ZNF395 Is an Activator of a Subset of IFN-Stimulated Genes

Activation of the interferon (IFN) pathway in response to infection with pathogens results in the induction of IFN-stimulated genes (ISGs) including proinflammatory cytokines, which mount the proper antiviral immune response. However, aberrant expression of these genes is pathogenic to the host. In addition to IFN-induced transcription factors non-IFN-regulated factors contribute to the transcriptional control of ISGs. Here, we show by genome wide expression analysis, siRNA-mediated suppression and Doxycycline-induced overexpression that the cellular transcription factor ZNF395 activates a subset of ISGs including the chemokines CXCL10 and CXCL11 in keratinocytes. We found that ZNF395 acts independently of IFN but enhances the IFN-induced expression of CXCL10 and CXCL11. Luciferase reporter assays revealed a requirement of intact NFκB-binding sites for ZNF395 to stimulate the CXCL10 promoter. The transcriptional activation of CXCL10 and CXCL11 by ZNF395 was abolished after inhibition of IKK by BMS-345541, which increased the stability of ZNF395. ZNF395 encodes at least two motifs that mediate the enhanced degradation of ZNF395 in response to IKK activation. Thus, IKK is required for ZNF395-mediated activation of transcription and enhances its turn-over to keep the activity of ZNF395 low. Our results support a previously unrecognized role of ZNF395 in the innate immune response and inflammation.

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MITF induces escape from innate immunity in melanoma

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-021-01916-8 ◽

2021 ◽

Vol 40 (1) ◽

Author(s):

Luis Sánchez-del-Campo ◽

Román Martí-Díaz ◽

María F. Montenegro ◽

Rebeca González-Guerrero ◽

Trinidad Hernández-Caselles ◽

...

Keyword(s):

Immune Response ◽

Immune System ◽

Nk Cells ◽

Nk Cell ◽

Luciferase Reporter ◽

Damage Repair ◽

Reporter Assays ◽

Underlying Mechanisms ◽

Nk Cytotoxicity

Abstract Background The application of immune-based therapies has revolutionized cancer treatment. Yet how the immune system responds to phenotypically heterogeneous populations within tumors is poorly understood. In melanoma, one of the major determinants of phenotypic identity is the lineage survival oncogene MITF that integrates diverse microenvironmental cues to coordinate melanoma survival, senescence bypass, differentiation, proliferation, invasion, metabolism and DNA damage repair. Whether MITF also controls the immune response is unknown. Methods By using several mouse melanoma models, we examine the potential role of MITF to modulate the anti-melanoma immune response. ChIP-seq data analysis, ChIP-qPCR, CRISPR-Cas9 genome editing, and luciferase reporter assays were utilized to identify ADAM10 as a direct MITF target gene. Western blotting, confocal microscopy, flow cytometry, and natural killer (NK) cytotoxicity assays were used to determine the underlying mechanisms by which MITF-driven phenotypic plasticity modulates melanoma NK cell-mediated killing. Results Here we show that MITF regulates expression of ADAM10, a key sheddase that cleaves the MICA/B family of ligands for NK cells. By controlling melanoma recognition by NK-cells MITF thereby controls the melanoma response to the innate immune system. Consequently, while melanoma MITFLow cells can be effectively suppressed by NK-mediated killing, MITF-expressing cells escape NK cell surveillance. Conclusion Our results reveal how modulation of MITF activity can impact the anti-melanoma immune response with implications for the application of anti-melanoma immunotherapies.

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Polo-like kinase 3 inhibits glucose metabolism in colorectal cancer by targeting HSP90/STAT3/HK2 signaling

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-019-1418-2 ◽

2019 ◽

Vol 38 (1) ◽

Cited By ~ 1

Author(s):

Baochi Ou ◽

Hongze Sun ◽

Jingkun Zhao ◽

Zhuoqing Xu ◽

Yuan Liu ◽

...

