpressing FTO increased cell viability and reduced apoptosis, but Nrf2 interference reversed these effects. Meanwhile, FTO overexpression decreased the m6A methylation of Nrf2 mRNA. Moreover, YTHDF2 bound to Nrf2 mRNA and decreased its stability. Furthermore, FTO overexpression reduced seizure frequency in rats and inhibited hippocampal neuron apoptosis via lowering the m6A methylation level of Nrf2 mRNA. Overexpressing FTO reduced m6A methylation of Nrf2 mRNA, increased cell viability, suppressed apoptosis, and slowed the progression of epileptic diseases, which is linked to YTHDF2 binding to m6A-modified Nrf2 and promoting its degradation, as well as downregulating Nrf2 expression in hippocampal neurons.

Keywords:  FTO; Nrf2; YTHDF2; epilepsy; m6A methylation

DOI:  https://doi.org/10.1002/syn.22270
J Biol Chem. 2023 May 03. pii: S0021-9258(23)01811-2. [Epub ahead of print] 104783

Fat mass and obesity-associated factor (FTO)-mediated N6-methyladenosine regulates spermatogenesis in an age-dependent manner.

Yifei Wu, Jincheng Li, Chenmeijie Li, Shuai Lu, Xiaoyu Wei, Yang Li, Wenjuan Xia, Chunfeng Qian, Zihang Wang, Mingxi Liu, Yayun Gu, Boxian Huang, Yueqiu Tan, Zhibin Hu.

  N6-methyladenosine (m6A) is the most prevalent reversible RNA modification in the mammalian transcriptome. It has recently been demonstrated that m6A is crucial for male germline development. Fat mass and obesity-associated factor (FTO), a known m6A demethylase, is widely expressed in human and mouse tissues and is involved in manifold biological processes and human diseases. However, the function of FTO in spermatogenesis and male fertility remains poorly understood. Here, we generated an Fto knockout mouse model using CRISPR/Cas9-mediated genome editing techniques to address this knowledge gap. Remarkably, we found that loss of Fto in mice caused spermatogenesis defects in an age-dependent manner, resulting from the attenuated proliferation ability of undifferentiated spermatogonia and increased male germ cell apoptosis. Further research showed that FTO plays a vital role in the modulation of spermatogenesis and Leydig cell maturation by regulating the translation of the androgen receptor in an m6A-dependent manner. In addition, we identified two functional mutations of FTO in male infertility patients, resulting in truncated FTO protein and increased m6A modification in vitro. Our results highlight the crucial effects of FTO on spermatogonia and Leydig cells for the long-term maintenance of spermatogenesis and expand our understanding of the function of m6A in male fertility.

Keywords:  FTO; age-dependent; androgen receptor; m6A; spermatogenesis

DOI:  https://doi.org/10.1016/j.jbc.2023.104783
Front Genet. 2023 ;14 1148510

RNA methylation pattern and immune microenvironment characteristics mediated by m6A regulator in ischemic stroke.

Kejuan Jia, Wenbo Xia, Qian Su, Shiqi Yang, Yanli Zhang, Xunran Ni, Zhiqiang Su, Delong Meng.

  Background: Ischemic stroke (IS) is a highly heterogeneous disease. Recent studies have shown that epigenetic variables affect the immune response. However, only a few studies have examined the relationship between IS and m6A immunoregulation. Therefore, we aim to explore the methylation of RNA mediated by m6A regulatory factor and the immune microenvironment characteristics of IS. Methods: Differentially expressed m6A regulators were detected in IS microarray datasets GSE22255 and GSE58294. We used a series of machine learning algorithms to identify key IS-related m6A regulators and validated them on blood samples of IS patients, oxygen-glucose deprivation/reoxygenation (OGD/R) microglia and GSE198710 independent data sets. Different m6A modification modes were determined and the patients were classified. In addition, we systematically associate these modification patterns with the characteristics of immune microenvironment, including infiltrating immune cells, immune function genes and immune response genes. Then we developed a model of m6A score to quantify the m6A modification in IS samples. Results: Through the analysis of the differences between the control group and IS patients, METTL16, LRPPRC, and RBM15 showed strong diagnostic significance in three independent data sets. In addition, qRT-PCR and Western blotting also confirmed that the expression of METTL16 and LRPPRC was downregulated and the expression of RBM15 was upregulated after ischemia. Two m6A modification modes and two m6A gene modification modes were also identified. m6A gene cluster A (high m6A value group) was positively correlated with acquired immunity, while m6A gene cluster B (low m6A value group) was positively correlated with innate immunity. Similarly, five immune-related hub genes were significantly associated with m6Acore (CD28, IFNG, LTF, LCN2, and MMP9). Conclusion: The modification of m6A is closely related to the immune microenvironment. The evaluation of individual m6A modification pattern may be helpful for future immunomodulatory therapy of anti-ischemic response.

