Noncoding RNA NR Research Articles

Long Noncoding RNA NR_030777 Alleviates Cobalt Nanoparticles-Induced Neurodegenerative Damage by Promoting Autophagosome-Lysosome Fusion.

Potential exposure to cobalt nanoparticles (CoNPs) occurs in various fields, including hard alloy industrial production, the increasing use of new energy lithium-ion batteries, and millions of patients with metal-on-metal joint prostheses. Evidence from human, animal, and in vitro experiments suggests a close relationship between CoNPs and neurotoxicity. However, a systematic assessment of central nervous system (CNS) impairment due to CoNPs exposure and the underlying molecular mechanisms is lacking. In this study, we found that CoNPs induced neurodegenerative damage both in vivo and in vitro, including cognitive impairment, β-amyloid deposition and Tau hyperphosphorylation. CoNPs promoted the formation of autophagosomes and impeding autophagosomal-lysosomal fusion in vivo and in vitro, leading to toxic protein accumulation. Moreover, CoNPs exposure reduced the level of transcription factor EB (TFEB) and the abundance of lysosome, causing a blockage in autophagosomal-lysosomal fusion. Interestingly, overexpression of long noncoding RNA NR_030777 mitigated CoNPs-induced neurodegenerative damage in both in vivo and in vitro models. Fluorescence in situ hybridization assay revealed that NR_030777 directly binds and stabilizes TFEB mRNA, alleviating the blockage of autophagosomal-lysosomal fusion and ultimately restoring neurodegeneration induced by CoNPs in vivo and in vitro. In summary, our study demonstrates that autophagic dysfunction is the main toxic mechanism of neurodegeneration upon CoNPs exposure and NR_030777 plays a crucial role in CoNPs-induced autophagic dysfunction. Additionally, the proposed adverse outcome pathway contributes to a better understanding of CNS toxicity assessment of CoNPs.

Negative regulation of HBG1/2 expression through S6K by long noncoding RNA NR_120526.

High levels of fetal hemoglobin (HbF) may alleviate clinical symptoms in patients with β-thalassemia. A previous study showed that the long noncoding RNA NR_120526 (lncRNA NR_120526) might be involved in regulating HbF levels (HBG1/2 gene expression). However, the function and mechanism by which NR_120526 regulates HbF expression remains unknown. Here, we investigated the effect of NR_120526 on HbF and its mechanism so as to provide an experimental basis for treating patients with β-thalassemia. Chromatin isolation by RNA purification-mass spectrometry (ChIRP-MS) assay, database query, and bioinformatics analysis were performed to explore the proteins that specifically bind to NR_120526 and their interactions. Chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) were used to determine whether NR_120526 directly regulates the expression of HBG1/2. The NR_120526 gene was knocked out (KO) using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology in K562 cells. Finally, quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to detect the messenger RNA (mRNA) and protein expressions of HBG1/2, ribosomal protein S6 kinase B1 (RPS6KB1, S6K), and Ras homologous family member A (RhoA), respectively. We found that NR_120526 interacts with ILF2, ILF3, and S6K. However, ILF2/ILF3 bound to NR_120526 did not interact with HBG1/2, suggesting that NR_120526 may regulate HBG1/2 expression indirectly. The qRT-PCR results showed no statistical difference in the mRNA expression levels of HBG1/2, S6K, and RhoA between the NR_120526-KO group and negative control (NC) group (P>0.05). However, Western blot results showed a significant increase in the protein levels of HBG1/2, S6K, and RhoA in the KO group (P<0.05). It was found that NR_120526 inhibited S6K, thereby downregulating RhoA and leading to decreased HBG1/2 expression. LncRNA NR_120526 negatively regulates the expression of HBG1/2 through S6K. These new findings provide mechanistic insights into the regulation of HbF and offer potential therapeutic targets for precision medicine in patients with β-thalassemia.

A host cell long noncoding RNA NR_033736 regulates type I interferon-mediated gene transcription and modulates intestinal epithelial anti-Cryptosporidium defense.

The gastrointestinal epithelium guides the immune system to differentiate between commensal and pathogenic microbiota, which relies on intimate links with the type I IFN signal pathway. Epithelial cells along the epithelium provide the front line of host defense against pathogen infection in the gastrointestinal tract. Increasing evidence supports the regulatory potential of long noncoding RNAs (lncRNAs) in immune defense but their role in regulating intestinal epithelial antimicrobial responses is still unclear. Cryptosporidium, a protozoan parasite that infects intestinal epithelial cells, is an important opportunistic pathogen in AIDS patients and a common cause of diarrhea in young children in developing countries. Recent advances in Cryptosporidium research have revealed a strong type I IFN response in infected intestinal epithelial cells. We previously identified a panel of host cell lncRNAs that are upregulated in murine intestinal epithelial cells following microbial challenge. One of these lncRNAs, NR_033736, is upregulated in intestinal epithelial cells following Cryptosporidium infection and displays a significant suppressive effect on type I IFN-controlled gene transcription in infected host cells. NR_033736 can be assembled into the ISGF3 complex and suppresses type I IFN-mediated gene transcription. Interestingly, upregulation of NR_033736 itself is triggered by the type I IFN signaling. Moreover, NR_033736 modulates epithelial anti-Cryptosporidium defense. Our data suggest that upregulation of NR_033736 provides negative feedback regulation of type I IFN signaling through suppression of type I IFN-controlled gene transcription, and consequently, contributing to fine-tuning of epithelial innate defense against microbial infection.

