[ZEB2 Regulates the Migration and Invasion of PANC-1 Pancreatic Cancer Cells: An Experimental Study].

Objective: To investigate the effects and mechanisms of zinc finger E-box binding homeobox transcription factor-2 ( ZEB2) on the proliferation, colony formation, migration, and invasion abilities and the epithelial-mesenchymal transition (EMT) of PANC-1 cells, a human pancreatic cancer cell line. Methods: Data on the expression of ZEB2 in pancreatic cancer tissues and paracancerous tissues from The Cancer Genome Atlas (TCGA) database were analyzed. PANC-1 pancreatic cancer cells were divided into si-NC group, si- ZEB2 group, pcDNA3.1 group, and pcDNA3.1- ZEB2 group. qRT-PCR and Western blot were conducted to confirm the effectiveness of ZEB2 knockdown or overexpression. CCK-8, colony formation, wound healing, and Transwell assays were conducted to examine the effects of ZEB2 on the proliferation, colony formation, migration, and invasion of PANC-1 cells. qRT-PCR and immunofluorescence assays were performed to examine the expression of E-cadherin and vimentin, the EMT markers, in the cells. Prediction of proteins interacting with ZEB2 was made through the STRING database. Results: TCGA database analysis showed that the expression level of ZEB2 in pancreatic cancer tissues was significantly higher than that in adjacent tissues ( P<0.05). Compared with those of cells in the control group, the proliferation, colony formation, migration, and invasion of cells in the si- ZEB2 group were decreased ( P<0.05). Compared with those of cells in the pcDNA3.1 group, the proliferation, colony formation, migration and invasion of cells in the pcDNA3.1- ZEB2 group were increased (all P<0.05). According to the results of qRT-PCR and immunofluorescence assays, compared with those of the si-NC group, the expression of E-cadherin mRNA, an epithelial marker, in the si- ZEB2 group increased, while the expression of vimentin mRNA, an mesenchymal marker, and the protein decreased. Compared with those of the pcDNA3.1 group, the expression of E-cadherin mRNA in the PANC-1 cells of the pcDNA3.1- ZEB2 group decreased, while the expression of vimentin mRNA and the protein increased (all P<0.05). Analysis with the STRING database predicted that 10 proteins had close interaction with ZEB2. Conclusion: Overexpression of ZEB2 promotes the migration, invasion, and the EMT process of PANC-1 pancreatic cancer cells.

RNA N6-methyladenosine modification mediates downregulation of NR4A1 to facilitate malignancy of cervical cancer.

BACKGROUND: N6-methyladenosine is the most abundant eukaryotic mRNA modification and alters a wide range of cellular processes in cancer. Therefore, defining the molecular details are critical for understanding the regulatory mechanism of m6A modification. RESULTS: We found that METTL3, a core m6A methyltransferase component, is upregulated and functions as an oncogene in cervical cancer. Mechanistically, METTL3 induces the degradation of m6A-modified transcripts of NR4A1 though YTHDF2-DDX6 pathway. In addition, NR4A1 overexpression attenuates the malignant progression through recruiting the LSD1/HDAC1/CoREST transcriptional repression complex to AKT1 promoter. CONCLUSIONS: Our findings reveal that m6A regulates cervical cancer cellular progression through manipulating NR4A1 pathway.

The cAMP-PKA signalling pathway regulates hyphal growth, conidiation, trap morphogenesis, stress tolerance, and autophagy in Arthrobotrys oligospora.

The cyclic adenosine monophosphate-protein kinase A (cAMP-PKA) signalling pathway is evolutionarily conserved in eukaryotes and plays a crucial role in defending against external environmental challenges, which can modulate the cellular response to external stimuli. Arthrobotrys oligospora is a typical nematode-trapping fungus that specializes in adhesive networks to kill nematodes. To elucidate the biological roles of the cAMP-PKA signalling pathway, we characterized the orthologous adenylate cyclase AoAcy, a regulatory subunit (AoPkaR), and two catalytic subunits (AoPkaC1 and AoPkaC2) of PKA in A. oligospora by gene disruption, transcriptome, and metabolome analyses. Deletion of Aoacy significantly reduced the levels of cAMP and arthrobotrisins. Results revealed that Aoacy, AopkaR, and AopkaC1 were involved in hyphal growth, trap morphogenesis, sporulation, stress resistance, and autophagy. In addition, Aoacy and AopkaC1 were involved in the regulation of mitochondrial morphology, thereby affecting energy metabolism, whereas AopkaC2 affected sporulation, nuclei, and autophagy. Multi-omics results showed that the cAMP-PKA signalling pathway regulated multiple metabolic and cellular processes. Collectively, these data highlight the indispensable role of cAMP-PKA signalling pathway in the growth, development, and pathogenicity of A. oligospora, and provide insights into the regulatory mechanisms of signalling pathways in sporulation, trap formation, and lifestyle transition.