Keywords:  immune microenvironment; ischemic stroke; m6A; m6A modification pattern; machine learning

DOI:  https://doi.org/10.3389/fgene.2023.1148510
Transl Res. 2023 Apr 28. pii: S1931-5244(23)00072-5. [Epub ahead of print]

ZBTB7C m6A modification incurred by METTL3 aberration promotes osteosarcoma progression.

Xueying An, Wenshu Wu, Lin Yang, Jian Dong, Bin Liu, Junxia Guo, Jianmei Chen, Baosheng Guo, Wangsen Cao, Qing Jiang.

  Aberrant N6-methyladenosine (m6A) modification of mRNAs contributes significantly to the epigenetic tumorigenesis, however, its precise role and the key targets in osteosarcoma (OS) are not defined. Here we reported that selective METTL3 (methyltransferase like 3) elevation and the consequential increase of m6A modification causally affect OS progression. The fast-growing OS cells displayed preferential upregulation of METTL3 and increased m6A modification. Conversely, m6A inhibition by 3-deazaadenosine, siRNA-mediated METTL3 knockdown or a METTL3-selective inhibitor by STM2457 effectively inhibits OS cell growth and induced OS cell apoptosis. Further investigation revealed that an oncogenic protein ZBTB7C was likely a critical m6A target that mediated the oncogenic effects. ZBTB7C mRNA contains a typical m6A motif of high confidence and its mRNA and protein were enriched with increased m6A modification in OS samples/cells. In an OS xenograft model, STM2457 or siRNA-mediated METTL3 knockdown effectively lowed ZBTB7C abundance. More importantly, the anti-OS effects of STM2457 were significantly reduced when ZBTB7C was overexpressed by lentivirus. Together, our results demonstrate that the METTL3 aberration and the resultant ZBTB7C m6A modification form an important epigenetic regulatory loop that promotes OS progression, and targeting the METTL3/ZBTB7C axis may provide novel insights into the potential strategies for OS therapy.

Keywords:  Apoptosis; METTL3; N6-methyladenosine; Osteosarcoma; ZBTB7C

DOI:  https://doi.org/10.1016/j.trsl.2023.04.005
Anal Chim Acta. 2023 Jun 15. pii: S0003-2670(23)00429-4. [Epub ahead of print]1260 341208

RNA demethylation-driven functional supramolecular structure for label-free detection of m6A modification eraser FTO in human breast tissues.

Wen-Xuan Li, Su Jiang, Wen-Jing Liu, Chun-Yang Zhang.

  Fat mass and obesity-associated enzyme (FTO) can dynamically regulate N6-methyladenosine modification, and it is engaged in various cellular functions. Herein, we demonstrate the RNA demethylation-driven functional supramolecular structure for label-free detection of m6A modification eraser FTO in human breast tissues. The presence of FTO catalyzes the removal of methyl group in m6A, causing the cleavage of demethylated DNA by DpnII and the release of DNA primer. The resultant DNA primer hybridizes with circular template to initiate isothermal rolling circle amplification (RCA), producing abundant long ssDNA polymers with repeating sequences of G-quadruplex. Subsequently, N-methylmesoporphyrin IX (NMM) is selectively embedded into G-quadruplex DNAzyme to form a supramolecular NMM-G-quadruplex structure for the generation of an amplified fluorescence signal. Benefiting from high selectivity of DpnII toward demethylated DNA, high amplification efficiency of RCA, and high signal-to-noise ratio of G-quadruplex-NMM system, this assay can sensitively detect FTO with a limit of detection (LOD) of 3.10 × 10-16 M, screen RNA demethylase inhibitors, quantify FTO activity in cancer cells, and discriminate FTO activity between breast cancer patient tissues and healthy person tissues. Importantly, this assay can be homogeneously conducted in a label-free manner, with great potential in RNA demethylases-related pathogenesis research and clinical diagnostics.

Keywords:  Clinical diagnostics; Fat mass and obesity-associated enzyme (FTO); Label-free assay; N-methylmesoporphyrin IX; Supramolecular structure

DOI:  https://doi.org/10.1016/j.aca.2023.341208
Front Genet. 2023 ;14 1156095

Comprehensive analysis of a novel RNA modifications-related model in the prognostic characterization, immune landscape and drug therapy of bladder cancer.

