scholarly journals LIM homeodomain transcription factor Isl1 affects urethral epithelium differentiation and apoptosis via Shh

2019 ◽  
Vol 10 (10) ◽  
Author(s):  
Tiantian Su ◽  
Hui Liu ◽  
Di Zhang ◽  
Guojin Xu ◽  
Jiali Liu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Luciferase Reporter ◽  
Knockout Mouse Model ◽  
Functional Studies ◽  
Human Disorders ◽  
Homeobox Transcription Factor ◽  
Reporter Assays ◽  
The Common ◽  
Mechanistic Basis ◽  
Genital Tubercle

Abstract Urethral hypoplasia, including failure of urethral tube closure, is one of the common phenotypes observed in hereditary human disorders, the mechanism of which remains unclear. The present study was thus designed to study the expression, functions, and related mechanisms of the LIM homeobox transcription factor Isl1 throughout mouse urethral development. Results showed that Isl1 was highly expressed in urethral epithelial cells and mesenchymal cells of the genital tubercle (GT). Functional studies were carried out by utilizing the tamoxifen-inducible Isl1-knockout mouse model. Histological and morphological results indicated that Isl1 deletion caused urethral hypoplasia and inhibited maturation of the complex urethral epithelium. In addition, we show that Isl1-deleted mice failed to maintain the progenitor cell population required for renewal of urethral epithelium during tubular morphogenesis and exhibited significantly increased cell death within the urethra. Dual-Luciferase reporter assays and yeast one-hybrid assays showed that ISL1 was essential for normal urethral development by directly targeting the Shh gene. Collectively, results presented here demonstrated that Isl1 plays a crucial role in mouse urethral development, thus increasing our potential for understanding the mechanistic basis of hereditary urethral hypoplasia.

scholarly journals Hsa_circ_0026628 promotes the development of colorectal cancer by targeting SP1 to activate the Wnt/β-catenin pathway

2021 ◽  
Vol 12 (9) ◽  
Author(s):  
Xuexiu Zhang ◽  
Jianning Yao ◽  
Haoling Shi ◽  
Bing Gao ◽  
Haining Zhou ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Transcription Factor ◽  
Circular Rna ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Transcriptional Level ◽  
Sp1 Transcription Factor ◽  
Reporter Assays ◽  
Sphere Formation

AbstractCircular RNAs (circRNAs) have been reported to play crucial roles in the progression of various cancers, including colorectal cancer (CRC). SP1 (Sp1 transcription factor) is a well-recognized oncogene in CRC and is deemed to trigger the Wnt/β-catenin pathway. The present study was designed to investigate the role of circRNAs which shared the same pre-mRNA with SP1 in CRC cells. We identified that hsa_circ_0026628 (circ_0026628), a circular RNA that originated from SP1 pre-mRNA, was upregulated in CRC cells. Sanger sequencing and agarose gel electrophoresis verified the circular characteristic of circ_0026628. Functional assays including CCK-8, colony formation, transwell, immunofluorescence staining, and sphere formation assay revealed the function of circ_0026628. RNA pull-down and mass spectrometry disclosed the proteins interacting with circ_0026628. Mechanistic assays including RIP, RNA pull-down, CoIP, ChIP, and luciferase reporter assays demonstrated the interplays between molecules. The results depicted that circ_0026628 functioned as a contributor to CRC cell proliferation, migration, EMT, and stemness. Mechanistically, circ_0026628 served as the endogenous sponge of miR-346 and FUS to elevate SP1 expression at the post-transcriptional level, thus strengthening the interaction between SP1 and β-catenin to activate the Wnt/β-catenin pathway. In turn, the downstream gene of Wnt/β-catenin signaling, SOX2 (SRY-box transcription factor 2), transcriptionally activated SP1 and therefore boosted circ_0026628 level. On the whole, SOX2-induced circ_0026628 sponged miR-346 and recruited FUS protein to augment SP1, triggering the downstream Wnt/β-catenin pathway to facilitate CRC progression.

