TGF-? induces miR-100 and miR-125b but blocks let-7a through LIN28B controlling PDAC progression.
ABSTRACT: TGF-?/Activin induces epithelial-to-mesenchymal transition and stemness in pancreatic ductal adenocarcinoma (PDAC). However, the microRNAs (miRNAs) regulated during this response have remained yet undetermined. Here, we show that TGF-? transcriptionally induces MIR100HG lncRNA, containing miR-100, miR-125b and let-7a in its intron, via SMAD2/3. Interestingly, we find that although the pro-tumourigenic miR-100 and miR-125b accordingly increase, the amount of anti-tumourigenic let-7a is unchanged, as TGF-? also induces LIN28B inhibiting its maturation. Notably, we demonstrate that inactivation of miR-125b or miR-100 affects the TGF-?-mediated response indicating that these miRNAs are important TGF-? effectors. We integrate AGO2-RIP-seq with RNA-seq to identify the global regulation exerted by these miRNAs in PDAC cells. Transcripts targeted by miR-125b and miR-100 significantly overlap and mainly inhibit p53 and cell-cell junctions' pathways. Together, we uncover that TGF-? induces an lncRNA, whose encoded miRNAs, miR-100, let-7a and miR-125b play opposing roles in controlling PDAC tumourigenesis.
Project description:In this study a microRNA (miRNA) signature was identified in a gemcitabine resistant pancreatic ductal adenocarcinoma (PDAC) cell line model (BxPC3-GZR) and this signature was further examined in advanced PDAC tumor specimens from The Cancer Genome Atlas (TCGA) database. BxPC3-GZR showed a mesenchymal phenotype, expressed high levels of CD44 and showed a highly significant deregulation of 17 miRNAs. Based on relevance to cancer, a seven-miRNA signature (miR-100, miR-125b, miR-155, miR-21, miR-205, miR-27b and miR-455-3p) was selected for further studies. A strong correlation was observed for six of the seven miRNAs in 43 advanced tumor specimens compared to normal pancreas tissue. To assess the functional relevance we initially focused on miRNA-125b, which is over-expressed in both the BxPC3-GZR model and advanced PDAC tumor specimens. Knockdown of miRNA-125b in BxPC3-GZR and Panc-1 cells caused a partial reversal of the mesenchymal phenotype and enhanced response to gemcitabine. Moreover, RNA-seq data from each of 40 advanced PDAC tumor specimens from the TCGA data base indicate a negative correlation between expression of miRNA-125b and five of six potential target genes (BAP1, BBC3, NEU1, BCL2, STARD13). Thus far, two of these target genes, BBC3 and NEU1, that are tumor suppressor genes but not yet studied in PDAC, appear to be functional targets of miR-125b since knockdown of miR125b caused their up regulation. These miRNAs and their molecular targets may serve as targets to enhance sensitivity to chemotherapy and reduce metastatic spread.
Project description:Breast cancer is still the most common and leading cause of cancer-related deaths in women worldwide. Long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) have shown key regulator roles in various cancers. Previous reports have identified miR-125b as a critical tumor suppressor in breast cancer. However, the role of lncRNAs in breast cancer is far from well-characterized. In this study, we identified a novel lncRNA LINC01787, which specifically binds pre-miR-125b, inhibits the binding between DICER and pre-miR-125b, represses the processing of pre-miR-125b by DICER, and therefore induces pre-miR-125b accumulation and represses mature miR-125b generation. Functional assays showed that LINC01787 promotes breast cancer cell proliferation and migration and breast cancer xenograft growth <i>in vivo</i>, which is abolished by the mutation of pre-miR-125b binding sites on LINC01787 or overexpression of miR-125b. Furthermore, LINC01787 is up-regulated in breast cancer tissues and is associated with advanced stages and poor survival. The expression of LINC01787 is inversely associated with that of miR-125b in breast cancer tissues. In conclusion, our findings identified a novel up-regulated and oncogenic lncRNA LINC01787 in breast cancer, which binds pre-miR-125b and represses mature miR-125b generation. Our data suggests LINC01787 as a potential prognostic biomarker and therapeutic target for breast cancer.