Ziying Zhang, Peng Chen, Jingping Yun.

  Background: Bladder cancer (BCa) is the leading reason for death among genitourinary malignancies. RNA modifications in tumors closely link to the immune microenvironment. Our study aimed to propose a promising model associated with the "writer" enzymes of five primary RNA adenosine modifications (including m6A, m6Am, m1A, APA, and A-to-I editing), thus characterizing the clinical outcome, immune landscape and therapeutic efficacy of BCa. Methods: Unsupervised clustering was employed to categorize BCa into different RNA modification patterns based on gene expression profiles of 34 RNA modification "writers". The RNA modification "writers" score (RMS) signature composed of RNA phenotype-associated differentially expressed genes (DEGs) was established using the least absolute shrinkage and selection operator (LASSO), which was evaluated in meta-GEO (including eight independent GEO datasets) training cohort and the TCGA-BLCA validation cohort. The hub genes in the RMS model were determined via weighted gene co-expression network analysis (WGCNA) and were further validated using human specimen. The potential applicability of the RMS model in predicting the therapeutic responsiveness was assessed through the Genomics of Drug Sensitivity in Cancer database and multiple immunotherapy datasets. Results: Two distinct RNA modification patterns were determined among 1,410 BCa samples from a meta-GEO cohort, showing radically varying clinical outcomes and biological characteristics. The RMS model comprising 14 RNA modification phenotype-associated prognostic DEGs positively correlated with the unsatisfactory outcome of BCa patients in meta-GEO training cohort (HR = 3.00, 95% CI = 2.19-4.12) and TCGA-BLCA validation cohort (HR = 1.53, 95% CI = 1.13-2.09). The infiltration of immunosuppressive cells and the activation of EMT, angiogenesis, IL-6/JAK/STAT3 signaling were markedly enriched in RMS-high group. A nomogram exhibited high prognostic prediction accuracy, with a concordance index of 0.785. The therapeutic effect of chemotherapeutic agents and antibody-drug conjugates was significantly different between RMS-low and -high groups. The combination of the RMS model and conventional characteristics (TMB, TNB and PD-L1) achieved an optimal AUC value of 0.828 in differentiating responders from non-responders to immunotherapy. Conclusion: We conferred the first landscape of five forms of RNA modifications in BCa and emphasized the excellent power of an RNA modifications-related model in evaluating BCa prognosis and immune landscape.

Keywords:  RNA modification; bladder cancer; immunotherapy efficacy; patient stratification; prognostic model

DOI:  https://doi.org/10.3389/fgene.2023.1156095
Oncogene. 2023 May 02.

Peritoneal high-fat environment promotes peritoneal metastasis of gastric cancer cells through activation of NSUN2-mediated ORAI2 m5C modification.

Kanghui Liu, Peng Xu, Jialun Lv, Han Ge, Zhengyuan Yan, Shansong Huang, Bowen Li, Hao Xu, Li Yang, Zekuan Xu, Diancai Zhang.

Peritoneal metastasis (PM) is an important metastatic modality of gastric cancer (GC).It is associated with poor prognosis. The underlying molecular mechanism of PM remains elusive. 5-Methylcytosine (m5C), a posttranscriptional RNA modification, involves in the progression of many tumors. However, its role in GC peritoneal metastasis remains unclear. In our study, transcriptome results suggested that NSUN2 expression was significantly upregulated in PM. And patients with high NSUN2 expression of PM predicted a worse prognosis. Mechanistically, NSUN2 regulates ORAI2 mRNA stability by m5C modification, thereby promoting ORAI2 expression and further promoting peritoneal metastasis and colonization of GC. YBX1 acts as a "reader" by binding to the ORAI2 m5C modification site. Following the uptake of fatty acids from omental adipocytes by GC cells, the transcription factor E2F1 was upregulated, which further promoted the expression of NSUN2 through cis-element. Briefly, these results revealed that peritoneal adipocytes provide fatty acid for GC cells, thus contributing to the elevation of E2F1 and NSUN2 through AMPK pathway, and upregulated NSUN2 activates the key gene ORAI2 through m5C modification, thereby promoting peritoneal metastasis and colonization of gastric cancer.

DOI: https://doi.org/10.1038/s41388-023-02707-5
Cancer Immunol Res. 2023 May 02. pii: CIR-22-0814. [Epub ahead of print]

KRT17 promotes T-lymphocyte infiltration through the YTHDF2-CXCL10 axis in colorectal cancer.