Keyword(s):

Colorectal Cancer ◽

Glucose Metabolism ◽

Transcriptional Activation ◽

Lactate Production ◽

Luciferase Reporter ◽

Transcriptional Level ◽

Atp Production ◽

Reporter Assays

Abstract Background Polo-like kinase 3 (PLK3) has been documented as a tumor suppressor in several types of malignancies. However, the role of PLK3 in colorectal cancer (CRC) progression and glucose metabolism remains to be known. Methods The expression of PLK3 in CRC tissues was determined by immunohistochemistry. Cells proliferation was examined by EdU, CCK-8 and in vivo analyses. Glucose metabolism was assessed by detecting lactate production, glucose uptake, mitochondrial respiration, extracellular acidification rate, oxygen consumption rate and ATP production. Chromatin immunoprecipitation, luciferase reporter assays and co-immunoprecipitation were performed to explore the signaling pathway. Specific targeting by miRNAs was determined by luciferase reporter assays and correlation with target protein expression. Results PLK3 was significantly downregulated in CRC tissues and its low expression was correlated with worse prognosis of patients. In vitro and in vivo experiments revealed that PLK3 contributed to growth inhibition of CRC cells. Furthermore, we demonstrated that PLK3 impeded glucose metabolism via targeting Hexokinase 2 (HK2) expression. Mechanically, PLK3 bound to Heat shock protein 90 (HSP90) and facilitated its degradation, which led to a significant decrease of phosphorylated STAT3. The downregulation of p-STAT3 further suppressed the transcriptional activation of HK2. Moreover, our investigations showed that PLK3 was directly targeted by miR-106b at post-transcriptional level in CRC cells. Conclusion This study suggests that PLK3 inhibits glucose metabolism by targeting HSP90/STAT3/HK2 signaling and PLK3 may serve as a potential therapeutic target in colorectal cancer.

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CircRNA_0058063 functions as a ceRNA in bladder cancer progression via targeting miR-486-3p/FOXP4 axis

10.21203/rs.2.17516/v2 ◽

2019 ◽

Author(s):

Haote Liang ◽

Hang Huang ◽

Yeping Li ◽

Yongyong Lu ◽

Tingyu Ye

Keyword(s):

Bladder Cancer ◽

Cancer Progression ◽

Luciferase Reporter ◽

Circular Rnas ◽

Aberrant Expression ◽

Normal Tissues ◽

Tumorigenesis And Progression ◽

Rna Immunoprecipitation ◽

Reporter Assays ◽

Proliferation And Invasion

Abstract Emerging evidences have uncovered critical regulatory roles of circular RNAs (circRNAs) function as dynamic scaffolding molecules in tumorigenesis and progression. However, the aberrant expression and clinical significance of hsa_circ_0058063 (circRNA_0058063) in bladder cancer (BC) remain poorly understood. circRNA expression was analyzed via a microarray in cancerous tissue and non-carcinoma tissues. Luciferase reporter assays and RNA immunoprecipitation (RIP) were both conducted to uncover the function of circRNA_0058063 in BC. circRNA_0058063 was overexpressed in BC tissues compared to adjacent normal tissues. Knockdown of circRNA_0058063 dramatically decreased cell proliferation and invasion, and promoted apoptosis in 5637 and BIU-87 cell lines. Furthermore, mechanistic investigations showed that circRNA_0058063 and FOXP4 could directly bind to miR-486-3p, demonstrating that circRNA_0058063 regulated FOXP4 expression by competitively binding to miR-486-3p. Taken together, circRNA_0058063 functions by sponging miR-486-3p in BC progression, which could be act as a new biomarker and further developed to be a therapeutic target in BC.

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ELK4 promotes the development of gastric cancer by inducing M2 polarization of macrophages through regulation of the KDM5A-PJA2-KSR1 axis

Journal of Translational Medicine ◽

10.1186/s12967-021-02915-1 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Lei Zheng ◽

Hongmei Xu ◽

Ya Di ◽

Lanlan Chen ◽

Jiao Liu ◽

...

Keyword(s):