scholarly journals Transcriptomic and metabolomic analysis provides insights into anthocyanin and pyocyanin accumulation in pear

2020 ◽  
Author(s):  
Zhen Zhang ◽  
Changping Tian ◽  
Ya Zhang ◽  
Chenzhiyu Li ◽  
Xi Li ◽  
...  
Keyword(s):  
Luciferase Reporter ◽  
Strongly Correlated ◽  
Mobility Shift ◽  
Data Set ◽  
Functional Studies ◽  
Nutritive Quality ◽  
Core Set ◽  
Reporter Assays ◽  
Arabidopsis Mutant

Abstract Background Pear is one of the most important fruit crops worldwide. Anthocyanins and procyanidins (PAs) are important secondary metabolites that affect the appearance and nutritive quality of pear. However, few studies have focused on the molecular mechanism underlying anthocyanin and PA accumulation in pear . Results We conducted metabolome and transcriptome analyses to identify candidate genes involved in anthocyanin and PA accumulation in young fruits of the pear cultivar ‘Clapp Favorite’ (CF) and its red mutation cultivar ‘Red Clapp Favorite’ ( RCF ). Gene–metabolite correlation analyses revealed a ‘core set’ of 20 genes that were strongly correlated with 10 anthocyanin and seven PA metabolites. Of these, PcGSTF12 was confirmed to be involved in anthocyanin and PA accumulation by complementation of the tt19-7 Arabidopsis mutant. Interestingly, PcGSTF12 was found to be responsible for the accumulation of procyanidin A3, but not petunidin 3, 5-diglucoside, opposite to the function of AtGSTs in Arabidopsis . Transformation with PcGSTF12 greatly promoted or repressed genes involved in anthocyanin and PA biosynthesis, regulation, and transport. Electrophoretic mobility shift and luciferase reporter assays confirmed positive regulation of PcGSTF12 by PcMYB114 . Conclusion These findings identify a core set of genes for anthocyanin and PA accumulation in pear. Of these, PcGSTF12 , was confirmed to be involved in anthocyanin and PA accumulation. Our results also identified an important anthocyanin and PA regulation node comprising two core genes, PcGSTF12 and PcMYB114 . These results provide novel insights into anthocyanin and PA accumulation in pear and represent a valuable data set to guide future functional studies and pear breeding.

scholarly journals miR-142-5p promotes cervical cancer progression by targeting LMX1A through Wnt/β-catenin pathway

Open Medicine ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. 224-236
Author(s):  
Lijuan Ke ◽  
Yanping Chen ◽  
Yiying Li ◽  
Zheng Chen ◽  
Yihui He ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Cancer Progression ◽  
Hela Cells ◽  
Luciferase Reporter ◽  
Carcinogenic Effect ◽  
Expression Levels ◽  
Normal Tissues ◽  
Homeobox Transcription Factor ◽  
Reporter Assays ◽  
Cancer Tissues

Abstract Background Previous work has shown that miR-142-5p in cervical cancer tissues increased significantly compared with adjacent normal tissues. However, the function and the mechanism of miR-142-5p in cervical cancer have not been reported. Methods Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to determine the gene expression levels. MTT, flow cytometry, and transwell assays were performed to explore the functions of miR-142-5p in HeLa cells. The potential target gene of miR-142-5p was investigated via luciferase reporter assays. The protein expression levels were analyzed by Western blotting. Results We found that miR-142-5p expression was elevated but LIM homeobox transcription factor 1 alpha (LMX1A) was decreased in cervical cancer tissues and cells. Overexpression of miR-142-5p or knockdown of LMX1A inhibited cell apoptosis, promoted cell proliferation, migration, invasion abilities, and activated the Wnt/β-catenin pathway. However, knockdown of miR-142-5p or overexpression of LMX1A showed opposite results. LMX1A was identified as a direct target of miR-142-5p by luciferase reporter assays. Finally, rescue experiments demonstrated that LMX1A overexpression attenuated the carcinogenic effect of miR-142-5p mimic on HeLa cells. Conclusions These findings suggested that miR-142-5p might be a cervical cancer oncogene and could serve as a potential therapeutic target for the treatment of cervical cancer.

