MicroRNA-203a inhibits breast cancer progression through the PI3K/Akt and Wnt pathways.

MicroRNA expression in breast cancer (BC) is explored both as a potential biomarker and for therapeutic purposes. Recent studies have revealed that miR-203a-3p is involved in BC, and importantly contributes to BC chemotherapy responses; however, the regulatory pathways of miR-203a in BC remain elusive. Hence, we aimed to investigate the miR-203a regulatory mechanisms and their potential functions in the progress of BC. To this end, the miR-203a potential involving pathways was predicted by databases analyzing its target genes. The relations between miR-203a, the phosphatidylinositol 3'-kinase (PI3K)-Akt, and Wnt signaling pathways were mechanistically investigated. Our results revealed that miR-203a inhibited the activation of the PI3K/Akt and Wnt pathways and reduced its downstream cell cycle signals, including Cyclin D1 and c-Myc. Moreover, the overexpression of miR-203a drastically arrested the cell cycle at subG1 and G1 phases, decreased the viability, proliferation, and migration, and increased apoptosis of BC cells. Therefore, miR-203a-3p may be considered a tumor suppressor factor and a potential biomarker or therapeutic target for BC.

CLEC19A overexpression inhibits tumor cell proliferation/migration and promotes apoptosis concomitant suppression of PI3K/AKT/NF-κB signaling pathway in glioblastoma multiforme.

BACKGROUND: GBM is the most frequent malignant primary brain tumor in humans. The CLEC19A is a member of the C-type lectin family, which has a high expression in brain tissue. Herein, we sought to carry out an in-depth analysis to pinpoint the role of CLEC19A expression in GBM. METHODS: To determine the localization of CLEC19A, this protein was detected using Western blot, Immunocytochemistry/Immunofluorescence, and confocal microscopy imaging. CLEC19A expression in glioma cells and tissues was evaluated by qRT-PCR. Cell viability, proliferation, migration, and apoptosis were examined through MTT assay, CFSE assay, colony formation, wound healing assay, transwell test, and flow cytometry respectively after CLEC19A overexpression. The effect of CLEC19A overexpression on the PI3K/AKT/NF-κB signaling pathway was investigated using Western blot. An in vivo experiment substantiated the in vitro results using the glioblastoma rat models. RESULTS: Our in-silico analysis using TCGA data and measuring CLEC19A expression level by qRT-PCR determined significantly lower expression of CLEC19A in human glioma tissues compared to healthy brain tissues. By employment of ICC/IF, confocal microscopy imaging, and Western blot we could show that CLEC19A is plausibly a secreted protein. Results obtained from several in vitro readouts showed that CLEC19A overexpression in U87 and C6 glioma cell lines is associated with the inhibition of cell proliferation, viability, and migration. Further, qRT-PCR and Western blot analysis showed CLEC19A overexpression could reduce the expression levels of PI3K, VEGFα, MMP2, and NF-κB and increase PTEN, TIMP3, RECK, and PDCD4 expression levels in glioma cell lines. Furthermore, flow cytometry results revealed that CLEC19A overexpression was associated with significant cell cycle arrest and promotion of apoptosis in glioma cell lines. Interestingly, using a glioma rat model we could substantiate that CLEC19A overexpression suppresses glioma tumor growth. CONCLUSIONS: To our knowledge, this is the first report providing in-silico, molecular, cellular, and in vivo evidences on the role of CLEC19A as a putative tumor suppressor gene in GBM. These results enhance our understanding of the role of CLEC19A in glioma and warrant further exploration of CLEC19A as a potential therapeutic target for GBM.

In-silico and in-vitro evidence suggest LINC01405 as a sponge for miR-29b and miR-497-5p, and a potential regulator of Wnt, PI3K, and TGFB signaling pathways in breast carcinoma.