Wenfeng Liang, Huashan Liu, Ziwei Zeng, Zhenxing Liang, Hao Xie, Wenxin Li, Li Xiong, Zhihang Liu, Mian Chen, Haiqing Jie, Xiaobin Zheng, Liang Huang, Liang Kang.

Poor infiltration of T lymphocytes has been regarded as a crucial mechanism of tumor immune escape. Here, we demonstrate a protective role of KRT17 in colorectal cancer (CRC), where KRT17 reversed the tumor immunosuppressive microenvironment by increasing T-lymphocyte infiltration. High-throughput RNA sequencing suggested KRT17 was significantly upregulated in deficient mismatch repair (dMMR) tumors compared to proficient mismatch repair (pMMR) tumors. In a CRC cohort of 446 cases, KRT17 expression positively correlated with better clinical outcomes. Krt17 overexpression decreased xenograft tumor growth in immune-competent mice. T-cell depletion in a murine model showed that the presence of T lymphocytes was necessary for Krt17-mediated disruption of tumorigenesis. Mass spectrometry and co-immunoprecipitation assays suggested KRT17 caused YTHDF2 degradation through the ubiquitin-proteasome system. Through high-throughput RNA immunoprecipitation sequencing, we found CXCL10 was the target gene of the N6-methyladenosine (m6A) 'reader' YTHDF2. KRT17 synergized with anti-PD-1 for better tumor control in an immunotherapy-resistant murine model. In a cohort of patients with CRC receiving pembrolizumab, high KRT17 expression was found within the tumors of responders. Collectively, we elucidated a critical role of KRT17 in CRC to prevent immune escape. These findings present new insights into potential therapeutic strategies and effective markers of immunotherapy reactivity against pMMR tumors.

DOI: https://doi.org/10.1158/2326-6066.CIR-22-0814
Immunol Lett. 2023 Apr 28. pii: S0165-2478(23)00068-8. [Epub ahead of print]

Mettl14 sustains FOXP3 expression to promote the differentiation and functions of induced-regulatory T cells via the mTOR signaling pathway.

Yanzhuo Liu, Yinglin Yuan, Zili Zhou, Xiaomei Jiang, Shu He, Fan Wei, Yuanyuan Cui, Lu Yang, Gaoping Zhao.

  Induced regulatory T cell (iTregs) can be generated in vitro. Thus, iTregs-based therapeutics are receiving increased attention for their potential to treat autoimmune diseases and prevent transplant rejection. However, iTregs fail to maintain FoxP3 expression and suppressive activity, which limits their clinical application. Increasing lines of evidence suggest that methyltransferase-like 14 (METTL14), a critical component of the m6A writer complex, regulates the stability and function of the Treg cells. However, beyond meeting the epigenetic modification of Treg cells, whether Mettl14 plays a role in the fate determination of iTregs is unclear. Here, we systemically investigated the potential function of METTL14 in iTregs differentiation and regulatory activity. In our study, iTregs were generated from CD4+ naïve T cells under iTreg-polarizing conditions, we found that the expression of METTL14 was increased in iTregs compared with CD4+ naïve T cells. Subsequently, the expression of METTL14 was knocked down by siRNA-METTL14 interference in CD4+ naïve T cells and cultured under iTreg-polarizing conditions. According to the results, Mettl14 deficiency resulted in the disruption of iTregs differentiation evidenced by the limited FoxP3 expression. Meanwhile, inflammatory cytokines such as IFN-γ and IL-17a were upregulated in cultured iTregs. We next determined the functional change in METTL14-deficient iTregs. The results of the colitis development in Rag1-/- mice and CFSE assays revealed that loss of METTL14 significantly compromised the suppressive function of iTregs in vivo and in vitro. We further checked the altered signaling pathway in METTL14-deficient iTregs. We found that reduced METTL14 leads to activation of the mTOR pathway with increased p-mTOR and p-p70S6K, which are known to modulate the suppressive function of iTregs. In conclusion, our study revealed that Mettl14 plays a critical role in the development and suppressive function of iTregs in vitro and could thus serve as a regulatory element for stabilizing iTregs in cell-based therapy.

Keywords:  N6-methyladenosine; differential; iTregs; mTOR; suppressive function

DOI:  https://doi.org/10.1016/j.imlet.2023.04.008
Mol Cell Biochem. 2023 Apr 29.