Gastric Cancer ◽

Transcriptional Activation ◽

Luciferase Reporter ◽

Loss Of Function ◽

Luciferase Reporter Gene ◽

M2 Polarization ◽

Lysine Specific Demethylase ◽

Dependent Inhibition ◽

Kinase Suppressor Of Ras ◽

The Stability

Abstract Background We tried to elaborate the molecular mechanism of ETS-like transcription factor 4 (ELK4) affecting gastric cancer (GC) progression through M2 polarization of macrophages mediated by lysine-specific demethylase 5A (KDM5A)-Praja2 (PJA2)-kinase suppressor of ras 1 (KSR1) axis. Methods GC expression dataset was obtained from GEO database, and the downstream regulatory mechanism of ELK4 was predicted. Tumor-associated macrophages (TAMs) were isolated from GC tissues. The interaction among ELK4, KDM5A, PJA2 and KSR1 was analyzed by dual luciferase reporter gene, ChIP and Co-IP assays. The stability of KSR1 protein was detected by cycloheximide (CHX) treatment. After TAMs were co-cultured with HGC-27 cells, HGC-27 cell biological processes were assessed through gain- and loss-of function assays. Tumorigenicity was detected by tumorigenicity test in nude mice. Results In GC and TAMs, ELK4, KDM5A and KSR1 were highly expressed, while PJA2 was lowly expressed. M2 polarization of macrophages promoted the development of GC. ELK4 activated KDM5A by transcription and promoted macrophage M2 polarization. KDM5A inhibited the expression of PJA2 by removing H3K4me3 of PJA2 promoter, which promoted M2 polarization of macrophages. PJA2 reduced KSR1 by ubiquitination. ELK4 promoted the proliferative, migrative and invasive potentials of GC cells as well as the growth of GC xenografts by regulating KSR1. Conclusion ELK4 may reduce the PJA2-dependent inhibition of KSR1 by transcriptional activation of KDM5A to promote M2 polarization of macrophages, thus promoting the development of GC.

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CircRNA 0058063 functions as a ceRNA in bladder cancer progression via targeting miR-486-3p/FOXP4 axis

10.21203/rs.2.17516/v1 ◽

2019 ◽

Author(s):

Haote Liang ◽

Hang Huang ◽

Yeping Li ◽

Yongyong Lu ◽

Tingyu Ye

Keyword(s):

Bladder Cancer ◽

Cancer Progression ◽

Luciferase Reporter ◽

Circular Rnas ◽

Aberrant Expression ◽

Normal Tissues ◽

Tumorigenesis And Progression ◽

Rna Immunoprecipitation ◽

Reporter Assays ◽

Proliferation And Invasion

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Reconstitution of the Jasmonate Signaling Pathway in Plant Protoplasts

Cells ◽

10.3390/cells8121532 ◽

2019 ◽

Vol 8 (12) ◽

pp. 1532 ◽

Cited By ~ 1

Author(s):

Ning Li ◽

Joachim F. Uhrig ◽

Corinna Thurow ◽

Li-Jun Huang ◽

Christiane Gatz

Keyword(s):

Signaling Pathway ◽

Function Analysis ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Protein Levels ◽

Activation Of Transcription ◽

Subsequent Degradation ◽

The Stability ◽

F Box Protein ◽

Ja Signaling

The phytohormone jasmonic acid (JA) plays an important role in various plant developmental processes and environmental adaptations. The JA signaling pathway has been well-elucidated in the reference plant Arabidopsis thaliana. It starts with the perception of the active JA derivative, jasmonoyl-isoleucine (JA-Ile), by the F-box protein COI1 which is part of the E3-ligase SCFCOI1. Binding of JA-Ile enables the interaction between COI1 and JAZ repressor proteins. Subsequent degradation of JAZ proteins leads to the activation of transcription factors like e.g., MYC2. Here we demonstrate that the pathway can be reconstituted in transiently transformed protoplasts. Analysis of the stability of a JAZ1-fLuc fusion protein as a function of COI1 transiently expressed in coi1 protoplasts allows structure function analysis of both JAZs and COI1. Using this system, we found that conserved cysteines in COI1 influence steady state COI1 protein levels. Using a luciferase reporter gene under the control of the JAZ1 promoter enable to address those features of JAZ1 that are required for MYC2 repression. Interestingly, the conserved TIFY-motif previously described to interact with NINJA to recruit the corepressor TOPLESS is not necessary for repression. This result is in favor of the alternative repression mode that proposes a direct competition between repressive JAZs and promotive MEDIATOR25 at MYC2. Finally, using protoplasts from the aos coi1 double mutant, which is deficient in JA synthesis and perception, we provide a system that has the potential to study the activity of different COI1 variants in the presence of different ligands.

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Aberrant expression of hsa_circ_0025036 in lung adenocarcinoma and its potential roles in regulating cell proliferation and apoptosis

Biological Chemistry ◽

10.1515/hsz-2018-0303 ◽

2018 ◽

Vol 399 (12) ◽

pp. 1457-1467 ◽

Cited By ~ 4

Author(s):

Shujun Wu ◽

Hui Li ◽

Chunya Lu ◽

Furui Zhang ◽

Huaqi Wang ◽

...

