Potential mechanisms of microRNA-141-3p to alleviate chronic inflammatory pain by downregulation of downstream target gene HMGB1: in vitro and in vivo studies

Abstract Background Glioma is considered one of the most common tumors and has a poor prognosis. Recently, microRNAs (miRNAs) have been reported to be strongly linked to various human tumors including glioma. In this study, we investigated a new anticancer miRNA, miR-346, to determine the effects and mechanism of miR-346 and its downstream target gene NFIB on tumors. Methods Lentivirus transfection, real-time PCR, western blotting, immunohistochemistry, cell proliferation assays, and mouse experiments were used to examine the relationship between miR-346 and its regulation of NFIB in glioma cells. Results The expression of miR-346 was downregulated in glioma cells. Overexpression of miR-346 arrested the cell cycle of glioma cells and inhibited their proliferation in vitro and in vivo. NFIB was a direct target of miR-346, whose expression was reduced by the miRNA. Overexpression of NFIB reversed all tested functions of miR-346. Conclusion miR-346 inhibited the growth of glioma cells by targeting NFIB and may be a new prognostic and diagnostic biomarker for glioma.

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GmBTB/POZ promotes the ubiquitination and degradation of LHP1 to regulate the response of soybean to Phytophthora sojae

Communications Biology ◽

10.1038/s42003-021-01907-7 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Chuanzhong Zhang ◽

Qun Cheng ◽

Huiyu Wang ◽

Hong Gao ◽

Xin Fang ◽

...

Keyword(s):

Root Rot ◽

Target Gene ◽

Nuclear Protein ◽

Wrky Transcription Factor ◽

Phytophthora Sojae ◽

Downstream Target ◽

Downstream Target Gene ◽

Transgenic Lines

AbstractPhytophthora sojae is a pathogen that causes stem and root rot in soybean (Glycine max [L.] Merr.). We previously demonstrated that GmBTB/POZ, a BTB/POZ domain-containing nuclear protein, enhances resistance to P. sojae in soybean, via a process that depends on salicylic acid (SA). Here, we demonstrate that GmBTB/POZ associates directly with soybean LIKE HETEROCHROMATIN PROTEIN1 (GmLHP1) in vitro and in vivo and promotes its ubiquitination and degradation. Both overexpression and RNA interference analysis of transgenic lines demonstrate that GmLHP1 negatively regulates the response of soybean to P. sojae by reducing SA levels and repressing GmPR1 expression. The WRKY transcription factor gene, GmWRKY40, a SA-induced gene in the SA signaling pathway, is targeted by GmLHP1, which represses its expression via at least two mechanisms (directly binding to its promoter and impairing SA accumulation). Furthermore, the nuclear localization of GmLHP1 is required for the GmLHP1-mediated negative regulation of immunity, SA levels and the suppression of GmWRKY40 expression. Finally, GmBTB/POZ releases GmLHP1-regulated GmWRKY40 suppression and increases resistance to P. sojae in GmLHP1-OE hairy roots. These findings uncover a regulatory mechanism by which GmBTB/POZ-GmLHP1 modulates resistance to P. sojae in soybean, likely by regulating the expression of downstream target gene GmWRKY40.

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Aryl Hydrocarbon Receptor: Its Regulation and Roles in Transformation and Tumorigenesis

Current Drug Targets ◽

10.2174/1389450120666181109092225 ◽

2019 ◽

Vol 20 (6) ◽

pp. 625-634 ◽

Cited By ~ 1

Author(s):

Xun Che ◽

Wei Dai

Keyword(s):

Target Gene ◽

Tumor Development ◽

Environmental Response ◽

Carcinogenic Effect ◽

Post Translational Modifications ◽

Downstream Target ◽

Aryl Hydrocarbon ◽

Downstream Target Gene ◽

Major Mediator

AhR is an environmental response gene that mediates cellular responses to a variety of xenobiotic compounds that frequently function as AhR ligands. Many AhR ligands are classified as carcinogens or pro-carcinogens. Thus, AhR itself acts as a major mediator of the carcinogenic effect of many xenobiotics in vivo. In this concise review, mechanisms by which AhR trans-activates downstream target gene expression, modulates immune responses, and mediates malignant transformation and tumor development are discussed. Moreover, activation of AhR by post-translational modifications and crosstalk with other transcription factors or signaling pathways are also summarized.

