MicroRNA-455-3p mediates GATA3 tumor suppression in mammary epithelial cells by inhibiting TGF-? signaling.
ABSTRACT: GATA3 is a basic and essential transcription factor that regulates many pathophysiological processes and is required for the development of mammary luminal epithelial cells. Loss-of-function GATA3 alterations in breast cancer are associated with poor prognosis. Here, we sought to understand the tumor-suppressive functions GATA3 normally performs. We discovered a role for GATA3 in suppressing epithelial-to-mesenchymal transition (EMT) in breast cancer by activating miR-455-3p expression. Enforced expression of miR-455-3p alone partially prevented EMT induced by transforming growth factor ? (TGF-?) both in cells and tumor xenografts by directly inhibiting key components of TGF-? signaling. Pathway and biochemical analyses showed that one miRNA-455-3p target, the TGF-?-induced protein ZEB1, recruits the Mi-2/nucleosome remodeling and deacetylase (NuRD) complex to the promotor region of miR-455 to strictly repress the GATA3-induced transcription of this microRNA. Considering that ZEB1 enhances TGF-? signaling, we delineated a double-feedback interaction between ZEB1 and miR-455-3p, in addition to the repressive effect of miR-455-3p on TGF-? signaling. Our study revealed that a feedback loop between these two axes, specifically GATA3-induced miR-455-3p expression, could repress ZEB1 and its recruitment of NuRD (MTA1) to suppress miR-455, which ultimately regulates TGF-? signaling. In conclusion, we identified that miR-455-3p plays a pivotal role in inhibiting the EMT and TGF-? signaling pathway and maintaining cell differentiation. This forms the basis of that miR-455-3p might be a promising therapeutic intervention for breast cancer.
Project description:MiR-455-3p has been reported to suppress the expression of ROCK2, HOXB5, hTERT, and RUNX2, which are involved in multiple biological processes including renal fibrosis, chondrogenic development and differentiation, Alzheimer’s disease, preeclampsia, and tumorigenesis. Here, we showed that miR-455-3p broadly inhibited TGF-beta-induced EMT by repressing the expression of Smad2, ZEB1, and HDAC2 in breast cancer cells, and that miR-455-3p loss-of-function promotes the acquisition of cell invasive properties and the ability to metastasize. These results indicate that miR-455-3p represents an important node that inhibits metastasis of breast cancer. Overall design: MiR-455-3p mimics or controls were transfected into MCF-7 cells (n=3 for each group) for 48 hours. Total RNA was isolated using Trizol reagent (Invitrogen, USA). Agilent 2100 Bioanalyzer (Agilent Technologies, USA) was used to evaluate the RNA quality.
Project description:Accumulating evidence indicates long noncoding RNAs (lncRNA) play a vital role in tumor progression. However, the role of linc00645-induced accelerated malignant behavior in glioblastoma (GBM) remains unknown. In the present study, linc00645 expression was significantly upregulated in GBM tissues and cell lines. High level of linc00645 was associated with poor overall survival in GBM patients. Knockdown of linc00645 suppressed the proliferation, stemness, migration, invasion, and reversed transforming growth factor (TGF)-?-induced motility of glioma cell lines. Furthermore, linc00645 directly interacted with miR-205-3p and upregulated of miR-205-3p impeded efficiently the increase of ZEB1 induced by linc00645 overexpression. Moreover, knockdown of linc00645 significantly suppressed the progression of glioma cells in vivo. miR-205-3p was a target of linc00645 and linc00645 modulates TGF-?-induced glioma cell migration and invasion via miR-205-3p. Taken together, our findings identified the linc00645/miR-205-3p/ZEB1 signaling axis as a key player in EMT of glioma cells triggered by TGF-?. These data elucidated that linc00645 plays an oncogenic role in glioma and it may serve as a prognostic biomarker and a potential therapeutic target for the treatment of glioma in humans.