scholarly journals HIV-1 Vpr protein upregulates microRNA-210-5p expression to induce G2 arrest by targeting TGIF2

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0261971
Author(s):  
Jialu Qiao ◽  
Qian Peng ◽  
Feng Qian ◽  
Qiang You ◽  
Lingyan Feng ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Transcription Factor ◽  
Gene Silencing ◽  
Luciferase Reporter ◽  
G2 Arrest ◽  
Host Interactions ◽  
Reporter Assays ◽  
New Evidence ◽  
Hiv 1

MicroRNAs (miRNAs) are important molecules that mediate virus-host interactions, mainly by regulating gene expression via gene silencing. Here, we demonstrated that HIV-1 infection upregulated miR-210-5p in HIV-1-inoculated cell lines and in the serum of HIV-1-infected individuals. Luciferase reporter assays and western blotting confirmed that a target protein of miR-210-5p, TGIF2, is regulated by HIV-1 infection. Furthermore, HIV-1 Vpr protein induced miR-210-5p expression. The use of a miR-210-5p inhibitor and TGIF2 overexpression showed that Vpr upregulated miR-210-5p and thereby downregulated TGIF2, which might be one of the mechanisms used by Vpr to induce G2 arrest. Moreover, we identified a transcription factor, NF-κB p50, which upregulated miR-210-5p in response to Vpr protein. In conclusion, we identified a mechanism whereby miR-210-5p, which is induced upon HIV-1 infection, targets TGIF2. This pathway was initiated by Vpr protein activating NF-κB p50, which promoted G2 arrest. These alterations orchestrated by miRNA provide new evidence on how HIV-1 interacts with its host during infection and increase our understanding of the mechanism by which Vpr regulates the cell cycle.

scholarly journals EGR1 modulated LncRNA HNF1A-AS1 drives glioblastoma progression via miR-22-3p/ENO1 axis

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Chunchun Ma ◽  
Hongliang Wang ◽  
Gang Zong ◽  
Jie He ◽  
Yuyang Wang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bioinformatic Analysis ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Clinical Samples ◽  
Regulation Mechanism ◽  
Functional Studies ◽  
Reporter Assays ◽  
Invasion Assays

AbstractAccumulating evidences revealed that long noncoding RNAs (lncRNAs) have been participated in cancer malignant progression, including glioblastoma multiforme (GBM). Despite much studies have found the precise biological role in the regulatory mechanisms of GBM, however the molecular mechanisms, particularly upstream mechanisms still need further elucidated. RT-QPCR, cell transfection, western blotting and bioinformatic analysis were executed to detect the expression of EGR1, HNF1A-AS1, miR-22-3p and ENO1 in GBM. Cell proliferation assay, colony formation assay, wound healing, migration and invasion assays were performed to detect the malignant characters of GBM cells. The molecular regulation mechanism was confirmed by luciferase reporter assay, ChIP and RIP. Finally, orthotopic mouse models were established to examine the effect of HNF1A-AS1 in vivo. In the current study, we analyzed clinical samples to show that the HNF1A-AS1 expression is upregulated and associated with poor patient survival in GBM. Functional studies revealed that HNF1A-AS1 knockdown markedly inhibits malignant phenotypes of GBM cells, whereas overexpression of HNF1A-AS1 exerts opposite effect. Mechanistically, the transcription factor EGR1 forced the HNF1A-AS1 expression by directly binding the promoter region of HNF1A-AS1. Furthermore, combined bioinformatics analysis with our mechanistic work, using luciferase reporter assays and RIP, we first demonstrated that HNF1A-AS1 functions as a competing endogenous RNA (ceRNA) with miR-22-3p to regulate ENO1 expression in GBM cells. HNF1A-AS1 directly binds to miR-22-3p and significantly inhibits miR-22-3p expression, while ENO1 expression was increased. miR-22-3p inhibitor offsets the HNF1A-AS1 silencing induced suppression in malignant behaviors of GBM cells. ENO1 was verified as a direct target of miR-22-3p and its expression levels was negatively with the prognosis in GBM patients. Taken together, our study illuminated the definite mechanism of HNF1A-AS1 in promoting GBM malignancy, and provided a novel therapeutic target for further clinical application.

