miR-125b enhances metastasis and progression of cancer via the TXNIP and HIF1α pathway in pancreatic cancer

2021 ◽  
pp. 1-12
Author(s):  
Pengli Wang ◽  
Dan Zheng ◽  
Hongyang Qi ◽  
Qi Gao
Keyword(s):  
Pancreatic Cancer ◽  
Hypoxia Inducible Factor ◽  
Immunofluorescence Assay ◽  
Luciferase Assay ◽  
Interacting Protein ◽  
Over Expression ◽  
Thioredoxin Interacting Protein ◽  
Cellular Processes ◽  
And Migration

BACKGROUND: MicroRNAs (miRNAs) play potential role in the development of various types of cancer conditions including pancreatic cancer (PC) targeting several cellular processes. Present study was aimed to evaluate function of miR-125b and the mechanism involved in PC. METHODS: Cell migration, MTT and BrdU study was done to establish the migration capability, cell viability and cell proliferation respectively. Binding sites for miR-125b were recognized by luciferase assay, expression of protein by western blot and immunofluorescence assay. In vivo study was done by BALB/c nude xenograft mice for evaluating the function of miR-125b. RESULTS: The study showed that expression of miR-125b was elevated in PC cells and tissues, and was correlated to proliferation and migration of cells. Also, over-expression of miR-125b encouraged migration, metastasis and proliferation of BxPC-3 cells, the suppression reversed it. We also noticed that thioredoxin-interacting protein (TXNIP) was the potential target of miR-125b. The outcomes also suggested that miR-125b governed the expression of TXNIP inversely via directly attaching to the 3′-UTR activating hypoxia-inducible factor 1α (HIF1α). Looking into the relation between HIF1α and TXNIP, we discovered that TXNIP caused the degradation and export of HIF1α by making a complex with it. CONCLUSION: The miR-125b-TXNIP-HIF1α pathway may serve useful strategy for diagnosing and treating PC.

scholarly journals Hypoxia-related gene FUT11 promoted pancreatic cancer progression via maintaining the stabilization of PDK1

2021 ◽  
Author(s):  
Wenpeng Cao ◽  
Zhirui Zeng ◽  
Runsang Pan ◽  
Zhiwei He ◽  
Hao Wu ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cell Growth ◽  
Western Blot ◽  
Cancer Progression ◽  
Hypoxia Inducible Factor ◽  
Pyruvate Dehydrogenase Kinase ◽  
Cell Counting ◽  
Luciferase Assay

Abstract Background Hypoxia participated in the occurrence and development of pancreatic cancer (PC). However, genes associated with hypoxia respond and their regulated mechanism in PC cells were unclear. The current research was aimed to illuminate the role and hypoxia regulated mechanism of fucosyltransferase 11 (FUT11) in the progression of PC. Methods After predicting FUT11 as a key hypoxia associated gene in PC using bioinformatics analysis. The expression of FUT11 in PC using quantitative real-time fluorescent PCR, western blot and immunohistochemistry. The effects of FUT11 on PC cells proliferation and migration under normoxia and hypoxia were detected using Cell Counting Kit 8, 5-ethynyl-2’-deoxyuridine assay, colony formation assay and transwell assay. Spleen capsule injected liver metastasis and subcutaneously injected model were performed to confirm the effects of FUT11 in vivo. Furthermore, western blot, luciferase assay and immunoprecipitation were performed to explore the regulated relationship among FUT11, hypoxia-inducible factor 1α (HIF1α) and pyruvate dehydrogenase kinase 1 (PDK1) in PC. Results FUT11 was markedly increased of PC cells in hypoxia, up-regulated in the PC clinical tissues, and predicted a poor outcome. Inhibition of FUT11 reduced PC cell growth and mobility of PC cells under normoxia and hypoxia conditions in vitro, and growth and mobility in vivo. FUT11 bind with PDK1 and regulated the expression PDK1 under normoxia and hypoxia. FUT11 knockdown significantly increased the degradation rate of PDK1 under hypoxia, while treatment with MG132 can relieve the degradation of PDK1 induced by FUT11 knockdown. Overexpression of PDK1 in PC cells under hypoxia conditions reversed the suppressiv impacts of FUT11 knockdown on PC cell growth and mobility. In addition, HIF1α bound to the enhancer of FUT11 and increased its expression, as well as co-expressing with FUT11 in PC tissues. Furthermore, overexpress of FUT11 partially rescued the suppressiv effects of HIF1α knockdown on PC cell growth and mobility in hypoxia conditions. Conclusion Our data further implicate that hypoxia-induced FUT11 in PC contributes to proliferation and metastasis by maintaining the stability of PDK1, and suggest FUT11 maybe a novel and effective target for treatment of pancreatic cancer.