BACKGROUND: Carcinoma of the breast, a prevailing factor in female mortality worldwide, involves dysregulation of lncRNAs and microRNAs. AIM: The main goal of this research was to predict and experimentally examine the LINC01405 expression status in breast cancer subtypes, along with investigation of its interaction with miR-29b and miR-497-5p that results in regulating PI3-Kinase, WNT, and TGF-beta signaling pathways. METHODS AND RESULTS: We performed a meta-analysis of five GEO datasets, encompassing microarray and RNA-seq data, to identify differentially expressed genes. The Cancer Genome Atlas transcriptome dataset was also analyzed to determine essential gene modules, associated with different stages of breast cancer by weighted gene co-expression networks. In addition, networks of drug-gene interactions were constructed to explore potential treatment options. LINC01405 as a microRNA sponge was chosen and examined. furthermore, downstream target genes were discovered. Experimental validation consisted of plasmid constructs used in cell culture experiments, RT-qPCR for expression analysis, and cell cycle assays. Our bioinformatics findings showed higher LINC01405 expression in Basal-like triple-negative breast carcinoma. In contrast, lower expression in Luminal samples was observed compared with normal samples, which was consistently observed in both breast cancer tissues and cell lines. LINC01405 expression level was correlated with miR-29b and miR-497 levels. The MDA-MB-231 cell line demonstrated higher LINC01405 expression and lower miR-29b and miR-497 expression levels. However, SKBR3 and MCF7 cells had lower LINC01405 expression and higher miR-29b and miR-497 levels, suggesting a regulatory role for LINC01405 as a competing endogenous RNA. This was experimentally confirmed when LINC01405 was overexpressed in SKBR3 cells, and the common target genes of miR-29b and miR-497 were upregulated. Additionally, LINC01405 upregulation led to the increased cell populations, proliferation, and upregulation of critical cancer-related genes, including AKT1, AKT3, mTOR, WNT3A, SMAD3, CYCLIN D1, CYCLIN D2, BCL2, and GSK3B. CONCLUSION: We revealed the differential expression of LINC01405 in several types of breast cancer and its role in regulating signaling pathways, potentially via scavenging miRNAs. These findings clarified the role of LINC01405 in breast cancer development and identified potential therapeutic targets.

Microbial regulation of feedbacks to ecosystem change.

Microbes are key biodiversity components of all ecosystems and control vital ecosystem functions. Although we have just begun to unravel the scales and factors that regulate microbial communities, their role in mediating ecosystem stability in response to disturbances remains underexplored. Here, we review evidence of how, when, and where microbes regulate or drive disturbance feedbacks. Negative feedbacks dampen the impacts of disturbance, which maintain ecosystem stability, whereas positive feedbacks instead erode stability by amplifying the disturbance. Here we describe the processes underlying the responses to disturbance using a hierarchy of functional traits, and we exemplify how these may drive biogeochemical feedbacks. We suggest that the feedback potential of functional traits at different hierarchical levels is contingent on the complexity and heterogeneity of the environment.

Functional analysis of a putative HER2-associated expressed enhancer, Her2-Enhancer1, in breast cancer cells.

HER-2/neu (HER2) is a member of the epidermal growth factor receptors family, encoding a protein with tyrosine kinase activity. Following the gene amplification or increased HER2 transcription, carcinogenesis has been observed in some cancers. Genetic and epigenetic changes occurring in enhancer sequences can deeply affect the expression and transcriptional regulation of downstream genes, which can cause some physiological and pathological changes, including tumor progression. A therapeutic approach that directly targets the genomic sequence alterations is of high importance, with low side effects on healthy cells. Here, we employed the CRISPR/Cas9 method to genetically knockout an expressed putative enhancer (GH17J039694; we coined it as Her2-Enhancer1) located within the HER2 gene, 17q12: 39,694,339-39,697,219 (UCSC-hg38). We then investigated the potential regulatory effect of Her2-Enhancer1 on HER2 and HER2-interacting genes. To evaluate the cis and trans effects of Her2-Enhancer1, genetic manipulation of this region was performed in HER2-positive and -negative breast cancer cells. Our bioinformatics and real-time PCR data revealed that this putative enhancer region is indeed expressed, and acts as an expressed enhancer. Further functional analysis on edited and unedited cells revealed a significant alteration in the expression of HER2 variants, as well as some other target genes of HER2. Moreover, the apoptosis rate was considerably elevated within the edited cells. As we expected, Western blot analysis confirmed a reduction in protein levels of HER2, GRB7, the gene interacting with HER2, and P-AKT in the PI3K/AKT pathway. Altogether, our findings revealed an enhancer regulatory role for Her2-Enhancer1 on HER2 and HER2-interacting genes; and that this region has a potential for targeted therapy of HER2-positive cancers.

Downregulated miR-495-3p in colorectal cancer targets TGFßR1, TGFßR2, SMAD4 and BUB1 genes and induces cell cycle arrest.