Project description:MicroRNAs (miRNAs, miR) play a key role in the pathogenesis of osteoarthritis (OA). Few studies have examined the regulatory role of P21-activated kinases (PAKs), a family of serine/threonine kinases, in OA. The aim of this study was to determine whether miR-455-3p can regulate cartilage degeneration in OA by targeting PAK2. MiR-455-3p knockout mice showed significant degeneration of the knee cartilage. MiR-455-3p expression increased and PAK2 expression decreased in the late stage of human adipose-derived stem cell (hADSC) chondrogenesis and in chondrocytes affected by OA. Furthermore, in both miR-455-3p-overexpressing chondrocytes and PAK2-suppressing chondrocytes, cartilage-specific genes were upregulated, and hypertrophy-related genes were downregulated. A luciferase reporter assay confirmed that miR-455-3p regulates PAK2 expression by directly targeting the 3'-untranslated regions (3'UTRs) of PAK2 mRNA. IPA-3, a PAK inhibitor, inhibited cartilage degeneration due to OA. Moreover, suppressing PAK2 promoted R-Smad activation in the TGF/Smad signaling pathway in chondrocytes. Altogether, our results suggest that miR-455-3p promotes TGF-?/Smad signaling in chondrocytes and inhibits cartilage degeneration by directly suppressing PAK2. These results thus indicate that miR-455-3p and PAK2 are novel potential therapeutic agents and targets, respectively, for the treatment of OA.
Project description:OBJECTIVE:The long non-coding RNA zinc finger E-box-binding homeobox 1 antisense 1 (ZEB1-AS1) acts as an oncogenic regulator in many human tumours. In the present study, we identify the role and potential molecular biological mechanisms of ZEB1-AS1 in colon adenocarcinoma (COAD). METHODS:QRT-PCR was used to detect the expression of ZEB1-AS1, miR-455-3p and p21-activated kinases 2 (PAK2) in COAD tissues. CCK8 assay, EdU assay, transwell assay and scratch wound assay were used to explore the biological function of ZEB1-AS1 in COAD cells. Bioinformatics, luciferase reporter assays and an RNA pull-down assay were used to demonstrate the mechanism of ZEB1-AS1. We further explore the role of ZEB1-AS1 in vivo though xenograft tumour assay. RESULTS:We found that ZEB1-AS1 expression was significantly up-regulated in COAD tissues, and high ZEB1-AS1 level was correlated with the poor prognosis of COAD patients. MiR-455-3p plays an anti-cancer role in COAD by targeting PAK2. We confirmed that ZEB1-AS1 promotes PAK2 expression by sponging miR-455-3p, thus facilitating COAD cell growth and metastasis. CONCLUSIONS:To sum up, this result illustrates the novel molecular mechanism of ZEB1-AS1 in COAD and provides a new target for the diagnosis and treatment of COAD patients.
Project description:Background:Pancreatic cancer (PC) is a highly invasive tumor with a poor prognosis, short overall survival rate and few chemotherapeutic choices. Despite the importance of finding ways to treat pancreatic cancer, the mechanisms of tumor progression have not been fully elucidated. microRNA-455-3p (miR-455-3p) has been reported to play an important role in several cancers, but its function in pancreatic cancer remains unclear. Methods:To investigate the biological functions, miRNAs mimics or inhibitors were transfected into pancreatic cancer cells. Flow cytometry was used to detect cell apoptosis. Wound healing and Transwell assays were employed to observe cell invasion and migration abilities. The expression of Bcl-2, Bax, caspase-3, E-cadherin, N-cadherin, Snail, ?-Catenin, c-Myc and Cyclin D1 were evaluated by qPCR and Western blot. Results:We confirmed that inhibition of miR-455-3p decreases cell apoptosis and increases cell migration, invasion and EMT of pancreatic cancer, whereas forced overexpression of miR-455-3p has the opposite effect. Furthermore, we demonstrated that the tumor suppression effects of miR-455-3p were partially reversed by TAZ overexpression. In addition, miR-455-3p led to inactivation of Wnt/?-catenin signaling in pancreatic cancer cells, and TAZ overexpression restored the inhibition of Wnt/?-catenin signaling. Conclusion:Taken together, our data demonstrated that miR-455-3p functions as an important tumor suppressor that suppresses the Wnt/?-catenin signaling pathway via TAZ to inhibit tumor progression in pancreatic cancer. We conclude that the miR-455-3p/TAZ/Wnt axis may be a potential therapeutic target for pancreatic cancer.