scholarly journals MiR-598 Suppresses Invasion and Migration by Negative Regulation of Derlin-1 and Epithelial-Mesenchymal Transition in Non-Small Cell Lung Cancer

2018 ◽  
Vol 47 (1) ◽  
pp. 245-256 ◽  
Author(s):  
Fengming Yang ◽  
Ke Wei ◽  
Zhiqiang Qin ◽  
Weitao Liu ◽  
Chuchu Shao ◽  
...  
Keyword(s):  
Cell Lines ◽  
Epithelial Mesenchymal Transition ◽  
Luciferase Assay ◽  
Small Cell Lung ◽  
Mesenchymal Transition ◽  
Invasion And Migration ◽  
Over Expression ◽  
And Migration

Background/Aims: MicroRNAs regulate a wide range of biological processes of non-small cell lung cancer (NSCLC). Although miR-598 has been reported to act as a suppressor in osteosarcoma and colorectal cancer, the physiological function of miR-598 in NSCLC remains unknown. In this study, the role of miR-598 in NSCLC was investigated. Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to estimate the expression of miR-598 and Derlin-1 (DERL1) in both NSCLC tissues and cell lines. Immunohistochemistry (IHC) analyzed the association between the miR-598 expression and epithelial-mesenchymal transition (EMT) hallmark genes (E-cadherin, Vimentin) by staining the tumors representative of the high- and low-expression groups. The effect of miR-598 and DERL1 on invasion and migration was determined in vitro using transwell and wound-healing assays. The molecular mechanism underlying the relevance between miR-598 and DERL1 was elucidated by luciferase assay and Western blot. Western blot assessed the expression levels of EMT hallmark genes in cell lines. Xenograft tumor formation assay was conducted as an in vivo experiment. Results: In this study, a relatively low level of miR-598 and high DERL1 expressions were found in NSCLC specimens and cell lines. IHC results established a positive correlation between the miR-598 expression and E-cadherin and a negative with Vimentin. DERL1 was verified as a direct target of miR-598 by luciferase assay. In vitro, the over-expression of miR-598 negatively regulated DERL1 and EMT for the suppression of invasion and migration. In vivo, the over-expression of miR-598 could inhibit tumor cell metastasis in NSCLC. Conclusions: These findings for the first time revealed that miR-598, as a tumor suppressor, negatively regulate DERL1 and EMT to suppress the invasion and migration in NSCLC, thereby putatively serving as a novel therapeutic target for NSCLC clinical treatment.

Identification of Thioredoxin-Interacting Protein 1 as a Hypoxia-Inducible Factor 1α-Induced Gene in Pancreatic Cancer

Pancreas ◽  
2008 ◽  
Vol 36 (2) ◽  
pp. 178-186 ◽  
Author(s):  
Amanda F. Baker ◽  
Mei Y. Koh ◽  
Ryan R. Williams ◽  
Brian James ◽  
Huamin Wang ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Hypoxia Inducible Factor ◽  
Interacting Protein ◽  
Hypoxia Inducible Factor 1Α ◽  
Thioredoxin Interacting Protein

scholarly journals NR5A2 transcriptional activation by BRD4 promotes pancreatic cancer progression by upregulating GDF15

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Feng Guo ◽  
Yingke Zhou ◽  
Hui Guo ◽  
Dianyun Ren ◽  
Xin Jin ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Cancer Progression ◽  
Transcriptional Activation ◽  
Ductal Adenocarcinoma ◽  
Pancreatic Cancer Cells ◽  
Gene Sets ◽  
And Migration