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Genetic evidence for the transcriptional-activating function of Homothorax during adult fly development

Development ◽

10.1242/dev.128.18.3405 ◽

2001 ◽

Vol 128 (18) ◽

pp. 3405-3413 ◽

Cited By ~ 4

Author(s):

Adi Inbal ◽

Naomi Halachmi ◽

Charna Dibner ◽

Dale Frank ◽

Adi Salzberg

Keyword(s):

Transcriptional Activity ◽

Target Genes ◽

Genetic Evidence ◽

In Vivo Studies ◽

Loss Of Function ◽

Native Form ◽

Downstream Target ◽

Activating Function

Homothorax (HTH) is a homeobox-containing protein, which plays multiple roles in the development of the embryo and the adult fly. HTH binds to the homeotic cofactor Extradenticle (EXD) and translocates it to the nucleus. Its function within the nucleus is less clear. It was shown, mainly by in vitro studies, that HTH can bind DNA as a part of ternary HTH/EXD/HOX complexes, but little is known about the transcription regulating function of HTH-containing complexes in the context of the developing fly. Here we present genetic evidence, from in vivo studies, for the transcriptional-activating function of HTH. The HTH protein was forced to act as a transcriptional repressor by fusing it to the Engrailed (EN) repression domain, or as a transcriptional activator, by fusing it to the VP16 activation domain, without perturbing its ability to translocate EXD to the nucleus. Expression of the repressing form of HTH in otherwise wild-type imaginal discs phenocopied hth loss of function. Thus, the repressing form was working as an antimorph, suggesting that normally HTH is required to activate the transcription of downstream target genes. This conclusion was further supported by the observation that the activating form of HTH caused typical hth gain-of-function phenotypes and could rescue hth loss-of-function phenotypes. Similar results were obtained with XMeis3, the Xenopus homologue of HTH, extending the known functional similarity between the two proteins. Competition experiments demonstrated that the repressing forms of HTH or XMeis3 worked as true antimorphs competing with the transcriptional activity of the native form of HTH. We also describe the phenotypic consequences of HTH antimorph activity in derivatives of the wing, labial and genital discs. Some of the described phenotypes, for example, a proboscis-to-leg transformation, were not previously associated with alterations in HTH activity. Observing the ability of HTH antimorphs to interfere with different developmental pathways may direct us to new targets of HTH. The HTH antimorph described in this work presents a new means by which the transcriptional activity of the endogenous HTH protein can be blocked in an inducible fashion in any desired cells or tissues without interfering with nuclear localization of EXD.

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MiR-30a-5p inhibits proliferation, migration and invasion of nasopharyngeal carcinoma cells by targeting NUCB2

Human & Experimental Toxicology ◽

10.1177/0960327121991913 ◽

2021 ◽

pp. 096032712199191

Author(s):

M Li ◽

Y Wang ◽

Q Zhao ◽

W Ma ◽

J Liu

Keyword(s):

Nasopharyngeal Carcinoma ◽

Tumor Growth ◽

Carcinoma Cells ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Binding Interaction ◽

Downstream Target ◽

Downstream Target Gene

Background: Nasopharyngeal carcinoma (NPC) is a malignant head and neck tumor arising in the nasopharynx. MicroRNAs (miRNAs) are elucidated to exert tumor-suppressing function in human cancers. Numerous studies have manifested that miR-30a-5p serves as an anti-oncogene in various cancers. Objective: To research the biological function and molecular mechanism of miR-30a-5p in NPC. Methods: The morphology of NPC tissues was revealed by H&E staining. Transwell and wound healing assays were applied to investigate the effects of miR-30a-5p on NPC cell migration. The binding interaction between miR-30a-5p and nucleobindin 2 (NUCB2) was identified by luciferase reporter assay. Xenograft nude mice were used to detect the influence of miR-30a-5p on NPC tumor growth. Results: MiR-30a-5p was downregulated in NPC tissues and cells. The overexpression ofmiR-30a-5p inhibited proliferation, migration and invasion abilities of NPC cells. Moreover, NUCB2 was revealed to be a downstream target gene of miR-30a-5p, and knockdown of NUCB2 repressed the malignant behaviors of NPC cells and tumor growth. Additionally, rescue experiments revealed that miR-30a-5p suppressed the proliferation, migration and invasion of NPC cells via targeting NUCB2 in vitro. Meanwhile, in vivo assays depicted that NUCB2 overexpression rescued the effects induced by miR-30a-5p upregulation on tumor growth. Conclusion: MiR-30a-5p modulates NPC progression by targeting NUCB2. These findings lay a foundation for exploring the clinical treatment of NPC.