scholarly journals Transcriptomic and metabolomic analysis provides insights into anthocyanin and pyocyanin accumulation in pear

2020 ◽  
Author(s):  
Zhen Zhang ◽  
Changping Tian ◽  
Ya Zhang ◽  
Chenzhiyu Li ◽  
Xi Li ◽  
...  
Keyword(s):  
Luciferase Reporter ◽  
Strongly Correlated ◽  
Mobility Shift ◽  
Data Set ◽  
Functional Studies ◽  
Nutritive Quality ◽  
Core Set ◽  
Reporter Assays ◽  
Arabidopsis Mutant

Abstract Background Pear is one of the most important fruit crops worldwide. Anthocyanins and procyanidins (PAs) are important secondary metabolites that affect the appearance and nutritive quality of pear. However, few studies have focused on the molecular mechanism underlying anthocyanin and PA accumulation in pear . Results We conducted metabolome and transcriptome analyses to identify candidate genes involved in anthocyanin and PA accumulation in young fruits of the pear cultivar ‘Clapp Favorite’ (CF) and its red mutation cultivar ‘Red Clapp Favorite’ ( RCF ). Gene–metabolite correlation analyses revealed a ‘core set’ of 20 genes that were strongly correlated with 10 anthocyanin and seven PA metabolites. Of these, PcGSTF12 was confirmed to be involved in anthocyanin and PA accumulation by complementation of the tt19-7 Arabidopsis mutant. Interestingly, PcGSTF12 was found to be responsible for the accumulation of procyanidin A3, but not petunidin 3, 5-diglucoside, opposite to the function of AtGSTs in Arabidopsis . Transformation with PcGSTF12 greatly promoted or repressed genes involved in anthocyanin and PA biosynthesis, regulation, and transport. Electrophoretic mobility shift and luciferase reporter assays confirmed positive regulation of PcGSTF12 by PcMYB114 . Conclusion These findings identify a core set of genes for anthocyanin and PA accumulation in pear. Of these, PcGSTF12 , was confirmed to be involved in anthocyanin and PA accumulation. Our results also identified an important anthocyanin and PA regulation node comprising two core genes, PcGSTF12 and PcMYB114 . These results provide novel insights into anthocyanin and PA accumulation in pear and represent a valuable data set to guide future functional studies and pear breeding.

scholarly journals Transcriptomic and metabolomic analysis provides insights into anthocyanin and pyocyanin accumulation in pear

2019 ◽  
Author(s):  
Shouqian Feng ◽  
Zhen Zhang ◽  
Changping Tian ◽  
Ya Zhang ◽  
Chenzhiyu Li ◽  
...  
Keyword(s):  
Luciferase Reporter ◽  
Mobility Shift ◽  
Data Set ◽  
Functional Studies ◽  
Nutritive Quality ◽  
Core Set ◽  
Reporter Assays ◽  
Arabidopsis Mutant ◽  
Core Genes