LncRNA GAS5 regulated by FTO-mediated m6A demethylation promotes autophagic cell death in NSCLC by targeting UPF1/BRD4 axis.

Yihui Fu, Lirong Liu, Haihong Wu, Yamei Zheng, Huijuan Zhan, Liang Li.

  Long non-coding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5) has been shown to be a regulator for many cancers, including non-small cell lung cancer (NSCLC). Therefore, its role and mechanism in the process of NSCLC deserve to be further revealed. The expression levels of GAS5, fat mass and obesity-associated protein (FTO) and bromodomain-containing protein 4 (BRD4) were detected by quantitative real-time PCR. Western blot analysis was used to examine the protein expression of FTO, BRD4, up-frameshift protein 1 (UPF1) and autophagy-related markers. Methylated RNA immunoprecipitation was used to assess the m6A level of GAS5 regulated by FTO. Cell proliferation and apoptosis were determined using MTT assay, EdU assay and flow cytometry. Autophagy ability was assessed by immunofluorescence staining and transmission electron microscope. Xenograft tumor model was constructed to explore the effects of FTO and GAS5 on NSCLC tumor growth in vivo. The interaction between UPF1 and GAS5 or BRD4 was confirmed by pull-down assay, RIP assay, dual-luciferase reporter assay, and chromatin immunoprecipitation. Fluorescent in situ hybridization was used to analyze the co-localization of GAS5 and UPF1. Actinomycin D treatment was employed to evaluate BRD4 mRNA stability. GAS5 was downregulated in NSCLC tissues and was associated with poor prognosis in NSCLC patients. FTO was highly expressed in NSCLC, and it inhibited GAS5 expression by reducing GAS5 m6A methylation level. GAS5 suppressed by FTO could promote the autophagic death of NSCLC cells in vitro and inhibit NSCLC tumor growth in vivo. In addition, GAS5 was able to interact with UPF1 to reduce the mRNA stability of BRD4. Knockdown of BRD4 reversed the inhibition of GAS5 or UPF1 silencing on the autophagic cell death of NSCLC. The findings of the study showed that lncRNA GAS5 mediated by FTO could contribute to the autophagic cell death of NSCLC by interacting with UPF1 to reduce BRD4 mRNA stability, suggesting that GAS5 might be a vital therapy target for NSCLC progression.

Keywords:  BRD4; FTO; GAS5; Non-small cell lung cancer; UPF1

DOI:  https://doi.org/10.1007/s11010-023-04748-6
Genome Biol. 2023 04 30. 24(1): 103

m6A readers ECT2/ECT3/ECT4 enhance mRNA stability through direct recruitment of the poly(A) binding proteins in Arabidopsis.

Peizhe Song, Lianhuan Wei, Zixin Chen, Zhihe Cai, Qiang Lu, Chunling Wang, Enlin Tian, Guifang Jia.

   BACKGROUND: RNA N6-methyladenosine (m6A) modification is critical for plant growth and crop yield. m6A reader proteins can recognize m6A modifications to facilitate the functions of m6A in gene regulation. ECT2, ECT3, and ECT4 are m6A readers that are known to redundantly regulate trichome branching and leaf growth, but their molecular functions remain unclear.
RESULTS: Here, we show that ECT2, ECT3, and ECT4 directly interact with each other in the cytoplasm and perform genetically redundant functions in abscisic acid (ABA) response regulation during seed germination and post-germination growth. We reveal that ECT2/ECT3/ECT4 promote the stabilization of their targeted m6A-modified mRNAs, but have no function in alternative polyadenylation and translation. We find that ECT2 directly interacts with the poly(A) binding proteins, PAB2 and PAB4, and maintains the stabilization of m6A-modified mRNAs. Disruption of ECT2/ECT3/ECT4 destabilizes mRNAs of ABA signaling-related genes, thereby promoting the accumulation of ABI5 and leading to ABA hypersensitivity.
CONCLUSION: Our study reveals a unified functional model of m6A mediated by m6A readers in plants. In this model, ECT2/ECT3/ECT4 promote stabilization of their target mRNAs in the cytoplasm.

Keywords:  N 6-Methyladenosine (m6A); PAB proteins; Stability; m6A readers ECT2/ECT3/ECT4

DOI:  https://doi.org/10.1186/s13059-023-02947-4
Hematol Oncol. 2023 May 01.

LncRNA FEZF1-AS1 accelerates multiple myeloma progression by regulating IGF2BP1/BZW2 signaling.