Project description:BACKGROUND:The plasticity of cancer stem cells (CSCs)/tumor-initiating cells (T-ICs) suggests that multiple CSC/T-IC subpopulations exist within a tumor and that multiple oncogenic pathways collaborate to maintain the CSC/T-IC state. Here, we aimed to identify potential therapeutic targets that concomitantly regulate multiple T-IC subpopulations and CSC/T-IC-associated pathways. METHODS:A chemoresistant patient-derived xenograft (PDX) model of human esophageal squamous cell carcinoma (ESCC) was employed to identify microRNAs that contribute to ESCC aggressiveness. The oncogenic effects of microRNA-455-3p (miR-455-3p) on ESCC chemoresistance and tumorigenesis were examined by in vivo and in vitro chemoresistance, tumorsphere formation, side-population, and in vivo limiting dilution assays. The roles of miR-455-3p in activation of the Wnt/?-catenin and transforming growth factor-? (TGF-?)/Smad pathways were determined by luciferase and RNA immunoprecipitation assays. RESULTS:We found that miR-455-3p played essential roles in ESCC chemoresistance and tumorigenesis. Treatment with a miR-455-3p antagomir dramatically chemosensitized ESCC cells and reduced the subpopulations of CD90+ and CD271+ T-ICs via deactivation of multiple stemness-associated pathways, including Wnt/?-catenin and TGF-? signaling. Importantly, miR-455-3p exhibited aberrant upregulation in various human cancer types, and was significantly associated with decreased overall survival of cancer patients. CONCLUSIONS:Our results demonstrate that miR-455-3p functions as an oncomiR in ESCC progression and may provide a potential therapeutic target to achieve better clinical outcomes in cancer patients.
Project description:p53 suppresses tumor progression and metastasis. Epithelial-mesenchymal transition (EMT) is a key process in tumor progression and metastasis. The transcription factors ZEB1 and ZEB2 promote EMT. Here, we show that p53 suppresses EMT by repressing expression of ZEB1 and ZEB2. By profiling 92 primary hepatocellular carcinomas (HCCs) and 9 HCC cell lines, we found that p53 up-regulates microRNAs (miRNAs), including miR-200 and miR-192 family members. The miR-200 family members transactivated by p53 then repress ZEB1/2 expression. p53-regulated miR-192 family members also repress ZEB2 expression. Inhibition or overexpression of the miRNAs affects p53-regulated EMT by altering ZEB1 and ZEB2 expression. Our findings indicate that p53 can regulate EMT, and that p53-regulated miRNAs are critical mediators of p53-regulated EMT.