Identification of downstream targets and signaling pathways of long non-coding RNA NR_002794 in human trophoblast cells

ABSTRACT Preeclampsia (PE) is a huge threat to pregnant women. Our previous study demonstrated that long non-coding RNA (lncRNA) NR_002794 was highly expressed in placentas of PE patients and could regulate the phenotypes of trophoblast cells. However, the downstream regulatory mechanisms of NR_002794 remain unknown. In this text, some potential downstream targets or signaling pathways of NR_002794 were identified through RNA sequencing (RNA-seq) and bioinformatics analysis in SWAN71 trophoblast cells. Western blot assay demonstrated that NR_002794 inactivated protein kinase B (AKT) and extracellular signal-regulated kinase 1/2 (ERK1/2) pathways and activated cell apoptotic signaling in SWAN71 cells. Both RNA-seq and reverse transcription-quantitative PCR (RT-qPCR) outcomes showed that NR_002794 up-regulation could notably inhibit the expression of C-C motif chemokine ligand 4 like 2 (CCL4L2), interleukin 15 receptor subunit alpha (IL15RA), interleukin 32 (IL32), and tyrosine kinase with immunoglobulin-like and EGF-like domains 1 (TIE1), while NR_002794 knockdown induced these gene expressions in SWAN71 cells. CCK-8, BrdU, Transwell, wound healing, and flow cytometry analyses showed that NR_002794 inhibited cell proliferation and migration and induced cell apoptosis through down-regulating TIE1 in SWAN71 cells. In conclusion, lncRNA NR_002794 could exert its functions by regulating AKT and ERK1/2 pathways and TIE1 expression in human trophoblast cells.

LncRNA NR_027471 Functions as a ceRNA for miRNA-8055 Leading to Suppression of Osteosarcoma by Regulating the Expression of TP53INP1.

Osteosarcoma is a malignancy with high aggressiveness and poor prognosis, which occurs mainly in children. The therapeutic strategy against osteosarcoma includes surgery combined with chemotherapy and radiotherapy. Although the treatment of osteosarcoma has been improved in recent years, there is a large proportion of patients with incurable osteosarcoma. Investigation of the mechanism of osteosarcoma progression would be of great help in discovering therapeutic targets for this disease. Long non-coding RNAs play critical roles in the pathogenesis of different types of cancer. The current study showed that long non-coding RNA NR_027471 was downregulated in osteosarcoma cells. In vitro and in vivo studies indicated that upregulation of NR_027471 impeded the viability, proliferation, and invasion of osteosarcoma, as well as induced cell cycle arrest at G1. In addition, binding of miR-8055 to NR_027471 was demonstrated, thereby influencing the expression of tumor protein p53 inducible nuclear protein 1 (TP53INP1). Knockdown of NR_027471 promoted epithelial–mesenchymal transition by inhibiting E-cadherin and increasing the expression of zinc finger E-box-binding homeobox 1 (ZEB1), Snail, and fibronectin. These results suggested that overexpression of NR_027471 upregulated TP53INP1 by sponging to miR-8055, leading to suppression of osteosarcoma cell proliferation and progression.

The long noncoding RNA NR_045363 involves cardiomyocyte apoptosis and cardiac repair via p53 signal pathway.

Long noncoding RNAs (lncRNAs) can participate in various biological behaviors, including regulating cell differentiation, proliferation, and apoptosis. The investigators have previously confirmed that highly conserved lncRNA NR_045363 controls cardiomyocyte (CM) proliferation and cardiac repair. The present study investigates the effects of NR_045363 on CM apoptosis. Seven-day-old mice were subjected to permanent left anterior descending coronary artery ligation (LAD), and the NR_045363 expression was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). The expression of NR_045363 in the MI group significantly exceeded the Sham group during the first week after the operation. The NR_045363 expression was knocked down in primary cultured CMs using an NR_045363-targeting lncRNA Smart silencer, and the apoptosis of CMs was analyzed by terminal-deoxynucleoitidyl transferase mediated nick end labeling and Annexin-V/PI double staining. These present results indicate that the NR_045363 knockdown significantly promoted the apoptosis of CMs. In order to investigate the underlying mechanism, RNA-sequencing (RNA-seq) was performed, and ingenuity pathway analysis (IPA) was used to analyze the RNA-seq results. The RNA-seq data revealed that a total of 2,291 genes were upregulated or downregulated in NR_045363 knockdown CMs, and the IPA analysis indicated that tumor protein 53 (p53) was the upstream regulator. In vivo, the NR_045363 overexpression through the AAV9 system improved the heart function after MI in 7-day-old mice and inhibited the CM apoptosis. These data suggest that NR_045363 is involved in CM apoptosis and that NR_045363 overexpression exerts positive effects on cardiac repair by alleviating CM apoptosis through the inhibition of the p53 pathway.