AbstractNR5A2 is a transcription factor regulating the expression of various oncogenes. However, the role of NR5A2 and the specific regulatory mechanism of NR5A2 in pancreatic ductal adenocarcinoma (PDAC) are not thoroughly studied. In our study, Western blotting, real-time PCR, and immunohistochemistry were conducted to assess the expression levels of different molecules. Wound-healing, MTS, colony formation, and transwell assays were employed to evaluate the malignant potential of pancreatic cancer cells. We demonstrated that NR5A2 acted as a negative prognostic biomarker in PDAC. NR5A2 silencing inhibited the proliferation and migration abilities of pancreatic cancer cells in vitro and in vivo. While NR5A2 overexpression markedly promoted both events in vitro. We further identified that NR5A2 was transcriptionally upregulated by BRD4 in pancreatic cancer cells and this was confirmed by Chromatin immunoprecipitation (ChIP) and ChIP-qPCR. Besides, transcriptome RNA sequencing (RNA-Seq) was performed to explore the cancer-promoting effects of NR5A2, we found that GDF15 is a component of multiple down-regulated tumor-promoting gene sets after NR5A2 was silenced. Next, we showed that NR5A2 enhanced the malignancy of pancreatic cancer cells by inducing the transcription of GDF15. Collectively, our findings suggest that NR5A2 expression is induced by BRD4. In turn, NR5A2 activates the transcription of GDF15, promoting pancreatic cancer progression. Therefore, NR5A2 and GDF15 could be promising therapeutic targets in pancreatic cancer.

scholarly journals A novel thermostable beetle luciferase based cytotoxicity assay

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Sunju Choi ◽  
Hittu Matta ◽  
Ramakrishnan Gopalakrishnan ◽  
Venkatesh Natarajan ◽  
Songjie Gong ◽  
...  
Keyword(s):  
Membrane Integrity ◽  
Cytotoxicity Assay ◽  
Luciferase Assay ◽  
Target Cells ◽  
Over Expression ◽  
Click Beetle ◽  
In Vivo Bioluminescence Imaging ◽  
Cost Efficient ◽  
Dying Cells

AbstractCytotoxicity assays are essential for the testing and development of novel immunotherapies for the treatment of cancer. We recently described a novel cytotoxicity assay, termed the Matador assay, which was based on marine luciferases and their engineered derivatives. In this study, we describe the development of a new cytotoxicity assay termed ‘Matador-Glo assay’ which takes advantage of a thermostable variant of Click Beetle Luciferase (Luc146-1H2). Matador-Glo assay utilizes Luc146-1H2 and D-luciferin as the luciferase-substrate pair for luminescence detection. The assay involves ectopic over-expression of Luc146-1H2 in the cytosol of target cells of interest. Upon damage to the membrane integrity, the Luc146-1H2 is either released from the dead and dying cells or its activity is preferentially measured in dead and dying cells. We demonstrate that this assay is simple, fast, specific, sensitive, cost-efficient, and not labor-intensive. We further demonstrate that the Matador-Glo assay can be combined with the marine luciferase-based Matador assay to develop a dual luciferase assay for cell death detection. Finally, we demonstrate that the Luc146-1H2 expressing target cells can also be used for in vivo bioluminescence imaging applications.

scholarly journals The pro-oncogenic adaptor CIN85 inhibits hypoxia-inducible factor prolyl hydroxylase-2

2018 ◽  
Author(s):  
Nina Kozlova ◽  
Daniela Mennerich ◽  
Anatoly Samoylenko ◽  
Elitsa Y. Dimova ◽  
Peppi Koivunen ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Survival Function ◽  
Hypoxia Inducible Factor ◽  
Adaptor Protein ◽  
Prolyl Hydroxylase ◽  
Binding Partner ◽  
And Migration ◽  
Hif Prolyl Hydroxylase ◽  
Promote Breast Cancer

SummaryThe EGFR-adaptor protein CIN85 has been shown to promote breast cancer malignancy and hypoxia-inducible factor (HIF) stability. However, the mechanisms underlying cancer promotion remain ill-defined. Here, we show that CIN85 is a novel binding partner of the main HIF-prolyl hydroxylase PHD2, but not of PHD1 or PHD3. Mechanistically, the N-terminal SH3 domains of CIN85 interact with the proline-arginine rich region within the N-terminus of PHD2, thereby inhibiting PHD2 activity and HIF-degradation. This activity is essential in vivo, as specific loss of the CIN85-PHD2 interaction in CRISPR/Cas9 edited cells affected growth and migration properties as well as tumor growth in mice. Overall, we discovered a previously unrecognized tumor growth checkpoint that is regulated by CIN85-PHD2, and uncovered an essential survival function in tumor cells linking growth factor adaptors with hypoxia signaling.

scholarly journals Positive feedback between lncRNA FLVCR1-AS1 and KLF10 may inhibit pancreatic cancer progression via the PTEN/AKT pathway