Mechanism of the improved Fe(III)/persulfate reaction by gallic acid for ibuprofen degradation.

Gallic acid (GA), a natural plant polyphenol, was applied as amendment of Fe(III)/persulfate (PS) system for ibuprofen (IBP) degradation in this study. The impacts of all agentia (GA, Fe(III), PS) concentration and initial pH values on IBP removal efficiency were investigated, and their corresponding observed pseudo-first-order rate constants (kobs) were calculated. The addition of GA has significantly improved elimination efficiency of IBP due to the enhanced Fe(III)/Fe(II) cycle. Electron paramagnetic resonance (EPR) results confirmed that SO4•-, HO• and O2•- were involved in GA/Fe(III)/PS system. However, quenching experiments further affirmed the impact of SO4•- and HO• towards IBP decomposition instead of O2•-, with contribution ratio to IBP removal was 69.12% and 30.88%, respectively. SO4•- was the main radicals formed by directly activation of PS with Fe(II), while HO• was the transformation product of SO4•-. Based on instrumental analysis (stopped-flow UV-vis spectrum and MS) and theoretical calculation, the potential reaction mechanism between GA and Fe(III) in the presence of PS was further proposed. GA complexed with Fe(III) firstly and the Fe(III)-GA complex was then converted into quinone substance, accompanied by the generation of Fe(II). Furthermore, the application of GA extended the optimal pH range to neutral as well, which made it a promising treatment in practical application.

Predicting colorectal cancer tumor mutational burden from histopathological images and clinical information using multi-modal deep learning.

MOTIVATION: Tumor mutational burden (TMB) is an indicator of the efficacy and prognosis of immune checkpoint therapy in colorectal cancer (CRC). In general, patients with higher TMB values are more likely to benefit from immunotherapy. Though whole-exome sequencing is considered the gold standard for determining TMB, it is difficult to be applied in clinical practice due to its high cost. There are also a few DNA panel-based methods to estimate TMB; however, their detection cost is also high, and the associated wet-lab experiments usually take days, which emphasize the need for faster and cheaper alternatives. RESULTS: In this study, we propose a multi-modal deep learning model based on a residual network (ResNet) and multi-modal compact bilinear pooling to predict TMB status (i.e. TMB high (TMB_H) or TMB low(TMB_L)) directly from histopathological images and clinical data. We applied the model to CRC data from The Cancer Genome Atlas and compared it with four other popular methods, namely, ResNet18, ResNet50, VGG19 and AlexNet. We tested different TMB thresholds, namely, percentiles of 10%, 14.3%, 15%, 16.3%, 20%, 30% and 50%, to differentiate TMB_H and TMB_L.For the percentile of 14.3% (i.e. TMB value 20) and ResNet18, our model achieved an area under the receiver operating characteristic curve of 0.817 after 5-fold cross-validation, which was better than that of other compared models. In addition, we also found that TMB values were significantly associated with the tumor stage and N and M stages. Our study shows that deep learning models can predict TMB status from histopathological images and clinical information only, which is worth clinical application.

Identification of prognostic alternative splicing signature in gastric cancer.

BACKGROUND: Aberrant alternative splicing (AS) events could be viewed as prognostic indicators in a large number of malignancies. This study aims to identify prognostic AS events, illuminate the function of the splicing variants biomarkers and provide reliable evidence for formulating public health strategies for gastric cancer (GC) surveillance. METHODS: RNA-Seq data, clinical information and percent spliced in (PSI) values were available in The cancer genome atlas (TCGA) and TCGA SpliceSeq data portal. A three-step regression method was conducted to identify prognostic AS events and construct multi-AS-based signatures. The associations between prognostic AS events and splicing factors were also investigated. RESULTS: We identified a total of 1,318 survival-related AS events in GC, parent genes of which were implicated in numerous oncogenic pathways. The final prognostic signatures stratified by seven types of AS events or not stratified performed well in risk prediction for GC patients. Moreover, five signatures based on AA, AD, AT, ES and RI events function as independent prognostic indicators after multivariate adjustment of other clinical variables. Splicing network also showed marked correlation between the expression of splicing factors and PSI value of AS events in GC patients. CONCLUSION: Our findings provide a landscape of AS events and regulatory network in GC, indicating that AS events might serve as prognostic biomarkers and therapeutic targets for GC.

The positive feedback loop of NHE1-ERK phosphorylation mediated by BRAFV600E mutation contributes to tumorigenesis and development of glioblastoma.