Xingxing Long, Feng Wen, Junjun Li, Xiaoqing Huang.

  Multiple myeloma (MM) is the second largest hematological tumor with clonal proliferation of malignant plasma cells. Growing reports have revealed that the dysregulation of long non-coding RNA (lncRNA) is involved in the MM progression. Nevertheless, lncRNA FEZF1 antisense RNA 1 (FEZF1-AS1) remain not deeply explored. The RNA transcripts and protein level of MM-associated molecule were measured by quantitative real-time polymerase chain reaction or western blot assays, respectively. The clinical correlation was analyzed by Pearson analysis. Molecular interactions among lncRNA FEZF1-AS1, basic leucine zipper and W2 domain 2 (BZW2) and insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) were verified by RNA immunoprecipitation and RNA pull-down assays. The survival of MM cells was detected by cell counting kit-8 and flow cytometry assays. Xenograft tumor in vivo was performed to assess tumor growth. The RNA transcripts of lncRNA FEZF1-AS1, BZW2 and IGF2BP1 were upregulated in MM samples compared to those in healthy donors. Knockdown of lncRNA FEZF1-AS1 could inhibit the proliferation and induce cell apoptosis in vitro and in vivo. Besides, lncRNA FEZF1-AS1 could maintain the stability of BZW2 mRNA by interacting IGF2BP1. Moreover, BZW2 silence also downregulated the proliferation but enhanced apoptosis of MM cells, while BZW2 overexpression had an opposite role, which dramatically reversed the regulatory roles of lncRNA FEZF1-AS1. Altogether, lncRNA FEZF1-AS1 facilitated MM development by regulating IGF2BP1/BZW2 signaling, suggesting that lncRNA FEZF1-AS1 might be a candidate for MM treatment.

Keywords:  BZW2; IGF2BP1; lncRNA FEZF1-AS1; multiple myeloma

DOI:  https://doi.org/10.1002/hon.3157
J Pathol Clin Res. 2023 May 06.

KLF2 inhibits colorectal cancer progression and metastasis by inducing ferroptosis via the PI3K/AKT signaling pathway.

Jia Li, Ji Ling Jiang, Yi Mei Chen, Wei Qi Lu.

  Krüppel-like factor 2 (KLF2) belongs to the zinc finger family and is thought to be a tumor suppressor gene due to its low expression in various cancer types. However, its functional role and molecular pathway involvement in colorectal cancer (CRC) are not well defined. Herein, we investigated the potential mechanism of KLF2 in CRC cell invasion, migration, and epithelial-mesenchymal transition (EMT). We utilized the TCGA and GEPIA databases to analyze the expression of KLF2 in CRC patients and its correlation with different CRC stages and CRC prognosis. RT-PCR, western blot, and immunohistochemistry assays were used to measure KLF2 expression. Gain-of-function assays were performed to evaluate the role of KLF2 in CRC progression. Moreover, mechanistic experiments were conducted to investigate the molecular mechanism and involved signaling pathways regulated by KLF2. Additionally, we also conducted a xenograft tumor assay to evaluate the role of KLF2 in tumorigenesis. KLF2 expression was low in CRC patient tissues and cell lines, and low expression of KLF2 was associated with poor CRC prognosis. Remarkably, overexpressing KLF2 significantly inhibited the invasion, migration, and EMT capabilities of CRC cells, and tumor growth in xenografts. Mechanistically, KLF2 overexpression induced ferroptosis in CRC cells by regulating glutathione peroxidase 4 expression. Moreover, this KLF2-dependent ferroptosis in CRC cells was mediated by inhibiting the PI3K/AKT signaling pathway that resulted in the suppression of invasion, migration, and EMT of CRC cells. We report for the first time that KLF2 acts as a tumor suppressor in CRC by inducing ferroptosis via inhibiting the PI3K/AKT signaling pathway, thus providing a new direction for CRC prognosis assessment and targeted therapy.

Keywords:  KLF2; colorectal cancer; ferroptosis; metastasis; pathway

DOI:  https://doi.org/10.1002/cjp2.325
Aging (Albany NY). 2023 May 03. 15

m6A-mediated upregulation of lncRNA-AC026356.1 promotes cancer stem cell maintenance in lung adenocarcinoma via activating Wnt signaling pathway.

Zhen Zhang, Xiaoning Tan, Ruoxia Wu, Tianhao Deng, Huazhong Wang, Xiulin Jiang, Puhua Zeng, Junqi Tang.