Abstract BackgroundPear is one of the most important fruit crops worldwide. Anthocyanins and procyanidins (PAs) are important secondary metabolic substances that affect the appearance and nutritive quality of pear. However, few studies have focused on the molecular mechanism underlying anthocyanin and PA accumulation in pear.ResultsWe conducted metabolome and transcriptome analyses to identify candidate genes involved in anthocyanin and PA accumulation in young pear fruits of ‘Clapp Favorite’ (CF) and its red mutation variety ‘Red Clapp Favorite’ (RCF). Gene–metabolite correlation analyses revealed a core set of 20 genes for pear anthocyanin and PA accumulation. One gene in the core set, PcGSTF12, was confirmed to be involved in anthocyanin and PA accumulation by complementation of the tt19-7 Arabidopsis mutant. Interestingly, PcGSTF12 was found to be responsible for procyanidin A3 accumulation, but not petunidin 3, 5-diglucoside accumulation, opposite to the function of AtGSTs in Arabidopsis. Transformation with PcGSTF12 greatly promoted or repressed genes involved in anthocyanin and PA biosynthesis, regulation, and transport. Electrophoretic mobility shift and luciferase reporter assays confirmed positive regulation of PcGSTF12 by PcMYB114.ConclusionThese findings identify a core set of genes for pear anthocyanin and PA accumulation. Of these, PcGSTF12, was confirmed to be involved in anthocyanin and PA accumulation. Furthermore, an important anthocyanin and PA regulation node was constructed by two core genes, PcGSTF12 and PcMYB114. These results provide novel insights into anthocyanin and PA accumulation in pear and represent a valuable data set to guide future functional studies and pear breeding.

scholarly journals Transcriptomic and metabolomic analysis provides insights into anthocyanin and procyanidin accumulation in pear

2020 ◽  
Author(s):  
Zhen Zhang ◽  
Changping Tian ◽  
Ya Zhang ◽  
Chenzhiyu Li ◽  
Xi Li ◽  
...  
Keyword(s):  
Luciferase Reporter ◽  
Strongly Correlated ◽  
Mobility Shift ◽  
Data Set ◽  
Functional Studies ◽  
Nutritive Quality ◽  
Core Set ◽  
Reporter Assays ◽  
Arabidopsis Mutant

Abstract Background Pear is one of the most important fruit crops worldwide. Anthocyanins and procyanidins (PAs) are important secondary metabolites that affect the appearance and nutritive quality of pear. However, few studies have focused on the molecular mechanism underlying anthocyanin and PA accumulation in pear . Results We conducted metabolome and transcriptome analyses to identify candidate genes involved in anthocyanin and PA accumulation in young fruits of the pear cultivar ‘Clapp Favorite’ (CF) and its red mutation cultivar ‘Red Clapp Favorite’ ( RCF ). Gene–metabolite correlation analyses revealed a ‘core set’ of 20 genes that were strongly correlated with 10 anthocyanin and seven PA metabolites. Of these, PcGSTF12 was confirmed to be involved in anthocyanin and PA accumulation by complementation of the tt19-7 Arabidopsis mutant. Interestingly, PcGSTF12 was found to be responsible for the accumulation of procyanidin A3, but not petunidin 3, 5-diglucoside, opposite to the function of AtGSTs in Arabidopsis . Transformation with PcGSTF12 greatly promoted or repressed genes involved in anthocyanin and PA biosynthesis, regulation, and transport. Electrophoretic mobility shift and luciferase reporter assays confirmed positive regulation of PcGSTF12 by PcMYB114 . Conclusion These findings identify a core set of genes for anthocyanin and PA accumulation in pear. Of these, PcGSTF12 , was confirmed to be involved in anthocyanin and PA accumulation. Our results also identified an important anthocyanin and PA regulation node comprising two core genes, PcGSTF12 and PcMYB114 . These results provide novel insights into anthocyanin and PA accumulation in pear and represent a valuable data set to guide future functional studies and pear breeding.

scholarly journals EGR2-mediated regulation of m6A reader IGF2BP proteins drive RCC tumorigenesis and metastasis via enhancing S1PR3 mRNA stabilization

2021 ◽  
Vol 12 (8) ◽  
Author(s):  
Yufan Ying ◽  
Xueyou Ma ◽  
Jiajie Fang ◽  
Shiming Chen ◽  
Weiyu Wang ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Renal Cell Cancer ◽  
Renal Cell ◽  
Growth Response ◽  
Bioinformatics Analysis ◽  
Cell Cancer ◽  
Early Growth ◽  
Luciferase Reporter ◽  
Early Growth Response ◽  
Reporter Assays