Abstract 1824: Long noncoding RNA NR_027340 promotes head and neck cancer cell proliferation and glycolysis via PFKP

Abstract Introduction: Long noncoding RNAs (lncRNAs) play crucial roles in various cancers. Recent study has indicated that lncRNA NR_027340 was significantly upregulated in head and neck cancer (HNC). However, its multifaceted roles in HNC are unclear. Here we demonstrate the role of lncRNA NR_027340 in HNC cell proliferation and glycolysis. Methods: qRT-PCR was used to detect lncRNA NR_027340 expression in HNC tissues, and the correlations between lncRNA NR_027340 and clinicopathologic features were then analyzed. RNA pull-down and mass spectrometry analysis were carried out to explore the lncRNA NR_027340-bound proteins. RNA immunoprecipitation (RIP) was used to confirm the association between lncRNA and relevant proteins. lncRNA NR_027340 knockout cell lines were generated using the CRISPR/Cas9. Colony formation, cell cycle analysis and flow cytometry were used to measure the proliferation of HNC cells. The production of lactate was observed to evaluate the glycolysis of HNC cells. Results: We found lncRNA NR_027340 was significantly up-regulated in HNCs and the elevated lncRNA NR_027340 was correlated with poor clinical prognosis. Mechanistically, we identified PFKP as the protein partner of lncRNA NR_027340 in the cytoplasm. In addition, we found that both lncRNA NR_027340 and PFKP depletion reduced the cell proliferation and glycolysis in vitro. The ingenuity pathway analysis and Western blot assay indicated that lncRNA NR_027340 targeted PFKP to regulate cell proliferation and glycolysis in HNC. Conclusion: These findings reveal a functional role for lncRNA NR_027340 in altering glycolysis and promoting proliferation in HNC cells, suggesting that lncRNA NR_027340 might serve as a prognostic and diagnostic indicator as well as a potential therapeutic target. Citation Format: Yifan Yang, Ru Wang, Ling Feng, Qian Shi, Chen Tan, Jugao Fang. Long noncoding RNA NR_027340 promotes head and neck cancer cell proliferation and glycolysis via PFKP [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1824.

A long noncoding RNA NR_045363 controls cardiomyocyte proliferation and cardiac repair

Long noncoding RNAs (lncRNAs) play important roles in the regulation of genes involved in cell proliferation. We have previously sought to more globally understand the differences of lncRNA expression between human fetal heart and adult heart to identify some functional lncRNAs which involve in the process of heart repair. We found that a highly conserved long noncoding RNA NR_045363 was mainly expressed in cardiomyocytes and rarely in non-cardiomyocytes. NR_045363 overexpression in 7-day-old mice heart could improve cardiac function and stimulate cardiomyocyte proliferation after myocardial infarction. Furthermore, NR_045363 knockdown inhibited proliferation of primary embryonic cardiomyocytes, while NR_045363 overexpression enhanced DNA synthesis and cytokinesis in neonatal cardiomyocytes in vitro. Mechanistic analysis revealed that NR_045363 promoted cardiomyocyte proliferation through interaction with miR-216a, which regulated the JAK2-STAT3 pathway. Our results showed that NR_045363 is a potent lncRNA modulator essential for cardiomyocyte proliferation.

Knockdown of Long Noncoding RNA NR_026689 Inhibits Proliferation and Invasion and Increases Apoptosis in Ovarian Carcinoma HO-8910PM Cells.

Ovarian cancer is one of the leading causes of gynecological cancer-related deaths worldwide. We investigated the role of a newly discovered long noncoding RNA, NR_026689, in cell proliferation, metastasis, and apoptosis in ovarian cancer cells. Our results showed that NR_026689 was overexpressed in both clinical ovarian cancer patients and cultured ovarian cancer cells. Knockdown of NR_026689 in HO-8910PM cells significantly decreased the cell proliferative rate and the ability to form colonies. Transwell assays revealed that depletion of NR_026689 suppressed cell migration ability by 68% and cell invasive capacity by 71% in HO-8910PM cells. Moreover, specific shRNAs against NR_026689 notably promoted cell apoptosis in HO-8910PM cells by upregulating the expression of proapoptotic proteins, including caspase 3, caspase 9, cytochrome C, and PARP. Our study suggests an oncogenic potential of NR_026689 in ovarian cancer and might provide novel clues for the diagnosis and treatment of ovarian cancer in the clinic.