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Jiewei Lin ◽  
Shuyu Zhai ◽  
Siyi Zou ◽  
Zhiwei Xu ◽  
Jun Zhang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Pancreatic Cancer ◽  
Cell Proliferation ◽  
Cancer Progression ◽  
Positive Feedback ◽  
Molecular Mechanisms ◽  
Tumor Tissues ◽  
And Migration

Abstract Background FLVCR1-AS1 is a key regulator of cancer progression. However, the biological functions and underlying molecular mechanisms of pancreatic cancer (PC) remain unknown. Methods FLVCR1-AS1 expression levels in 77 PC tissues and matched non-tumor tissues were analyzed by qRT-PCR. Moreover, the role of FLVCR1-AS1 in PC cell proliferation, cell cycle, and migration was verified via functional in vitro and in vivo experiments. Further, the potential competitive endogenous RNA (ceRNA) network between FLVCR1-AS1 and KLF10, as well as FLVCR1-AS1 transcription levels, were investigated. Results FLVCR1-AS1 expression was low in both PC tissues and PC cell lines, and FLVCR1-AS1 downregulation was associated with a worse prognosis in patients with PC. Functional experiments demonstrated that FLVCR1-AS1 overexpression significantly suppressed PC cell proliferation, cell cycle, and migration both in vitro and in vivo. Mechanistic investigations revealed that FLVCR1-AS1 acts as a ceRNA to sequester miR-513c-5p or miR-514b-5p from the sponging KLF10 mRNA, thereby relieving their suppressive effects on KLF10 expression. Additionally, FLVCR1-AS1 was shown to be a direct transcriptional target of KLF10. Conclusions Our research suggests that FLVCR1-AS1 plays a tumor-suppressive role in PC by inhibiting proliferation, cell cycle, and migration through a positive feedback loop with KLF10, thereby providing a novel therapeutic strategy for PC treatment.

scholarly journals Over-expression of ANP32E is associated with poor prognosis of pancreatic cancer and promotes cell proliferation and migration through regulating β-catenin

2020 ◽  
Author(s):  
Jianwei Zhang ◽  
Zhongmin Lan ◽  
Guotong Qiu ◽  
Hu Ren ◽  
Yajie Zhao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Pancreatic Cancer ◽  
Cell Proliferation ◽  
Poor Prognosis ◽  
Cell Cycle Progression ◽  
Over Expression ◽  
Cell Proliferation And Migration ◽  
And Migration ◽  
Cancer Tissues ◽  
Proliferation And Migration

Abstract Background: Pancreatic cancer is a malignant tumor with high mortality. Acidic nuclear phosphoprotein 32 family member E (ANP32E), a specific H2A.Z chaperone, has been shown to contribute to breast cancer development. However, the significance of ANP32E in pancreatic cancer is poorly understood. This study aimed to investigate the role of ANP32E in pancreatic cancer. Methods: The expression of ANP32E in 179 pancreatic cancer tissues and 171 normal tissues, and the correlation between ANP32E expression and patients’ survival were analyzed from the TCGA database. ANP32E was over-expressed and silenced using lentivirus. siRNA was used to knock down β-catenin. CCK8, colony formation, cell cycle and transwell experiments were performed to determine cell proliferation and migration. qRT-PCR and Western blot were conducted to detect mRNA and protein expression. Results: ANP32E was up-regulated in pancreatic cancer tissues and cells. Up-regulation of ANP32E predicted poor prognosis in pancreatic cancer patients. Lentivirus-mediated knockdown of ANP32E suppressed the proliferation, colony growth and migration of PANC1 and MIA cells. By contrast, ANP32E over-expression promoted the proliferation and migration of both cells. In addition, ANP32E accelerated the cell cycle progression in PANC1 and MIA cells. Molecular experiments showed that ANP32E activated β-catenin/cyclin D1 signaling. Silencing of β-catenin reduced cell proliferation and migration in ANP32E over-expressed cells. Conclusion: Our results propose that ANP32E functions as an oncogene in pancreatic cancer via activating β-catenin.

scholarly journals Arabidopsis ALIX is required for the endosomal localization of the deubiquitinating enzyme AMSH3