BACKGROUND: Hsa-miR-495 (miR-495) has been extensively investigated in cancer initiation and progression. On the other hand, our bioinformatics analysis suggested that miR-495 exerts its effects through targeting of TGFß signaling components. METHODS & RESULTS: In order to investigate such an effect, miR-495 precursor was overexpressed in HEK293T, SW480, and HCT116 cells, which was followed by downregulation of TGFßR1, TGFßR2, SMAD4, and BUB1 putative target genes, detected by RT-qPCR. Also, luciferase assay supported the direct interaction of miR-495 with 3'UTR sequences of TGFßR1, TGFßR2, SMAD4, and BUB1 genes. Furthermore, a negative correlation of expression between miR-495-3p and some of these target genes was deduced in a set of colorectal and breast cancer cell lines. Then, flow cytometry analysis showed that the overexpression of miR-495 in HCT116 and HEK293T resulted in an arrest at the G1 phase. Consistently, western blotting analysis showed a significant reduction of the Cyclin D1 protein in the cells overexpressing miR-495, pointing to downregulation of the TGFß signaling pathway and cell cycle arrest. Finally, microarray data analysis showed that miR-495-3p is significantly downregulated in colorectal tumors, compared to the normal pairs. CONCLUSIONS: Overall, the results of the current study introduced miR-495-3p as a cell cycle progression suppressor, which may negatively regulate TGFßR1, TGFßR2, SMAD4, and BUB1 genes. This finding suggests miR-495-3p as a tumor suppressor candidate for further evaluation.

Exogenous melatonin induces phenolic compounds production in Linum album cells by altering nitric oxide and salicylic acid.

Melatonin is a pleiotropic molecule that can influence various aspects of plant performance. Recent studies have exhibited that it mediates plant defensive responses, probably through managing redox homeostasis. We tried to track the regulatory effects of melatonin on the antioxidant machinery of Linum album cell culture. To this, different concentrations of melatonin were applied, and the oxidative status of cells was investigated by measuring the levels of oxidative molecules and antioxidant agents. The results showed that H2O2 content did not change at the low melatonin levels, while it increased at the high concentrations. It can be correlated with the low melatonin dosages capacity to remove excessive amounts of H2O2, while the high melatonin dosages exhibit toxicity effects. In contrast, the NO enhancement occurred at 50 µM melatonin, proposing its role in triggering melatonin-induced defensive responses. The MDA results stated that NO led to oxidative stress in melatonin-treated cells at 50 µM melatonin. Antioxidant enzyme POD was activated by melatonin treatment, while SOD enzyme behaved reversely which can explain the changes in the H2O2 level. In addition, the analysis of the phenolics profile showed that the contents of phenolic acids, flavonoids, and lignans enhanced following an increase in PAL enzyme activity. The increased level of phenolic hormone SA can indicate that melatonin affects the defensive responses in L. album cells through a SA-dependent pathway. In general, it seems that melatonin, by modulating NO and SA levels, can induce the activity of antioxidant enzymes and the production of phenolics, especially lignans, in L. album cells.

LINC02381 suppresses cell proliferation and promotes apoptosis via attenuating IGF1R/PI3K/AKT signaling pathway in breast cancer.

Identification of the genes and genetic networks involved in breast cancer development is a major need for prevention and therapy. LINC02381 (lncRNA) has already been introduced as a tumor suppressor in colorectal and gastric cancers. Here, we intended to investigate its potential functional effects on breast cancer. In the analysis performed on RNA-Seq and microarray data, the LINC02381 lncRNA was found to be significantly downregulated in the breast tumors and associated with poor survival of the patients. Then, the differential expression of LINC02381 was confirmed in breast tumor tissues and cancer cell lines using RT-qPCR. Overexpression of LINC02381 resulted in reduced IGF1R and p-AKT expression levels which indicates decreased PI3K pathway activity, detected by RT-qPCR and western blotting. At the cellular level, LINC02381 overexpression was followed by a decreased proliferation rate of transfected breast cell lines, detected by PI flow cytometry, RT-qPCR, colony formation, and MTT assays. Consistently, the results of Annexin-V/PI flow cytometry, RT-qPCR, caspase3/7 activity, and AO/EB-H33342/PI dual staining revealed that LINC02381 overexpression induced apoptosis and cell death. The reduced migration rate of these cells was also verified through wound healing assay and RT-qPCR against the EMT-involved genes. Our data show that LINC02381 exerts its tumor suppressor effect at least partly through attenuation of the IGF1R/PI3K/AKT signaling pathway, which originated from IGF1R downregulation.