Overexpression of long non-coding RNA NR_036575.1 contributes to the proliferation and migration of papillary thyroid cancer.

Current evidence suggests that the human genome produces a large number of non-coding RNAs, including microRNAs and long non-coding RNAs (lncRNAs). Generally, lncRNAs are defined as RNA transcripts longer than 200 nucleotides that are not transcribed into proteins. In recent years, lncRNAs have been reported to play oncogenic roles in tumourigenesis. However, minimal research has been performed on the expression and clinicopathological significance of lncRNAs in papillary thyroid cancer (PTC). In the present study, we investigated not only the expression and clinicopathological significance of a novel lncRNA, NR_036575.1, in PTC tissues and adjacent non-cancerous tissues but also its potential function in TPC1 cells. The expression levels of the lncRNA NR_036575.1 in 83 pairs of PTC tissues and adjacent non-cancerous tissues were detected via quantitative real-time polymerase chain reaction. The relationships between the expression levels and clinicopathological characteristics of the lncRNA NR_036575.1 were analysed. In addition, we established two receiver operating characteristic (ROC) curves to assess the diagnostic value of NR_036575.1 expression. Cell Counting Kit-8 and transwell assays were used to assess cell proliferation and migration, respectively. The expression levels of the lncRNA NR_036575.1 were significantly higher in PTC tissues than in adjacent non-cancerous tissues. High NR_036575.1 expression was associated with extrathyroidal extension (ETE) (P=0.011) and tumour size (P=0.006). The ROC curves indicated that NR_036575.1 could potentially serve as a biomarker for identifying PTC and related, non-cancerous diseases (sensitivity, 80.7%; specificity, 88%), as well as for differentiating between PTC with or without ETE (sensitivity, 57.8%; specificity, 86.7%). NR_036575.1 knock-down significantly inhibited the proliferation and migration of TPC1 cells. Our findings are the first to describe lncRNA NR_036575.1 overexpression in PTC. NR_036575.1 expression was associated with both ETE and tumour size. In addition, NR_036575.1 modulation could regulate TPC1 cell proliferation and migration. The results of our study suggest that NR_036575.1 could be applied as a potential biomarker and a novel therapeutic target for PTC patients.

Identification of a long non-coding RNA NR_026689 associated with lung carcinogenesis induced by NNK.

Long non-coding RNAs (lncRNA) are thought to be important epigenetic regulators involved in the development of a variety of cancers. Alterations in lncRNA expression are associated with exposure to chemical carcinogens. However, it is still unclear whether lncRNA expression during lung carcinogenesis is induced by chemical carcinogens. In this study, using NNK-induced rat lung cancer model established by our previous study, we determined the lncRNA expression profiles, and an alteration in lncRNA expression was observed in lung cancer tissues and blood in the NNK treatment group. Using quantitative reverse-transcription PCR (qRT-PCR), five differentially expressed lncRNAs were further detected and validated. We identified a novel lncRNA, NR_026689, which showed increased expression in lung cancer tissues induced by NNK and the alteration of lncRNA NR_026689 was specifically observed in lung tissue. The level of NR_026689 was determined and significantly increased in rat whole blood at the 10th and 20th week after NNK treatment to evaluate it as a potential early marker for lung cancer. Together, these findings suggest that lncRNA NR_026689 may be a potential early biomarker for lung cancer and is associated with lung carcinogenesis induced by NNK.

Long non-coding RNA NR_045623 and NR_028291 involved in benzene hematotoxicity in occupationally benzene-exposed workers

Benzene is an established human hematotoxicant and leukemogen. New insights into the pathogenesis of benzene hematotoxicity are urgently needed. Long non-coding RNA (lncRNA) widely participate in various physiological and pathological processes. It has been shown that lncRNA plays an important role in hematologic malignancy tumorigenesis. However, the expression and biological function of lncRNA during benzene hematotoxicity progress remain largely unknown. An integrated analysis of differentially expressed lncRNA and mRNA was performed to identify genes which were likely to be critical for benzene hematotoxicity through Microarray analysis. Dynamic gene network analysis of the differentially expressed lncRNA and mRNA was constructed and two main lncRNA (NR_045623 and NR_028291) were discovered and two key lncRNA subnets were involved in immune responses, hematopoiesis, B cell receptor signaling pathway and chronic myeloid leukemia. These findings suggested that NR_045623 and NR_028291 might be the key genes associated with benzene hematotoxicity.