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Sonic hedgehog regulates proliferation and differentiation of mesenchymal cells in the mouse metanephric kidney

Development ◽

10.1242/dev.129.22.5301 ◽

2002 ◽

Vol 129 (22) ◽

pp. 5301-5312 ◽

Cited By ~ 5

Author(s):

Jing Yu ◽

Thomas J. Carroll ◽

Andrew P. McMahon

Keyword(s):

Smooth Muscle ◽

Sonic Hedgehog ◽

Target Gene ◽

Kidney Development ◽

Metanephric Mesenchyme ◽

Downstream Target ◽

Proliferation And Differentiation ◽

Collecting Ducts ◽

Downstream Target Gene

Signaling by the ureteric bud epithelium is essential for survival,proliferation and differentiation of the metanephric mesenchyme during kidney development. Most studies that have addressed ureteric signaling have focused on the proximal, branching, ureteric epithelium. We demonstrate that sonic hedgehog is expressed in the ureteric epithelium of the distal, non-branching medullary collecting ducts and continues into the epithelium of the ureter— the urinary outflow tract that connects the kidney with the bladder. Upregulation of patched 1, the sonic hedgehog receptor and a downstream target gene of the signaling pathway in the mesenchyme surrounding the distal collecting ducts and the ureter suggests that sonic hedgehog acts as a paracrine signal. In vivo and in vitro analyses demonstrate that sonic hedgehog promotes mesenchymal cell proliferation, regulates the timing of differentiation of smooth muscle progenitor cells, and sets the pattern of mesenchymal differentiation through its dose-dependent inhibition of smooth muscle formation. In addition, we also show that bone morphogenetic protein 4 is a downstream target gene of sonic hedgehog signaling in kidney stroma and ureteral mesenchyme, but does not mediate the effects of sonic hedgehog in the control of mesenchymal proliferation.

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OP0246 MIR-214-3P PROTECTS AGAINST OSTEOARTHRITIS BY DIRECTLY TARGETING NF-ĸB PATHWAY

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2020-eular.5731 ◽

2020 ◽

Vol 79 (Suppl 1) ◽

pp. 155.2-155

Author(s):

Y. Cao ◽

S. Tang ◽

X. Nie ◽

W. Han ◽

Z. Zhu ◽

...

Keyword(s):

