An in vivo GA‐ and ABA‐responsive dual‐luciferase reporter system for simultaneous detection of GA and ABA responses, hormone crosstalk, and heat‐stress response in rice

Abstract Background: Gliomas are the most common and malignant tumors in the brain of humans, and the prognosis of glioma patient is very poor. MicroRNAs (miRNAs) play critical roles in different types of cancer by regulating gene expression at the posttranscriptional levels. Although miR-218 has been reported to be downregulated in gliomas, its role in gliomas still remains largely unknown. Methods: MiR-218 expression in gliomas and normal brain tissues (control subjects) were analyzed using TCGA dataset. The biological roles of miR-218 in glioma cells were determined by a series of in vitro and in vivo studies. The dual-luciferase reporter system was performed to identify potential targets of miR-218. The regulatory effect of miR-218 on TNC/AKT/AP-1/TGFβ1 pathway was evaluated by dual-luciferase reporter system and western blot.Results: We demonstrated miR-218 was significantly downregulated in gliomas compared to control subjects, and exerted a potent tumor suppressor in glioma cells by inhibiting cell proliferation, colony formation, migration, invasion and tumorigenic potential in nude mice, as well as inducing cell cycle arrest and apoptosis.Mechanistically, miR-218 inhibited malignant phenotypes of glioma cells by binding to the 3’ UTR of its target TNC and subsequently repressing its expression. As a result, it could reduce AKT phosphorylation and subsequently inhibit transcriptional activity of AP-1 by reducing JNK phosphorylation, downregulating the expression of TGFβ1, while TGFβ1 is able to, in turn, activate the TNC/AKT/AP-1 signaling axis.Conclusions: Our data uncover a previously unknown tumor suppressor role of miR-218 in glioma by blocking the TNC/AKT/AP-1/TGFβ1 positive feedback loop.

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Quantitative Assessment of Pdx1 Promoter Activity in Vivo Using a Secreted Luciferase Reporter System

Endocrinology ◽

10.1210/en.2012-2248 ◽

2013 ◽

Vol 154 (11) ◽

pp. 4388-4395 ◽

Cited By ~ 7

Author(s):

Wataru Nishimura ◽

Koki Eto ◽

Atsushi Miki ◽

Motohito Goto ◽

Miho Kawaguchi ◽

...

Keyword(s):

Promoter Activity ◽

Quantitative Assessment ◽

Luciferase Reporter ◽

Reporter System ◽

Luciferase Reporter System

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SET-9 and SET-26 are H3K4me3 readers and play critical roles in germline development and longevity

eLife ◽

10.7554/elife.34970 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 6

Author(s):

Wenke Wang ◽

Amaresh Chaturbedi ◽

Minghui Wang ◽

Serim An ◽

Satheeja Santhi Velayudhan ◽

...

Keyword(s):

Heat Stress ◽

Stress Response ◽

Binding Sites ◽

Target Genes ◽

Germline Development ◽

Heat Stress Response ◽

C Elegans ◽

Redundant Functions

C. elegans SET-9 and SET-26 are highly homologous paralogs that share redundant functions in germline development, but SET-26 alone plays a key role in longevity and heat stress response. Whereas SET-26 is broadly expressed, SET-9 is only detectable in the germline, which likely accounts for their different biological roles. SET-9 and SET-26 bind to H3K4me3 with adjacent acetylation marks in vitro and in vivo. In the soma, SET-26 acts through DAF-16 to modulate longevity. In the germline, SET-9 and SET-26 restrict H3K4me3 domains around SET-9 and SET-26 binding sites, and regulate the expression of specific target genes, with critical consequence on germline development. SET-9 and SET-26 are highly conserved and our findings provide new insights into the functions of these H3K4me3 readers in germline development and longevity.

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MiR-218 inhibits malignant phenotypes of glioma by targeting TNC/AKT/AP-1/TGFβ1 feedback signaling loop

10.21203/rs.3.rs-16349/v2 ◽

2020 ◽

Author(s):

Siwen Dang ◽

Rui Zhang ◽

Sijia Tian ◽

Banjun Ruan ◽

Peng Hou ◽

...