EVADR ceRNA transcript variants upregulate WNT and PI3K signaling pathways in SW480 and HCT116 cells by sponging miR-7 and miR-29b.

Long noncoding RNAs are cancer regulators and EVADR-lncRNA is highly upregulated in colorectal cancer (CRC). Accordingly, we aimed to functionally characterize the EVADR in CRC-originated cells. Firstly, during the amplification of EVADR full-length cDNA (named EVADR-v1), a novel/shorter variant (EVADR-v2) was discovered. Then, RT-qPCR analysis confirmed that EVADR is upregulated in tumors, consistent with RNA-seq analysis. Interestingly, bioinformatics analysis and dual-luciferase assay verified that EVADR sponges miR-7 and miR-29b. When both EVADR-v1/-v2 variants were overexpressed in SW480/HCT116 cells, miR-7 and miR-29b target genes (involved in the WNT/PI3K signaling) were upregulated. Furthermore, EVADR-v1/-v2 overexpression resulted in elevated PI3K activity (verified by western blotting and RT-qPCR) and upregulation of WNT signaling (confirmed by western blotting, TopFlash assay, and RT-qPCR). Consistently, overexpression of EVADR-v1/-v2 variants was followed by increased cell cycle progression, viability and migration as well as reduced early/late apoptotic rate, and Bax/Bcl2 ratio of the CRC cells, detected by the cell cycle analysis, MTT, wound-healing, Annexin-V/PI, and RT-qPCR methods, respectively. Overall, we introduced two oncogenic transcript variants for EVADR that by sponging miR-7/miR-29b, upregulate WNT and PI3K signaling. Given the crucial role of these pathways in CRC, EVADR may present potential therapy use.

ADAMTS9-AS1 Long Non­coding RNA Sponges miR­128 and miR-150 to Regulate Ras/MAPK Signaling Pathway in Glioma.

Glioma is a malignancy of the central nervous system with a poor prognosis. Therefore, the elaboration of its molecular features creates therapeutic opportunities. Looking for the regulatory non-coding RNAs (lncRNAs and miRNAs) that are involved in glioma incidence/progression, RNA-seq analysis introduced upregulated ADAMTS9-AS1 as a bona fide candidate that sponges miR-128 and miR-150 and shows the negative correlation of expression with them. Then, RT-qPCR verified the upregulation of ADAMTS9-AS1 in glioma tissues and cell lines. Furthermore, dual-luciferase assay supported that cytoplasmic ADAMTS9-AS1 is capable of sponging miR-128 and miR-150, which are known as regulators of Ras/MAPK, PI3K, and Wnt pathways. Following the overexpression of ADAMTS9-AS1 in 1321N1 and U87 glioma cells, tyrosine kinase receptors (IGF1R and TrkC), as well as Wnt receptors (Lrp6 and Fzd) were upregulated, detected by RT-qPCR. Furthermore, downstream genes of both Ras/MAPK and Wnt pathways were upregulated. Finally following the ADAMTS9-AS1 overexpression, upregulation of Ras/MAPK and Wnt signaling pathways was verified through western blotting and Top/Fop flash assay, respectively. At the cellular level, ADAMTS9-AS1 overexpression brought about reduced sub-G1 cell population, increased proliferation rate, reduced apoptosis level, increased migration rate, shortened Bax/Bcl2 ratio, induced EMT, and stemness characteristics of transfected cells, detected by flow cytometry, MTT assay, scratch test, and RT-qPCR. Overall, these results introduced ADAMTS9-AS1 as an oncogene that upregulates Ras/MAPK and Wnt pathways through sponging of the miR-128 and miR-150 in glioma cells. The outcome of ADAMTS9-AS1 expression is more aggression of the glioma cells through increased EMT and stemness characteristics. These features candidate ADAMTS9-AS1 locus for glioma therapy. As a result, we discovered the oncogenic properties of ADAMTS9-AS1 in glioma cancer. It sponges miR-128 and miR-150 and subsequently overstimulates RAS/MAPK and Wnt signaling pathways, particularly at the receptors level. Thus, ADAMTS9-AS1 increases proliferation, migration, and stemness in glioma cell lines. A schematic representation showing the functional effect of ADAMTS9-AS1.

ErbB4-encoded novel miRNAs act as tumor suppressors by regulating ErbB/PI3K signaling.