Keyword(s):

Cell Proliferation ◽

Lung Adenocarcinoma ◽

Luciferase Reporter ◽

Clinicopathological Parameters ◽

Circular Rnas ◽

Aberrant Expression ◽

Close Link ◽

Adenocarcinoma Cells ◽

Proliferation And Apoptosis ◽

Reporter Assays

AbstractAs the most common histological subtype of lung cancer, lung adenocarcinoma remains a tremendous risk to public health, which requires ceaseless efforts to elucidate the potential diagnostic and therapeutic strategies. Circular RNAs (circRNAs) have been identified with emerging roles in tumorigenesis and development. Our preliminary work noticed that hsa_circ_0025036 was significantly upregulated in lung adenocarcinoma tissues. However, its specific roles in lung adenocarcinoma remain unclear. The results in this study revealed that hsa_circ_0025036 existed as a circular form and was aberrantly upregulated in lung adenocarcinoma tissues via quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Its expression level exhibited a close link with aggressive clinicopathological parameters including cancer differentiation, TNM stage and lymph node metastasis. hsa_circ_0025036 knockdown significantly suppressed cell proliferation and promoted cell apoptosis in A549 and Calu-3 cells. Moreover, hsa_circ_0025036/miR-198/SHMT1&TGF-αaxis was identified via bioinformatics analysis and Dual-Luciferase Reporter assays. miR-198 inhibitors reversed the function of hsa_circ_0025036 knockdown. hsa_circ_0025036 knockdown exerted similar effects with miR-198 upregulation on cell proliferation and apoptosis. In conclusion, we demonstrate that hsa_circ_0025036 regulates cell proliferation and apoptosis in lung adenocarcinoma cells probably via hsa_circ_0025036/miR-198/SHMT1&TGF-αaxis. hsa_circ_0025036 may serve as a potential prognostic biomarker and a therapeutic target for lung adenocarcinoma.

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Inhibition of NF-κB Activation in Combination with Bcl-2 Expression Allows for Persistence of First-Generation Adenovirus Vectors in the Mouse Liver

Journal of Virology ◽

10.1128/jvi.72.11.9267-9277.1998 ◽

1998 ◽

Vol 72 (11) ◽

pp. 9267-9277 ◽

Cited By ~ 59

Author(s):

André Lieber ◽

Chen-Yi He ◽

Leonard Meuse ◽

Charis Himeda ◽

Christopher Wilson ◽

...

Keyword(s):

Gene Expression ◽

Immune Response ◽

Transgenic Mice ◽

Transcriptional Activation ◽

First Generation ◽

Viral Gene ◽

Gene Products ◽

Antiviral Immune Response ◽

Innate Defense ◽

Adenovirus Vectors

ABSTRACT NF-κB is a key regulator of the innate antiviral immune response, due in part to its transcriptional activation of cytokines and adhesion molecules, which, in turn, function in chemotaxis and activation of inflammatory cells. We reported earlier that viral gene expression in hepatocytes transduced with first-generation (E1-deleted) adenoviruses induced NF-κB activation, elevation of serum cytokines, and hepatocellular apoptosis during the first days postinfusion. These events did not occur in mice infused with an adenovirus vector deleted for E1, E2, E3, and late gene expression. In the present study, we used an adenovirus expressing an IκBα supersuppressor (Ad.IκBM) andbcl-2 transgenic mice to unravel the role of virus-induced NF-κB activation and apoptosis in the clearance of recombinant adenovirus vectors from the liver. The combined action of IκBM and Bcl-2 allowed for vector persistence in livers of C57BL/6 × C3H mice. In the absence of Bcl-2, IκBM expression in mouse livers significantly reduced NF-κB activation, cytokine expression, leukocyte infiltration, and the humoral immune response against the transgene product; however, this was not sufficient to prevent the decline of vector DNA in transduced cells. Infusion of Ad.IκBM caused extended apoptosis predominantly in periportal liver regions, indicating that NF-κB activation may protect transduced hepatocytes from apoptosis induced by adenovirus gene products. To confer vector persistence, bcl-2 transgene expression was required to block virus-induced apoptosis if NF-κB protection was inactivated by IκBM. Expression of gene products involved in early stages of apoptotic pathways was up-regulated in response to virus infusion inbcl-2 transgenic mice, which may represent a compensatory effect. Our study supports the idea that the suppression of innate defense mechanisms improves vector persistence.