  LncRNA plays a pivotal role in the stemness and drug resistance of lung cancer. Here, we found that lncRNA-AC026356.1 was upregulated in stem spheres and chemo-resistant lung cancer cells. Our fish assay also shows that AC026356.1 was predominantly located in the cytoplasm of lung cancer cells and does not have protein-coding potential. Silencing AC026356.1 significantly inhibited proliferation and migration but increased apoptosis in A549-cisplatin (DDP) cells. Additionally, IGF2BP2 and the lncRNA-AC026356.1 positively regulated the proliferation and stemness of stem-like lung cancer cells. Further mechanistic investigation revealed that METTL14/IGF2BP2-mediated m6A modification and stabilization of the AC026356.1 RNA. Functional analysis corroborated that AC026356.1 acted as a downstream target of METTL14/IGF2BP2 and AC026356.1 silencing could block the oncogenicity of lung cancer stem-like cells. AC026356.1 expression was correlated with immune cell infiltration and T cell exhaustion. Compared with paired adjacent normal tissues, lung cancer specimens exhibited consistently upregulated METTL14/IGF2BP2/AC026356.1. M6A-modified METTL14/IGF2BP2/AC026356.1 loop may serve as a potential therapeutic target and prognostic predictor for lung cancer therapy and diagnosis in the clinic.

Keywords:  T cell exhaustion; cancer stem cell; cisplatin resistance; immune checkpoint inhibitors; immunotherapy; lncRNA

DOI:  https://doi.org/10.18632/aging.204689
BMC Urol. 2023 May 03. 23(1): 82

NUF2 is correlated with a poor prognosis and immune infiltration in clear cell renal cell carcinoma.

Baishu Zheng, Shunde Wang, Xiaoyu Yuan, Junyong Zhang, Zhongjie Shen, Chengguo Ge.

   BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is one of the most common malignancies. Recently, immunotherapy has been considered a promising treatment for metastatic ccRCC. NUF2 is a crucial component of the Ndc80 complex. NUF2 can stabilize microtubule attachment and is closely related to cell apoptosis and proliferation. This research is dedicated to investigating the role of NUF2 in ccRCC and the possible mechanisms.
METHODS: First, analysis of NUF2 mRNA expression levels in ccRCC and normal tissues by The Cancer Genome Atlas (TCGA) database and further verified by analysis of independent multiple microarray data sets in the Gene Expression Omnibus (GEO) database. Moreover, we evaluated and identified correlations between NUF2 expression, clinicopathologic variable, and overall survival (OS) in ccRCC by various methods. We investigated the relationship between NUF2 and tumor immune infiltration and the expression of corresponding immune cell markers via the Gene Expression Profiling Interactive Analysis (GEPIA) and Tumor Immune Estimation Resource (TIMER) databases. Then, we performed functional enrichment analysis of NUF2 co-expressed genes using R software and protein-protein interactions (PPIs) using the search tool used to retrieve interacting genes/proteins (STRING) databases.
RESULTS: We discovered that NUF2 mRNA expression was upregulated in ccRCC tissues and was associated with sex, grade, pathological stage, lymph node metastasis, and worse prognosis. In addition, NUF2 was positively linked to tumor immune cells in ccRCC. Moreover, NUF2 was closely related to genetic markers of different immune cells. Finally, functional enrichment and protein-protein interaction (PPI) analysis suggested that NUF2 and its closely related genes may be involved in the regulation of the cell cycle and mitosis. Our results suggested that NUF2 is correlated with a poor prognosis and immune infiltration in ccRCC.

Keywords:  Clear cell renal cell carcinoma; Immune cells; NUF2; Tumor immune infiltration

DOI:  https://doi.org/10.1186/s12894-023-01258-x
Nucleic Acids Res. 2023 May 01. pii: gkad300. [Epub ahead of print]

Low RNA stability signifies increased post-transcriptional regulation of cell identity genes.

Yanqiang Li, Yang Yi, Jie Lv, Xinlei Gao, Yang Yu, Sahana Suresh Babu, Ivone Bruno, Dongyu Zhao, Bo Xia, Weiqun Peng, Jun Zhu, Hong Chen, Lili Zhang, Qi Cao, Kaifu Chen.