2015 ◽  
Vol 112 (40) ◽  
pp. E5543-E5551 ◽  
Author(s):  
Kamila Kalinowska ◽  
Marie-Kristin Nagel ◽  
Kaija Goodman ◽  
Laura Cuyas ◽  
Franziska Anzenberger ◽  
...  
Keyword(s):  
Cellular Level ◽  
Deubiquitinating Enzymes ◽  
Interacting Protein ◽  
Endosomal Sorting ◽  
Cellular Processes ◽  
Late Endosomes ◽  
Knockout Mutants ◽  
Important Regulator

Ubiquitination is a signal for various cellular processes, including for endocytic degradation of plasma membrane cargos. Ubiquitinating as well as deubiquitinating enzymes (DUBs) can regulate these processes by modifying the ubiquitination status of target protein. Although accumulating evidence points to the important regulatory role of DUBs, the molecular basis of their regulation is still not well understood. Associated molecule with the SH3 domain of signal transduction adaptor molecule (STAM) (AMSH) is a conserved metalloprotease DUB in eukaryotes. AMSH proteins interact with components of the endosomal sorting complex required for transport (ESCRT) and are implicated in intracellular trafficking. To investigate how the function of AMSH is regulated at the cellular level, we carried out an interaction screen for the Arabidopsis AMSH proteins and identified the Arabidopsis homolog of apoptosis-linked gene-2 interacting protein X (ALIX) as a protein interacting with AMSH3 in vitro and in vivo. Analysis of alix knockout mutants in Arabidopsis showed that ALIX is essential for plant growth and development and that ALIX is important for the biogenesis of the vacuole and multivesicular bodies (MVBs). Cell biological analysis revealed that ALIX and AMSH3 colocalize on late endosomes. Although ALIX did not stimulate AMSH3 activity in vitro, in the absence of ALIX, AMSH3 localization on endosomes was abolished. Taken together, our data indicate that ALIX could function as an important regulator for AMSH3 function at the late endosomes.

Abstract 11: Thioredoxin-Interacting Protein Is Required for VEGF-Induced Angiogenesis Through Regulation of VEGFR2 Internalization

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Shin-Young Park ◽  
Chen Yan ◽  
Bradford C Berk
Keyword(s):  
Umbilical Vein ◽  
Fold Increase ◽  
Tube Formation ◽  
In Vivo Studies ◽  
Vegf Receptor ◽  
Cell Fractionation ◽  
Interacting Protein ◽  
Thioredoxin Interacting Protein

Introduction— Thioredoxin-interacting protein (TXNIP) is an arrestin-like scaffold protein. We have shown previously that it is necessary for the transactivation of the vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) as well as promoting the migration and survival of endothelial cells (ECs). However, its roles in VEGF-induced angiogenesis and in vivo studies of TXNIP function have not been elucidated. Hypothesis— TXNIP regulates VEGF-mediated angiogenesis through modulation of angiogenic signaling pathways in ECs. Methods and Results— To determine the functions of TXNIP in ECs, we generated endothelial-specific TXNIP knockout (EC-TXNIP KO) mice (TXNIPflox/flox: Tie2-Cre/+). These mice displayed impaired capillary growth of the retinal vasculature compared to control mice. Furthermore, aortic rings from EC-TXNIP KO mice exhibited fewer and shorter vascular sprouts than those in control mice. To investigate the role of TXNIP in the regulation of VEGF-induced angiogenesis, we determined the subcellular localization of TXNIP in human umbilical vein EC (HUVEC). Immunofluorescence and cell fractionation studies revealed that upon VEGF stimulation (10ng/ml). TXNIP translocated from cytoplasm to the plasma membrane. There was a 9 fold increase of membrane associated TXNIP with a peak at 15 minutes compared to non-VEGF treatment cells. We hypothesized that membrane associated TXNIP may modulate VEGFR2 internalization and thereby affect VEGF-induced signaling and angiogenesis. To investigate this, we performed in vitro cell surface biotinylation assays in HUVEC. VEGFR2 internalization was decreased by 65% in TXNIP siRNA knockdown cells compared to control siRNA treated cells following VEGF stimulation. Consistent with this result, VEGF-induced phosphorylation of VEGFR2, PLCγ and ERK1/2 was decreased by knockdown of TXNIP. Significantly, TXNIP knockdown inhibited VEGF-induced proliferation and tube formation in vitro. Conclusion— Our results suggest that TXNIP can modulate VEGF-induced angiogenesis and signaling by regulation of VEGFR2 internalization.

Sign in / Sign up

Export Citation Format

Share Document