AbstractEmerging discoveries of dynamic and reversible N6-methyladenosine (m6A) modification on RNA in mammals have revealed the key roles of the modification in human tumorigenesis. As known m6A readers, insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) are upregulated in most cancers and mediates the enhancement of m6A-modified mRNAs stability. However, the mechanisms of IGF2BPs in renal cell cancer (RCC) still remain unclear. Bioinformatic analysis and RT-qPCR were performed to evaluate the expression of IGF2BPs and m6A writer Wilms tumor 1-associating protein (WTAP) in RCC samples and its correlation with patient prognosis. In vitro, in vivo biological assays were performed to investigate the functions of IGF2BPs and WTAP in RCC. Chromatin immunoprecipitation-qPCR (ChIP-qPCR) combined with bioinformatics analysis and following western blot assay, dual-luciferase reporter assays were performed to validate the regulatory relationships between transcription factor (TF) early growth response 2 (EGR2) and potential target genes IGF2BPs. RNA sequencing (RNA-seq), methylated RNA immunoprecipitation-qPCR (MERIP-qPCR), RIP-qPCR, m6A dot blot, and dual-luciferase reporter assays combined with bioinformatics analysis were employed to screen and validate the direct targets of IGF2BPs and WTAP. Here, we showed that early growth response 2 (EGR2) transcription factor could increase IGF2BPs expression in RCC. IGF2BPs in turn regulated sphingosine-1-phosphate receptor 3 (S1PR3) expression in an m6A-dependent manner by enhancing the stability of S1PR3 mRNA. They also promoted kidney tumorigenesis via PI3K/AKT pathway. Furthermore, IGF2BPs and WTAP upregulation predicted poor overall survival in RCC. Our studies showed that the EGR2/IGF2BPs regulatory axis and m6A-dependent regulation of S1PR3-driven RCC tumorigenesis, which enrich the m6A-modulated regulatory network in renal cell cancer. Together, our findings provide new evidence for the role of N6-methyladenosine modification in RCC.

scholarly journals The POU Transcription Factor POU-M2 Regulates Vitellogenin Receptor Gene Expression in the Silkworm, Bombyx mori

Genes ◽  
2020 ◽  
Vol 11 (4) ◽  
pp. 394
Author(s):  
Guanwang Shen ◽  
Enxiang Chen ◽  
Xiaocun Ji ◽  
Lina Liu ◽  
Jianqiu Liu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Bombyx Mori ◽  
Receptor Gene ◽  
Expression Patterns ◽  
Egg Laying ◽  
Luciferase Reporter ◽  
Mobility Shift ◽  
Reverse Transcription Pcr ◽  
Vitellogenin Receptor ◽  
Reporter Assays

Vitellogenin receptors (VgRs) play critical roles in egg formation by transporting vitellogenin (Vg) into oocytes in insects. Although the function of VgR in insects is well studied, the transcriptional regulation of this gene is still unclear. Here, we cloned the promoter of the VgR gene from Bombyx mori (BmVgR), and predicted many POU cis-response elements (CREs) in its promoter. Electrophoretic mobility shift and chromatin immunoprecipitation assays showed that the POU transcription factor POU-M2 bound directly to the CREs of the promoter. Overexpression of POU-M2 in an ovarian cell line (BmNs) enhanced BmVgR transcription and promoter activity detected by quantitative reverse transcription PCR and luciferase reporter assays. Analyses of expression patterns indicated that POU-M2 was expressed in ovary at day two of wandering stage initially, followed by BmVgR. RNA interference of POU-M2 significantly reduced the transcription of BmVgR in ovary and egg-laying rate. Our results suggest a novel function for the POU factor in silkworm oogenesis by its involvement in BmVgR regulation and expands the understanding of POU factors in insect VgR expression.

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