Project description:Although regulation of stem cell homeostasis by microRNAs (miRNAs) is well studied, it is unclear how individual miRNAs genomically encoded within an organized polycistron can interact to induce an integrated phenotype. miR-99a/100, let-7, and miR-125b paralogs are encoded in two tricistrons on human chromosomes 11 and 21. They are highly expressed in hematopoietic stem cells (HSCs) and acute megakaryoblastic leukemia (AMKL), an aggressive form of leukemia with poor prognosis. Here, we show that miR-99a/100?125b tricistrons are transcribed as a polycistronic message transactivated by the homeobox transcription factor HOXA10. Integrative analysis of global gene expression profiling, miRNA target prediction, and pathway architecture revealed that miR-99a/100, let-7, and miR-125b functionally converge at the combinatorial block of the transforming growth factor ? (TGF?) pathway by targeting four receptor subunits and two SMAD signaling transducers. In addition, down-regulation of tumor suppressor genes adenomatous polyposis coli (APC)/APC2 stabilizes active ?-catenin and enhances Wnt signaling. By switching the balance between Wnt and TGF? signaling, the concerted action of these tricistronic miRNAs promoted sustained expansion of murine and human HSCs in vitro or in vivo while favoring megakaryocytic differentiation. Hence, our study explains the high phylogenetic conservation of the miR-99a/100?125b tricistrons controlling stem cell homeostasis, the deregulation of which contributes to the development of AMKL.
Project description:OBJECTIVE:The study aimed to explore the correlations of long non-coding RNA MALAT1 (lncRNA MALAT1) and its targets microRNA (miR)-125b, miR-133, miR-146a, and miR-203 with acute exacerbation risk, inflammation, and disease severity of chronic obstructive pulmonary disease (COPD). METHODS:Plasma samples were obtained from 120 acute exacerbation COPD (AECOPD) patients, 120 stable COPD patients, and 120 healthy controls (HCs). RT-qPCR was conducted to detect lncRNA MALAT1 expression and its target miRNAs, and ELISA was performed to detect the inflammatory cytokines. RESULTS:LncRNA MALAT1 was highest in AECOPD patients followed by stable COPD patients and then HCs, which distinguished AECOPD patients from HCs (AUC: 0.969, 95% CI: 0.951-0.987) and stable COPD patients (AUC: 0.846, 95% CI: 0.798-0.894). Furthermore, lncRNA MALAT1 positively correlated with GOLD stage in both AECOPD and stable COPD patients. Regarding inflammatory cytokines, lncRNA MALAT1 positively correlated with tumor necrosis factor-? (TNF-?), interleukin (IL)-1?, IL-6, IL-8, IL-17, and IL-23 in both AECOPD and stable COPD patients. Besides, lncRNA MALAT1 negatively correlated with miR-125b, miR-146a, and miR-203 in AECOPD patients and reversely correlated with miR-125b and miR-146a in stable COPD patients. Notably, miR-125b, miR-133, miR-146a, and miR-203 were the lowest in AECOPD patients, followed by stable COPD patients, and then HCs; miR-125b, miR-133, miR-146a, and miR-203 negatively correlated with inflammation and GOLD stage in AECOPD and stable COPD patients. CONCLUSION:LncRNA MALAT1 exhibits clinical implications in acute exacerbation risk prediction and management of COPD via the inner-correlation with its targets miR-125b, miR-146a, and miR-203.