Project description:<h4>Background</h4>Lung fibroblasts are involved in extracellular matrix homeostasis, which is mainly regulated by transforming growth factor-beta (TGF-?), and are therefore crucial in lung tissue repair and remodeling. Abnormal repair and remodeling has been observed in lung diseases like COPD. As miRNA levels can be influenced by TGF-?, we hypothesized that TGF-? influences miRNA expression in lung fibroblasts, thereby affecting their function.<h4>Materials and methods</h4>We investigated TGF-?1-induced miRNA expression changes in 9 control primary parenchymal lung fibroblasts using miRNA arrays. TGF-?1-induced miRNA expression changes were validated and replicated in an independent set of lung fibroblasts composted of 10 controls and 15 COPD patients using qRT-PCR. Ago2-immunoprecipitation followed by mRNA expression profiling was used to identify the miRNA-targetomes of unstimulated and TGF-?1-stimulated primary lung fibroblasts (n = 2). The genes affected by TGF-?1-modulated miRNAs were identified by comparing the miRNA targetomes of unstimulated and TGF-?1-stimulated fibroblasts.<h4>Results</h4>Twenty-nine miRNAs were significantly differentially expressed after TGF-?1 stimulation (FDR<0.05). The TGF-?1-induced miR-455-3p and miR-21-3p expression changes were validated and replicated, with in addition, lower miR-455-3p levels in COPD (p<0.05). We identified 964 and 945 genes in the miRNA-targetomes of unstimulated and TGF-?1-stimulated lung fibroblasts, respectively. The TGF-? and Wnt pathways were significantly enriched among the Ago2-IP enriched and predicted targets of miR-455-3p and miR-21-3p. The miR-455-3p target genes HN1, NGF, STRADB, DLD and ANO3 and the miR-21-3p target genes HHEX, CHORDC1 and ZBTB49 were consistently more enriched after TGF-?1 stimulation.<h4>Conclusion</h4>Two miRNAs, miR-455-3p and miR-21-3p, were induced by TGF-?1 in lung fibroblasts. The significant Ago2-IP enrichment of targets of these miRNAs related to the TGF-? and/or Wnt pathways (NGF, DLD, HHEX) in TGF-?1-stimulated fibroblasts suggest a role for these miRNAs in lung diseases by affecting lung fibroblast function.
Project description:Breast cancer is the most common malignancy, and metastasis is the main cause of cancer-associated mortality in women worldwide. Transforming growth factor-? (TGF-?) signaling, an inducer of epithelial-to-mesenchymal transition (EMT), plays an important role in breast cancer metastasis. Abnormal expression of miR-543 is associated with tumorigenesis and progression of various human cancers; however, the knowledge about the role of miR-543 in breast cancer metastasis is still unknown. In this study, we demonstrated that miR-543 inhibits the EMT-like phenotype and TGF-?-induced breast cancer metastasis both in vitro and in vivo by targeting ZNF281. ZNF281 transactivates the EMT-related transcription factor ZEB1 and Snail. Furthermore, both ZEB1 and Snail can transcriptionally suppress miR-543 expression. Taken together, our data uncover the ZNF281-miR-543 feedback loop and provide a mechanism to extend the understanding of TGF-? network complexity.
Project description:Accumulating literatures have indicated that long non-coding RNAs (lncRNAs) are crucial molecules in tumor progression in various human cancers, including colorectal cancer (CRC). However, the clinical significance and regulatory mechanism of a vast majority of lncRNAs in CRC remain to be determined. The current study aimed to explore the function and molecular mechanism of lncRNA AC010789.1 in CRC progression. AC010789.1 found to be overexpressed in CRC tissues and cells. High expression of AC010789.1 was associated with lymph node metastasis and poor prognosis. Moreover, AC010789.1 silencing inhibited proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) in vitro as well as tumorigenesis and metastasis in vivo. Mechanistically, we demonstrated that repression of AC010789.1 promoted miR-432-3p expression, and miR-432-3p directly binds to ZEB1. We then proved the anti-tumor role of miR-432-3p in CRC, showing that the inhibitory effect of AC010789.1 knockdown on CRC cells was achieved by the upregulation of miR-432-3p but downregulation of ZEB1. We also established that silencing AC010789.1 suppressed the Wnt/?-catenin signaling pathway. However, this inhibitory effect was partially counteracted by inhibition of miR-432-3p. In summary, these results reveal that silencing AC010789.1 suppresses CRC progression via miR-432-3p-mediated ZEB1 downregulation and suppression of the Wnt/?-catenin signaling pathway, highlighting a potentially promising strategy for CRC treatment.