BACKGROUND: ErbB/PI3K signaling is widely recognized as a critical modulator of malignancy and miRNAs have been found to play a crucial role in the regulation of this pathway. OBJECTIVE: This study aimed to identify novel miRNAs related to the ErbBs loci and investigate the functional effects of these miRNAs on ErbB/PI3K signaling in cancer progression. MATERIALS AND METHODS: Bioinformatics tools and RNA-seq data were used to discover novel miRNAs in breast and colon cancer cells. Gene expression levels were determined using RT-qPCR. Western blotting and dual-luciferase assays were used to identify the regulatory mechanism between ErbB4-miR1/2 and related genes. The effects of ErbB4-miR1/2 on cell proliferation, viability, ROS production, and migration were assessed by PI-flow cytometry, colony formation, MTT, ROS, scratch, and transwell assays in SKBR3 and SW480 cells. RESULTS: MicroRNA prediction tools, RNA-seq data, RT-qPCR, and sequencing results identified ErbB4-miR1 and ErbB4-miR2 (ErbB4-miR1/2) as novel miRNAs encoded by ErbB4 gene. ErbB4-miR1/2 were downregulated in breast and colon tumor tissues and also in different cancerous cells. RT-qPCR and dual-luciferase assays revealed that ErbB2 and ErbB3 genes are regulated by ErbB4-miR1/2. Consistently, a decrease in the p-AKT/AKT protein ratio verified the suppressive effect of ErbB4-miR1/2 on ErbB/PI3K activity. Furthermore, ErbB4-miR1/2 overexpression suppressed cell proliferation, viability, and migration, and increased ROS production. CONCLUSIONS: ErbB4-miR1/2 are novel tumor suppressor miRNAs which attenuate ErbB/PI3K signaling in breast and colon cancer cells.

LINC02381-ceRNA exerts its oncogenic effect through regulation of IGF1R signaling pathway in glioma.

PURPOSE: LncRNAs play essential roles in the cellular and molecular biology of glioma. Some LncRNAs exert their role through sponging miRNAs and regulating multiple signaling pathways. LINC02381 is involved in several cancer types as either oncogene or tumor suppressor. Here, we intended to find the molecular mechanisms of the LINC02381 effect during the glioma progression in related cell lines. METHODS AND RESULTS: RNA-seq data analysis indicated the oncogenic characteristics of LINC02381, and RT-qPCR results confirmed its upregulation compared to normal tissues. Besides its expression was relatively stronger in invasive glioma cell lines. Furthermore, in silico analysis revealed LINC02381 is concentrated in the cytoplasm and predicted its sponging effect against miR-128 and miR-150, which was verified through dual luciferase assay. When LINC02381 was overexpressed in 1321N1, U87, and A172 cell lines, IGF1R and TrkC receptors as well as their downstream pathways (PI3K and RAS/MAPK), were upregulated, detected by RT-qPCR, and verified by western analysis. Consistently, LINC02381 overexpression was followed by an increased proliferation rate of transfected glioma cell lines, detected by flow cytometry and MTT assay, and RT-qPCR. It also resulted in elevated EMT and stemness markers expression level, increased migration rate, and reduced apoptosis rate, detected by RT-qPCR, western analysis, scratch test, and Annexin/PI flow cytometry analysis, respectively. CONCLUSION: The overall results indicated that LINC02381 exerts its oncogenic effect in glioma cells through sponging miR-128 and miR-150 to upregulate the IGF1R signaling pathway. Our results introduce LINC02381 and miR-128, and miR-150 as potential prognosis and therapy targets for the treatment of glioma.

OCC-1D regulates Wnt signaling pathway: potential role of long noncoding RNA in colorectal cancer.

BACKGROUND: Aberrant activation of the Wnt signaling pathway is observed in most colorectal cancers (CRC). OCC-1D is a splice variant of OCC-1 gene which is considered as a long noncoding RNA (lncRNA) due to lacking the translational initiation codon of the gene. Here, we sought supporting evidence for the effects of OCC-1D on the Wnt pathway and cell cycle progression in CRC. METHODS AND RESULTS: TOP/FOPflash assay and qRT-PCR indicated that expression alterations of OCC-1D could change Wnt signaling activity in colon cancer cells. Consistently, immunocytochemistry results showed the effect of OCC-1D overexpression on nuclear localization of ß-catenin proteins in SW480 cells. Flow cytometry, wound healing and MTT assay confirmed the cell cycle stimulatory effects of OCC-1D in CRC-originated cell lines (SW480 and HCT116). qRT-PCR revealed a positive correlation between the expression level of OCC-1D and its neighboring gene, APPL2. Two distinct tests, downregulation of APPL2 mRNA by using shRNA and Wnt signaling inhibition by using small molecule, along with OCC-1D overexpression confirmed that OCC-1D lncRNA exerts its effect on Wnt signaling pathway through expression modulation of APPL2 gene. CONCLUSIONS: Collectively, we suggested the putative regulatory effects of OCC-1D lncRNA on cell cycle progression and Wnt signaling activation through enhancing the APPL2 gene transcription.