Project description:UNLABELLED:Extracellular vesicles (EV), including exosomes and shed vesicles, have been implicated in intercellular communication; however, their biomarker potential is less clear. Therefore, EVs derived from MCF7 and MCF10A cells were analyzed to identify unique miRNA (miR) profiles that distinguish their origin. One characteristic common to the miR profiles of MCF7 EVs and their parent cells is the high abundance of miR-21, let-7a, miR-100, and miR-125b, and low levels of miR-205. A second characteristic is the high abundance of "miRNA-like" tRNA fragments, which is unique to the MCF7 EVs, and is not found in comparing the cellular profiles. In addition, correlations were examined in the MCF7 cellular expression levels of these five miRs and two tRNA-derived miRNAs, miR-720 and miR-1274b, and compared with the correlations in MCF7 EV levels. Interestingly, correlations in the cellular expression of miR-125b, miR-100, and let-7a are mirrored in the EVs. In contrast, correlations in tRNA-derived miRNA levels are found only in the EVs. The findings suggest that EV miR clusters can be defined based on functional miR interactions related to correlated cellular expression levels or physical miR interactions, for example, aggregation due to comparable binding affinities to common targets. IMPLICATIONS:These results point to using high levels of tRNA-derived small RNA fragments in combination with known miR signatures of tumors to distinguish tumor-derived EVs in circulation from EVs derived from other cell sources. Such biomarkers would be unique to the EVs where high abundances of tRNA fragments are amplified with respect to their cellular levels.
Project description:MiR-100 and miR-125b are lost in many cancers and have potential function as tumor suppressors. Using both primary prostatic epithelial cultures and laser capture-microdissected prostate epithelium from 45 patients enrolled in a vitamin D3 randomized trial, we identified miR-100 and -125b as targets of 1,25-dihydroxyvitamin D3 (1,25D). In patients, miR-100 and -125b levels were significantly lower in tumor tissue than in benign prostate. Similarly, miR-100 and -125b were lower in primary prostate cancer cells than in cells derived from benign prostate. Prostatic concentrations of 1,25D positively correlated with these miRNA levels in both prostate cancer and benign epithelium, showing that patients with prostate cancer may still benefit from vitamin D3. In cell assays, upregulation of these miRNAs by 1,25D was vitamin D receptor dependent. Transfection of pre-miR-100 and pre-miR-125b in the presence or absence of 1,25D decreased invasiveness of cancer cell, RWPE-2. Pre-miR-100 and pre-miR-125b decreased proliferation in primary cells and cancer cells respectively. Pre-miR-125b transfection suppressed migration and clonal growth of prostate cancer cells, whereas knockdown of miR-125b in normal cells increased migration indicates a tumor suppressor function. 1,25D suppressed expression of previously bona fide mRNA targets of these miRNAs, E2F3 and Plk1, in a miRNA-dependent manner. Together, these findings show that vitamin D3 supplementation augments tumor suppressive miRNAs in patient prostate tissue, providing evidence that miRNAs could be key physiologic mediators of vitamin D3 activity in prevention and early treatment of prostate cancer.
Project description:Over the past few years, it has become evident that the distinctive pattern of miRNA expression seen in embryonic stem cells (ESCs) contributes to important signals in the choice of the cell fate. Thus, the identification of miRNAs and their targets, whose expression is linked to a specific step of differentiation, as well as the modulation of these miRNAs, may prove useful in the learning of how ESC potential is regulated. In this context, we have studied the expression profile of miRNAs during neural differentiation of ESCs. We have found that miR-125b is upregulated in the first steps of neural differentiation of ESCs. This miRNA targets the BMP4 co-receptor, Dies1, and, in turn, regulates the balance between BMP4 and Nodal/Activin signaling. The ectopic expression of miR-125b blocks ESC differentiation at the epiblast stage, and this arrest is rescued by restoring the expression of Dies1. Finally, opposite to miR-125a, whose expression is under the control of the BMP4, miR-125b is not directly regulated by Transforming Growth Factor beta (TGF?) signals. These results highlight a new important role of miR-125b in the regulation of the transition from ESCs to the epiblast stage and add a new level of control on TGF? signaling in ESCs.