Signaling Pathway ◽

Therapeutic Potential ◽

Inhibitory Effect ◽

In Vivo Studies ◽

Luciferase Reporter ◽

Rt Pcr ◽

Human Cartilage ◽

Downstream Target

Background:Osteoarthritis (OA) is a degenerative disease associated with changes in the articular cartilage and bone, severely affecting patients’ mobility and quality of life. Multiple factors including mechanical stress, metabolic alteration and inflammatory mediators are involved in the complex pathogenesis of OA[1]. Interventions targeting these pathogenic factors may contribute to the treatment of OA. MiRNAs are single strand non-coding small RNAs, which are regulated in chondrogenesis and OA[2,3]. Recent studies demonstrated that miRNAs are involved in the regulation of NF-κB signaling pathway by different mechanisms[4]. These interactions suggest that NF-κB related miRNAs may be used as potential biomarkers and drug therapeutic targets in clinical treatment of OA. However, the relationship between miR-214-3p and NF-κB pathway remains poorly understood in OA.Objectives:This study aimed to test the expression and biological function of miR-214-3p in OA, and explore its mechanism in osteoarthritic chondrocytes.Methods:Articular primary chondrocytes were isolated from human cartilage samples, which were acquired from patients with end-stage knee OA at the time of total knee replacement surgery (n = 27), according to protocols approved by the Ethic Committee of Zhujiang Hospital. Real time PCR (RT-PCR) and in situ hybridization (ISH) were used to detect the expression of miR-214-3p in OA and non-OA cartilage tissues. Interference of miR-214-3p was conducted using inhibitor, while overexpression of miR-214-3p was performed with mimics. Metabolism of extracellular matrix was detected by RT-PCR, western blotting and immunofluorescence in vitro. Flow cytometry were conducted to determine cell apoptosis. A luciferase reporter assay, was used to evaluate the interaction between miR-214-3p and its downstream target. Human chondrocytes were cotransfected with miR-214-3p and the IKBKB-overexpressing plasmid to confirm the interaction between miR-214-3p and NF-ĸB pathway. For in vivo studies, experimental OA was induced in 12-week-old male C57BL/6J mice by destabilization of the medial meniscus (DMM) surgery with miR-214-3p agomir intra-articular (IA) injection (once weekly for 12 days) or by IA injection (once weekly for 12 days) of miR-214-3p antiagomir. Mice were sacrificed 10 weeks after the first IA injection, and subjected to histological analyses.Results:MiR-214-3p was significantly reduced in human OA cartilage. The decreased expression of miR-214-3p in the OA cartilage tissues was directly associated with excessive apoptosis and imbalance between anabolic and catabolic factors of ECM. Mechanistically, we determined that miR-214-3p directly targeted IKBKB/IKK-b and thereby suppressed the activation of NF-ĸB pathway. IKBKB overexpression attenuated the inhibitory effect of miR-214-3p on NF-ĸB pathway. Furthermore, inhibition of miR-214-3p in mice joints triggered spontaneous cartilage loss and OA development, while IA injection of miRNA-214-3p agomir alleviated OA in the DMM mouse model.Conclusion:Our results reveal an important role of miR-214-3p in OA progression. MiR-214-3p was down-regulated while IKBKB was upregulated in OA. MiR-214-3p inhibits the NF-kB signaling pathway and suppresses the progression of OA through targeting IKBKB. Thus, miR-214-3p maybe a therapeutic target for OA.References:[1]Glyn-Jones S, Palmer AJR, Agricola R, Price AJ, Vincent TL, Weinans H, Carr AJ:Osteoarthritis.The Lancet2015,386(9991):376-387.[2]Nugent M:MicroRNAs: exploring new horizons in osteoarthritis.Osteoarthritis and cartilage2016,24(4):573-580.[3]Vicente R, Noel D, Pers YM, Apparailly F, Jorgensen C:Deregulation and therapeutic potential of microRNAs in arthritic diseases.Nature reviews Rheumatology2016,12(4):211-220.[4]Xu B, Li YY, Ma J, Pei FX:Roles of microRNA and signaling pathway in osteoarthritis pathogenesis.Journal of Zhejiang University Science B2016,17(3):200-208.Disclosure of Interests:None declared

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KIAA1429 and ALKBH5 Oppositely Influence Aortic Dissection Progression via Regulating the Maturation of Pri-miR-143-3p in an m6A-Dependent Manner

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.668377 ◽

2021 ◽

Vol 9 ◽

Author(s):

Peng Wang ◽

Zhiwei Wang ◽

Min Zhang ◽

Qi Wu ◽

Feng Shi ◽

...

Keyword(s):

Aortic Dissection ◽

Dependent Manner ◽

Loss Of Function ◽

Therapeutic Approaches ◽

Downstream Target ◽

Downstream Target Gene ◽

Molecular Mechanism Of Action ◽

New Research

Despite decades of study into aortic dissection (AD), a lethal cardiovascular emergency due to a tear in the aorta intima or bleeding within the aortic wall, leading to the separation of the different layers of it, the factors that influence its progression and the deeper regulatory mechanisms remain poorly understood. Nowadays, with the maturity of N6-methyladenosine (m6A) sequence technology, m6A modification, one type of RNA epigenesis, has gradually become a new research hotspot for epigenetic molecular regulation. Especially recently, increasing evidence has revealed that m6A modification functions as a pivotal post-transcriptional modification to influence the progression of multiple diseases. Based on these findings, it is reasonable to speculate that m6A modification may affect the onset and progression of AD. To explore the validity of our conjecture and to elucidate its underlying molecular mechanism of action, we conducted the present study. In this study, we found that KIAA1429 is downregulated while ALKBH5 is upregulated in aortic tissues from AD patients. Furthermore, gain- and loss-of-function studies showed that KIAA1429 and ALKBH5 can oppositely regulate HASMC proliferation, HAEC apoptosis, and AD progression in AngII-infused mice. Mechanistically, we demonstrated that KIAA1429/ALKBH5-mediated m6A modifications can regulate the processing of pri-miR-143-3p through interacting with the microprocessor protein DGCR8, thus indirectly regulating the downstream target gene of mature miR-143-3p, DDX6, to perform their biological functions in vitro and in vivo. Our findings have revealed a novel connection between m6A modification and AD progression and may provide a novel molecular basis for subsequent researchers to search for novel therapeutic approaches to improve the health of patients struggling with AD.