Molecular and cellular evidence for hsa-miR-1254 suppressor effect against HER2 signaling in breast cancer.

HER2 signaling upregulation is a hallmark of breast cancer which is the second cause of cancer-related death in women. Here, we were looking for the candidate microRNAs which is capable of regulating the HER2 receptor and the genes of its downstream. To this aim, preliminary bioinformatics analysis introduced hsa-miR-1254 (miR-1254) as a potential common regulator of HER2, HER3, PIK3R2, and AKT1 genes. Then, reverse-transcription quantitative polymerase chain reaction (RT-qPCR) analysis indicated a lower expression level of miR-1254 in breast cancer specimens, compared to their normal pairs. Furthermore, overexpression of miR-1254 resulted in HER2, HER3, PIK3R2, and AKT1 genes downregulation, detected by RT-qPCR and confirmed by western blot analysis and enzyme-linked immunosorbent assay test against AKT1, BAX, FADD, and HER2 protein levels in SKBR3 cells. Dual-luciferase assay also supported direct interaction of miR-1254 with MREs within 3' untranslated region sequences of HER2, HER3, PIK3R2, and AKT1 target genes. Overexpression of miR-1254 in SKBR3 cells was followed by increased BAX/BCL2 expression ratio, detected by RT-qPCR, and increased proportion of G1 cell population, detected by flow cytometry. Corroborated by cell cycle analysis, MTT, Annexin V-FITC, and Live-Dead cell staining assays, overexpression of miR-1254 in MDA-MB-231 cells showed opposing results following the overexpression of miR-1254. Taken together, results indicated that miR-1254 acts as cell-type-specific tumor suppressor that targets HER2, HER3, PIK3R2, and AKT1 transcripts. These results suggest miR-1254 as a potential therapeutic candidate for breast cancer subtypes.

The conserved long non-coding RNA CARMA regulates cardiomyocyte differentiation.

AIMS: Production of functional cardiomyocytes from pluripotent stem cells requires tight control of the differentiation process. Long non-coding RNAs (lncRNAs) exert critical regulatory functions in cell specification during development. In this study, we designed an integrated approach to identify lncRNAs implicated in cardiogenesis in differentiating human embryonic stem cells (ESCs). METHODS AND RESULTS: We identified CARMA (CARdiomyocyte Maturation-Associated lncRNA), a conserved lncRNA controlling cardiomyocyte differentiation and maturation in human ESCs. CARMA is located adjacent to MIR-1-1HG, the host gene for two cardiogenic miRNAs: MIR1-1 and MIR-133a2, and transcribed in an antisense orientation. The expression of CARMA and the miRNAs are negatively correlated, and CARMA knockdown increases MIR1-1 and MIR-133a2 expression. In addition, CARMA possesses MIR-133a2 binding sites, suggesting the lncRNA could be also a target of miRNA action. Upon CARMA down-regulation, MIR-133a2 target protein-coding genes are coordinately down-regulated. Among those, we found RBPJ, the gene encoding the effector of the NOTCH pathway. NOTCH has been shown to control a binary cell fate decision between the mesoderm and the neuroectoderm lineages, and NOTCH inhibition leads to enhanced cardiomyocyte differentiation at the expense of neuroectodermal derivatives. Interestingly, two lncRNAs, linc1230 and linc1335, which are known repressors of neuroectodermal specification, were found up-regulated upon Notch1 silencing in ESCs. Forced expression of either linc1230 or linc1335 improved ESC-derived cardiomyocyte production. These two lncRNAs were also found up-regulated following CARMA knockdown in ESCs. CONCLUSIONS: Altogether, these data suggest the existence of a network, implicating three newly identified lncRNAs, the two myomirs MIR1-1 and MIR-133a2 and the NOTCH signalling pathway, for the coordinated regulation of cardiogenic differentiation in ESCs.