Keyword(s):

Tumor Suppressor ◽

Regulation Of Gene Expression ◽

Glioma Cells ◽

Luciferase Reporter ◽

Normal Brain ◽

Reporter System ◽

Control Subjects ◽

Luciferase Reporter System

Abstract Background: Gliomas are the most malignant and common tumors in human brains, and the prognosis of glioma patient is very poor. MicroRNAs (miRNAs) play critical roles in different types of cancer by performing posttranscriptional regulation of gene expression. Although miR-218 has been demonstrated decreased in gliomas, its role in gliomas still remains largely unknown. Methods: MiR-218 expression were analyzed in gliomas and normal brain tissues (control subjects) using TCGA dataset. A series of in vitro and in vivo studies was performed to determine the biological roles of miR-218 in glioma cells. Potential targets of miR-218 were identified using dual-luciferase reporter system. Western blot and dual-luciferase reporter system were performed to evaluate the regulatory effect of miR-218 on TNC/AKT/AP-1/TGFβ1 pathway.Results: We demonstrated miR-218 was significantly downregulated in gliomas compared to control subjects, and played potent tumor suppressor roles in glioma cells by inhibited cell proliferation, colony formation, migration, invasion and tumorigenic potential in nude mice, as well as inducing cell cycle arrest and apoptosis.Mechanistically, miR-218 inhibited malignant phenotypes of glioma cells by binding to the 3’ UTR of its target TNC and subsequently repressing its expression. As a result, it could reduce AKT phosphorylation and subsequently inhibit transcriptional activity of AP-1 by reducing JNK phosphorylation, downregulating the expression of TGFβ1, while TGFβ1 is able to, in turn, activate the TNC/AKT/AP-1 signaling axis.Conclusions: Our data uncover a previously unknown tumor suppressor role of miR-218 by blocking the TNC/AKT/AP-1/TGFβ1 positive feedback loop in glioma.

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miR-30a as Potential Therapeutic by Targeting TET1 through Regulation of the Hydroxymethlation of Drp-1 Promoter in IPF

10.20944/preprints201701.0074.v1 ◽

2017 ◽

Author(s):

Songzi Zhang ◽

Huizhu Liu ◽

Yuxia Liu ◽

Jie Zhang ◽

Hongbo Li ◽

...

Keyword(s):

Therapeutic Target ◽

Target Gene ◽

Mitochondrial Fission ◽

Luciferase Reporter ◽

Biological Processes ◽

Reporter System ◽

Potential Therapeutic Target ◽

Direct Target ◽

Luciferase Reporter System

Several recent studies have indicated that miR-30a plays critical roles in various biological processes and diseases. However, the mechanism of miR-30a participation in the regulation of idiopathic pulmonary fibrosis (IPF) is ambiguous. Our previous study demonstrated that miR-30a may function as a novel therapeutic target for lung ﬁbrosis by blocking mitochondrial fission, which is dependent on dynamin-related protein-1 (Drp-1). However, the regulatory mechanism between miR-30a and Drp-1 has yet to be investigated. In addition, whether miR-30a can act as a potential therapeutic has not been verified in vivo. In this study, the miR-30a expression in IPF patients was evaluated. Computational analysis and a dual luciferase reporter system assay were used to identify the target gene of miR-30a, and cell transfection was used to confirm this relationship. Ten-eleven translocation 1 (TET1) was validated as a direct target of miR-30a, and the transfection of miR-30a mimic/inhibitor significantly reduced/increased the expression of TET1 protein. Further experiment verified that the interference on TET1(siRNA) could inhibit the hydroxymethlation of the Drp-1 promoter. Finally, miR-30a agomir was designed and applied to identify and validate the therapeutic effect of miR-30a in vivo. Our study demonstrated that miR-30a could inhibit the TET1 expression by base pairing with complementary sites in the 3′ untranslated region to regulate the hydroxymethlation of the Drp-1 promoter. Furthermore, miR-30a could act as a potential therapeutic target for IPF.

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