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LncRNA OTUD6B-AS1 Induces Cisplatin Resistance in Cervical Cancer Cells Through Up-Regulating Cyclin D2 via miR-206

Frontiers in Oncology ◽

10.3389/fonc.2021.777220 ◽

2021 ◽

Vol 11 ◽

Author(s):

Hui Hou ◽

Rong Yu ◽

Haiping Zhao ◽

Hao Yang ◽

Yuchong Hu ◽

...

Keyword(s):

Cervical Cancer ◽

Cancer Cells ◽

Cisplatin Resistance ◽

Luciferase Reporter ◽

Cyclin D2 ◽

Aberrant Expression ◽

Qrt Pcr ◽

Cervical Cancer Cells ◽

Reporter Assays ◽

Underlying Mechanisms

Cervical cancer is one of the most common gynecological cancers. Cisplatin resistance remains a major hurdle in the successful treatment of cervical cancer. Aberrant expression of long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) are implicated in cisplatin resistance. However, the regulatory functions of lncRNAs and miRNAs in cervical cancer cisplatin resistance and the underlying mechanisms are still elusive. Our qRT-PCR assays verified that miR-206 levels were down-regulated in cisplatin-resistant cervical cancer cells. The introduction of miR-206 sensitized cisplatin-resistant cervical cancer cells to cisplatin. Our qRT-PCR and luciferase reporter assays showed that Cyclin D2 (CCND2) was the direct target for miR-206 in cervical cancer cells. The cisplatin-resistant cervical cancer cells expressed higher CCND2 expression than the parental cells, whereas inhibition of CCND2 could sensitize the resistant cells to cisplatin treatment. Furthermore, we demonstrated that lncRNA OTUD6B-AS1 was up-regulated in cisplatin-resistant cervical cancer cells, and knocking down OTUD6B-AS1 expression induced re-acquirement of chemosensitivity to cisplatin in cervical cancer cells. We also showed that OTUD6B-AS1 up-regulated the expression of CCND2 by sponging miR-206. Low miR-206 and high OTUD6B-AS1 expression were associated with significantly poorer overall survival. Taken together, these results suggest that OTUD6B-AS1-mediated down-regulation of miR-206 increases CCND2 expression, leading to cisplatin resistance. Modulation of these molecules may be a therapeutic approach for cisplatin-resistant cervical cancer.

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CircRNA_0058063 functions as a ceRNA in bladder cancer progression via targeting miR-486-3p/FOXP4 axis

Bioscience Reports ◽

10.1042/bsr20193484 ◽

2020 ◽

Vol 40 (3) ◽

Cited By ~ 4

Author(s):

Haote Liang ◽

Hang Huang ◽

Yeping Li ◽

Yongyong Lu ◽

Tingyu Ye

Keyword(s):

Bladder Cancer ◽

Cancer Progression ◽

Luciferase Reporter ◽

Circular Rnas ◽

Aberrant Expression ◽

Normal Tissues ◽

Tumorigenesis And Progression ◽

Rna Immunoprecipitation ◽

Reporter Assays ◽

Proliferation And Invasion

Abstract Emerging evidence has uncovered critical regulatory roles of circular RNAs (circRNAs) function as dynamic scaffolding molecules in tumorigenesis and progression. However, the aberrant expression and clinical significance of hsa_circ_0058063 (circRNA_0058063) in bladder cancer (BC) remain poorly understood. circRNA expression was analyzed via a microarray in cancerous tissue and non-carcinoma tissues. Luciferase reporter assays and RNA immunoprecipitation (RIP) were both conducted to uncover the function of circRNA_0058063 in BC. circRNA_0058063 was overexpressed in BC tissues compared with adjacent normal tissues. Knockdown of circRNA_0058063 dramatically decreased cell proliferation and invasion, and promoted apoptosis in 5637 and BIU-87 cell lines. Furthermore, mechanistic investigations showed that circRNA_0058063 and FOXP4 could directly bind to miR-486-3p, demonstrating that circRNA_0058063 regulated FOXP4 expression by competitively binding to miR-486-3p. Taken together, circRNA_0058063 functions by sponging miR-486-3p in BC progression, which could act as a new biomarker and further developed to be a therapeutic target in BC.

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