MiRNA-Wnt signaling regulatory network in colorectal cancer.

Colorectal cancer (CRC) is one of the common malignancies worldwide and the Wnt signaling pathway is recognized as the main disrupted pathway in this malignancy. MicroRNAs (miRNAs) are recognized to contribute to the pathogenesis of CRC by triggering or impeding the Wnt signaling pathway. In addition, transcriptional regulation of miRNAs by canonical Wnt signaling also participates in CRC cell progression. In this review, we present comprehensive literature of the existing data on the interaction of miRNAs and Wnt signaling that could be useful in future studies in the field of CRC management.

Hsa-miR-186-5p regulates TGFß signaling pathway through expression suppression of SMAD6 and SMAD7 genes in colorectal cancer.

TGFß signaling is a known pathway to be involved in colorectal cancer (CRC) progression and miRNAs play crucial roles by regulating different components of this pathway. Hence, finding the link between miRNAs and the pathway could be beneficial for CRC therapy. Array data indicated that miR-186-5p is a differentially expressed miRNA in colorectal Tumor/Normal tissues and bioinformatics tools predicted SMAD6/7 (inhibitory SMADs) as bona fide targets of this miRNA. Here, we intended to investigate the regulatory effect of the miR-186-5p expression on TGFß signaling in CRC. Firstly, the miR-186-5p overexpression in HCT116 cells resulted in a significant reduction of SMAD6/7 expression, measured through RT-qPCR. Then, the direct interactions of miR-186-5p with SMAD6/7 3'UTRs were supported through dual luciferase assay. Furthermore, miR-186-5p overexpression suppressed proliferation, cell viability, and migration while, it increased apoptosis in CRC cells, assessed by cell cycle, MTT, scratch and Annexin V/PI apoptosis assays. Consistently, miR-186-5p overexpression resulted in reduced CyclinD1 protein using western blot, and also resulted in increased P21 and decreased c-Myc expression. Overall, these results introduced miR-186-5p as a cell cycle suppressor through downregulation of SMAD6/7 expression. Thus, miR-186-5p might be served as a novel tumor suppressive biomarker and therapeutic target in CRC treatment.

LncRNAs in Cardiomyocyte Maturation: New Window for Cardiac Regenerative Medicine.

Cardiomyocyte (CM) maturation, which is characterized by structural, functional, and metabolic specializations, is the last phase of CM development that prepares the cells for efficient and forceful contraction throughout life. Over the past decades, CM maturation has gained increased attention due to the fact that pluripotent stem cell-derived CMs are structurally, transcriptionally, and functionally immature and embryonic-like, which causes a defect in cell replacement therapy. The current challenge is to discover and understand the molecular mechanisms, which control the CM maturation process. Currently, emerging shreds of evidence emphasize the role of long noncoding RNAs (lncRNAs) in regulating different aspects of CM maturation, including myofibril maturation, electrophysiology, and Ca2+ handling maturation, metabolic maturation and proliferation to hypertrophy transition. Here, we describe the structural and functional characteristics of mature CMs. Furthermore, this review highlights the lncRNAs as crucial regulators of different aspects in CM maturation, which have the potential to be used for mature CM production. With the current advances in oligonucleotide delivery; lncRNAs may serve as putative therapeutic targets to produce highly mature CMs for research and regenerative medicine.

Implication of TrkC-miR2 in neurotrophin signalling pathway regulation through NGFR transcript targeting.

TrkC and NGFR neurotrophin receptors are associated with cell death, cancer and differentiation. TrkC-miR2, which is located in TrkC gene, is known to regulate Wnt signalling pathway, and its influence on other signalling pathways is under investigation. Here, through RT-qPCR, dual-luciferase assay and Western blotting we reveal that TrkC-miR2 targets NGFR. Overexpression of TrkC-miR2 also affected TrkA, TrkC, NFKB, BCL2 and Akt2 expressions involved in neurotrophin signalling pathway, and elevated survival rate of HEK293t and U87 cells was distinguished by flow cytometry and MTT assay. Consistently, an opposite expression correlation was obtained between TrkC-miR2 and NGFR or TrkC for the duration of NT2 differentiation. Meanwhile, TrkC-miR2 down-regulation attenuated NT2 differentiation into neural-like cells. Overall, here we present in silico and experimental evidence showing TrkC-miR2 as a new controller in regulation of neurotrophin signalling pathway.