Project description:Multiple Endocrine Neoplasia type 1 (MEN1) is an autosomal dominant disorder characterised by the combined occurrence of parathyroid, pituitary and pancreatic islet tumours, and is due to mutations of the MEN1 gene, which encodes the tumour suppressor protein menin. Menin has multiple roles in genome stability, transcription, cell division and proliferation, but its mechanistic roles in tumourigenesis remain to be fully elucidated. MicroRNAs (miRNA) are non-coding single stranded RNAs that post-transcriptionally regulate gene expression and have been associated with tumour development, although the contribution of miRNAs to MEN1-associated tumourigenesis and their relationship with menin expression are not fully understood. Alterations in miRNA expression, including downregulation of three putative 'tumour suppressor' miRNAs, miR-15a, miR-16-1 and let-7a, have been reported in several tumour types including non-MEN1 pituitary adenomas. We have therefore investigated the expression of miR-15a, miR-16-1 and let-7a in pituitary tumours that developed after 12 months of age in female mice with heterozygous knock out of the Men1 gene (Men1+/- mice). The miRNAs miR-15a, miR-16-1 and let-7a were significantly downregulated in pituitary tumours (by 2.3-fold, p<0.05; 2.1-fold p<0.01 and 1.6-fold p<0.05, respectively) of Men1+/- mice, compared to normal wild type pituitaries. MiR-15a and miR-16-1 expression inversely correlated with expression of cyclin D1, a known pro-tumourigenic target of these miRNAs, and knock down of menin in a human cancer cell line (HeLa), and AtT20 mouse pituitary cell line resulted in significantly decreased expression of miR-15a (p<0.05), indicating that the decrease in miR-15a may be a direct result of lost menin expression.
Project description:BACKGROUND:MicroRNAs (miRNAs) are endogenous, small (21-25 nucleotide), non-coding RNAs that play important roles in numerous biological processes. Koi carp exhibit diverse color patterns, making it an ideal subject for studying the genetics of pigmentation. However, the influence of miRNAs on skin color regulation and variation in Koi carp is poorly understood. RESULTS:Herein, we performed small RNA (sRNA) analysis of the three main skin colors in Koi carp by Illumina sequencing. The results revealed 330, 397, and 335 conserved miRNAs (belonging to 81 families) and 340, 353, and 351 candidate miRNAs in black, red, and white libraries, respectively. A total of 164 differentially expressed miRNAs (DEMs) and 14 overlapping DEMs were identified, including miR-196a, miR-125b, miR-202, miR-205-5p, miR-200b, and etc. Target prediction and functional analysis of color-related miRNAs such as miR-200b, miR-206, and miR-196a highlighted putative target genes, including Mitf, Mc1r, Foxd3, and Sox10 that are potentially related to pigmentation. Determination of reference miRNAs for relative quantification showed that let-7a was the most abundant single reference gene, and let-7a and miR-26b was the most abundant combination. CONCLUSIONS:The findings provide novel insight into the molecular mechanisms determining skin color differentiation in Koi carp, and serve as a valuable reference for future studies on tissue-specific miRNA abundance in Koi carp.
Project description:BACKGROUND: MicroRNAs (miRNAs) are involved in essential biological activities, and have been reported to exhibit differential expression profiles in various cancers. Our previous study demonstrated that intercellular adhesion molecule-2 (ICAM2) inhibition induces radiosensitisation in oral squamous cell carcinoma (OSCC) cells. Thus, we hypothesised that certain miRNAs play crucial roles in radioresistance in OSCC by regulating ICAM2 expression. METHODS: Because predicted target gene analyses revealed that microRNA-125b (miR-125b) potentially regulates ICAM2 mRNA expression, we examined the association between miR-125b and radioresistance. The expression of miR-125b was investigated by real-time quantitative reverse transcriptase-PCR. For a functional analysis, miR-125b was transfected to OSCC-derived cells. RESULTS: A downregulated expression of miR-125b was found in OSCC-derived cell lines and OSCC samples. The miR-125b-transfected cells showed a decreased proliferation rate, enhanced radiosensitivity to X-ray irradiation and diminished ICAM2 mRNA expression. Moreover, miR-125b expression correlated with OSCC tumour staging and survival. CONCLUSION: These findings suggested that the downregulated miR-125b expression was associated with proliferation and radioresistance mechanisms, probably through ICAM2 signalling. Thus, controlling the expression or activity of miR-125b might contribute to suppressing proliferation and overcoming radioresistance in OSCC.