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GNA13, a Novel Component of the GPR84/Beta-Catenin Signaling Axis in AML Stem Cells, Regulates the Maintenance of MLLAF9 Leukemogenesis

Blood ◽

10.1182/blood.v124.21.3577.3577 ◽

2014 ◽

Vol 124 (21) ◽

pp. 3577-3577

Author(s):

Philipp A Dietrich ◽

Murray D Norris ◽

Jenny Yingzi Wang

Keyword(s):

Stem Cells ◽

Drug Targets ◽

Conflicts Of Interest ◽

In Vivo Studies ◽

Brdu Incorporation ◽

Beta Catenin ◽

Downstream Target ◽

Signaling Axis

Abstract Beta-catenin is required for the establishment of leukemic stem cells (LSC) in mixed lineage leukemia (MLL)-rearranged acute myeloid leukemia (AML). Targeted inhibition of beta-catenin signaling has been hampered by the lack of pathway components amenable to pharmacological inhibition. Our recent study has identified a new beta-catenin regulator, GPR84, a member of the G protein-coupled receptor family that represents a highly tractable class of drug targets. Since no GPR84-specific anti-cancer compound has been developed to date, we aim to further delineate the GPR84/beta-catenin axis in order to inhibit other components of this signaling pathway pharmacologically. Preliminary data from our lab has implicated GNA13 as a potential downstream target gene of GPR84. In this study we sought to further examine the involvement of GNA13 in the GPR84/beta-catenin signaling axis. Functional studies showed that shRNA-mediated ablation of GNA13 significantly decreased the capacity of MLLAF9-LSC to form colonies in vitro (P=0.0184). We further demonstrated that GNA13 inhibition in LSC induced a significant G1 cell cycle arrest in vitro (P=0.0192). Subsequent in vivo studies showed that GNA13 downregulation in MLLAF9-LSC significantly delayed the onset of AML (P=0.0016) and significantly decreased LSC proliferation (P=0.0066). Hence GNA13 inhibition mimics the previously described GPR84-deficient phenotype in MLLAF9-LSC (Blood 2013;122:3781). In contrast, the GPR84-deficient phenotype could be rescued in MLLAF9-LSC by GNA13 overexpression in vitro (P=0.0130) and in vivo (P=0.0012). Further analysis showed that there was a significant increase in the mitotic index of GPR84-deficient LSC transduced with GNA13 cDNA in vitro, evidenced by an increase in G2/M-phase accumulation (P=0.0030) and p-histone H3 (P=0.0040). In addition, our data demonstrated a significant increase in BrdU incorporation of GPR84-deficient LSC overexpressing GNA13 in vivo (P<0.0001). In conclusion, our studies have identified GNA13, a novel component of the GPR84/beta-catenin axis. Since this pathway is pivotal for MLLAF9 AML maintenance by sustaining LSC self-renewal, targeting the GPR84/GNA13/beta-catenin signaling axis may provide a novel and promising strategy for improving the therapy and outcome of AML patients. Disclosures No relevant conflicts of interest to declare.

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Long noncoding RNA PVT1 promoted gallbladder cancer proliferation by epigenetically suppressing miR-18b-5p via DNA methylation

Cell Death and Disease ◽

10.1038/s41419-020-03080-x ◽

2020 ◽

Vol 11 (10) ◽

Author(s):

Longyang Jin ◽

Qiang Cai ◽

Shouhua Wang ◽

Shuqing Wang ◽

Jiandong Wang ◽

...

Keyword(s):

Dna Methylation ◽

Gallbladder Cancer ◽

Noncoding Rna ◽

Biliary Tree ◽

Cancer Proliferation ◽

Downstream Target ◽

Downstream Target Gene ◽

Proliferation In Vitro

Abstract Gallbladder cancer (GBC) accounts for 85–90% malignancies of the biliary tree worldwide. Considerable evidence has demonstrated that dysregulation of lncRNAs is involved in the progression of cancer. LncRNA PVT1 has been reported to play important roles in various cancers, but its role in gallbladder cancer remains unknown. In the present study, we found that PVT1 was upregulated in GBC tissues and cells, and its upregulation was related with poor prognosis in GBC patients. PVT1 promoted GBC cells proliferation in vitro and in vivo. Mechanistically, PVT1 recruited DNMT1 via EZH2 to the miR-18b-5p DNA promoter and suppressed the transcription of miR-18b-5p through DNA methylation. Moreover, HIF1A was proved to be the downstream target gene of miR-18b-5p and PVT1 regulated GBC cells proliferation via HIF1A. In conclusion, our studies clarified the PVT1/miR-18b-5p/HIF1A regulation axis and indicated that PVT1 could be a potential therapeutic target for GBC.

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