Cell identity genes are distinct from other genes with respect to the epigenetic mechanisms to activate their transcription, e.g. by super-enhancers and broad H3K4me3 domains. However, it remains unclear whether their post-transcriptional regulation is also unique. We performed a systematic analysis of transcriptome-wide RNA stability in nine cell types and found that unstable transcripts were enriched in cell identity-related pathways while stable transcripts were enriched in housekeeping pathways. Joint analyses of RNA stability and chromatin state revealed significant enrichment of super-enhancers and broad H3K4me3 domains at the gene loci of unstable transcripts. Intriguingly, the RNA m6A methyltransferase, METTL3, preferentially binds to chromatin at super-enhancers, broad H3K4me3 domains and their associated genes. METTL3 binding intensity is positively correlated with RNA m6A methylation and negatively correlated with RNA stability of cell identity genes, probably due to co-transcriptional m6A modifications promoting RNA decay. Nanopore direct RNA-sequencing showed that METTL3 knockdown has a stronger effect on RNA m6A and mRNA stability for cell identity genes. Our data suggest a run-and-brake model, where cell identity genes undergo both frequent transcription and fast RNA decay to achieve precise regulation of RNA expression.

DOI: https://doi.org/10.1093/nar/gkad300
bioRxiv. 2023 Apr 24. pii: 2023.04.21.537835. [Epub ahead of print]

Myocardial IGF2R is a critical mediator of inflammation and fibrosis after ischemia-reperfusion injury.

Zhichao Wu, Wei Huang, Xingyu He, Suchandrima Dutta, Christian Paul, Guo-Chang Fan, Onur Kanisicak, Meifeng Xu, Jialiang Liang, Yigang Wang.

Ischemia-reperfusion (I/R) injury is a common occurrence in various surgical procedures used to treat heart diseases. However, the role of insulin-like growth factor 2 receptor (IGF2R) during the process of myocardial I/R remains unclear. Therefore, this study aims to investigate the expression, distribution, and functionality of IGF2R in various I/R-associated models (such as reoxygenation, revascularization, and heart transplant). Loss-of-function studies (including myocardial conditional knockout and CRISPR interference) were performed to clarify the role of IGF2R in I/R injuries. Following hypoxia, IGF2R expression increased, but this effect was reversed upon restoration of oxygen levels. Loss of myocardial IGF2R was found to enhance the cardiac contractile functions, and reduced cell infiltration or cardiac fibrosis of I/R mouse models compared to the genotype control. CRISPR-inhibition of IGF2R decreased cell apoptotic death under hypoxia. RNA sequencing analysis indicated that myocardial IGF2R played a critical role in regulating the inflammatory response, innate immune response, and apoptotic process following I/R. Integrated analysis of the mRNA profiling, pulldown assays, and mass spectrometry identified granulocyte-specific factors as potential targets of myocardial IGF2R in the injured heart. In conclusion, myocardial IGF2R emerges as a promising therapeutic target to ameliorate inflammation or fibrosis following I/R injuries.

DOI: https://doi.org/10.1101/2023.04.21.537835
J Oncol. 2023 ;2023 1105042

SH3BGRL Suppresses Liver Tumor Progression through Enhanced ATG5-Dependent Autophagy.

Abdulmomen Ali Mohammed Saleh, Farhan Haider, Haimei Lv, Bin Liu, Jing Xiao, Mingming Zhang, Yuzhen Zheng, Shulan Yang, Haihe Wang.

SH3BGRL, an adaptor protein, is upregulated in breast cancers and indicates its tumorigenic role. But the function of SH3BGRL in other types of cancers is largely unknown. Here, we modulate SH3BGRL expression level in two liver cancer cells and conduct both in vitro and in vivo analyses of SH3BGRL in cell proliferation and tumorigenesis. Results demonstrate that SH3BGRL notably inhibits cell proliferation and arrests the cell cycle in both LO2 and HepG2 cells. Molecularly, SH3BGRL upregulates the expression of ATG5 from proteasome degradation as well as the inhibitions of Src activation and its downstream ERK and AKT signaling pathways, which eventually enhance autophagic cell death. The xenograft mouse model reveals that SH3BGRL overexpression can efficiently suppress tumorigenesis in vivo, while the additional silencing ATG5 in SH3BGRL-overexpressing cells attenuates the inhibitory effect of SH3BGRL on both hepatic tumor cell proliferation and tumorigenicity in vivo. The relevance of SH3BGRL downregulation in liver cancers and their progression is validated based on the large-scale tumor data. Taken together, our results clarify the suppressive role of SH3BGRL in tumorigenesis of liver cancer, which would be of help to the diagnosis of liver cancer, while either promoting the autophagy of liver cancer cells or inhibiting the downstream signaling induced from SH3BGRL downregulation would be a promising therapy.

DOI: https://doi.org/10.1155/2023/1105042