The v-raf murine sarcoma viral oncogene homolog B1 (BRAF) activating mutation V600E (BRAFV600E) is involved in glioblastoma multiforme (GBM). Na/H exchanger 1 (NHE1), a main pH regulator affecting cell microenvironment, is hyper-expressed in GBM. However, the relationship between BRAFV600E signal pathway and NHE1 in GMB cells remains unclear. This study found that NHE1 was a downstream target of BRAFV600E and an upstream factor of extracellular signal-regulated kinase (ERK). In addition, there was a positive feedback loop between NHE1-ERK phosphorylation under regulation of BRAFV600E mutation contributing to the proliferation and invasion of GBM cells. Moreover, the proliferation and invasion abilities of BRAFV600E-mutant and BRAF wild type GBM cells were all suppressed by the NHE1 inhibitor, BRAFV600E inhibitor and combination of them. The inhibitory effect of combination of the two inhibitors was better than each single drug both in vitro and in vivo. Combination of BRAFV600E and NHE1 inhibitors could be considered as a new therapeutic regimen for GBM, especially for GBM with BRAFV600E.

microRNA-10a-5p overexpression suppresses malignancy of colon cancer by regulating human liver cancer fibroblasts.

The crosstalk between tumor and stroma plays a critical role in cancer metastasis. However, the function of miR-10a-5p on liver fibroblasts in the metastatic microenvironment of colon cancer (CC) and the effect of activated fibroblasts on CC cells are still unclear. In our study, miR-10a-5p overexpression inhibited the proliferation, migration, and IL-6/IL-8 level of LX-2 cells and human liver cancer fibroblasts (HLCFs). Moreover, miR-10a-5p had lower expression in HLCFs than in human liver normal fibroblasts (HLNFs). The conditioned medium (CM) from LX-2 cells with miR-10a-5p overexpression or HLNFs could inhibit the invasion, migration, and stemness of CC SW480 cells, whereas HLCFs CM could promote these malignant phenotypes of SW480 cells. The present study illustrates the effect of miR-10a-5p on the liver fibroblasts and the altered liver fibroblasts in the microenvironment on CC cells induced by miR-10a-5p, which may aid the understanding of the mechanisms underlying CC liver metastasis.

Circ-sirt1 inhibits growth and invasion of gastric cancer by sponging miR-132-3p/miR-212-3p and upregulating sirt1 expression.

circRNAs have been considered as a rising factor in cancers. However, the roles and mechanisms of circ-sirt1 in gastric cancer (GC) remain largely unknown. In this study, we found that the expressions of sirt1 and circ-sirt1 are decreased in tissues or serums of GC patients by real-time quantitative PCR (RT-qPCR). The expressions of miR-132-3p/miR-212-3p showed an opposite tendency in these samples. The co-transfection of miR-132-3p/miR-212-3p mimics counteracted the enhancement of sirt1 expression induced by circ-sirt1. The results of cell colony-formation assay and transwell assays demonstrated that the proliferation, migration, and invasion activities of BGC-823 cells were inhibited by circ-sirt1 overexpression or miR-132-3p/miR-212-3p knockdown, respectively. The xenograft tumor model result indicated that the circ-sirt1 overexpression suppressed the tumor growth of BGC-823 cells. The regulation of miR-132-3p/miR-212-3p between circ-sirt1 and sirt1 was verified in the mice tumor tissues. Thus, circ-sirt1 inhibited tumor growth and invasion probably by sponging miR-132-3p/miR-212-3p and upregulating sirt1 expression in GC. These findings may provide a theoretical basis for the classification of GC and a novel therapeutic target for GC patients.

Exosomal­miR­10a derived from colorectal cancer cells suppresses migration of human lung fibroblasts, and expression of IL­6, IL­8 and IL­1ß.

MicroRNAs (miRs) carried in exosomes serve an important role in the pre­metastatic microenvironment and in intercellular interactions. However, the function of exosomal­miR­10a derived from primary colorectal cancer (CRC) cells on fibroblasts in the lung metastatic microenvironment of patients with CRC remains unclear. Reverse transcription­quantitative PCR was performed using samples from patients with CRC, and demonstrated that the expression levels of miR­10a were significantly lower in serum and cancer tissue samples from patients with CRC compared with in serum from healthy individuals and paired non­cancerous tissues, respectively. In addition, the expression levels of miR­10a were inversely associated with the invasion depth of CRC. Exosomal­miR­10a derived from CRC cells reduced the proliferative and migratory activities of primary normal human lung fibroblasts (NHLFs), and the expression levels of IL­6, IL­8 and IL­1ß in NHLFs. The present study provided insight into the phenotypic alterations of NHLFs induced by exosomal­miR­10a derived from CRC cells, which may aid understanding of the mechanism underlying the process of CRC lung metastasis.

Correction to: miR-3928v is induced by HBx via NF-κB/EGR1 and contributes to hepatocellular carcinoma malignancy by down-regulating VDAC3.

An amendment to this paper has been published and can be accessed via the original article.

Correction to: miR-484 suppresses proliferation and epithelial-mesenchymal transition by targeting ZEB1 and SMAD2 in cervical cancer cells.

[This corrects the article DOI: 10.1186/s12935-017-0407-9.].

WSTF acetylation by MOF promotes WSTF activities and oncogenic functions.

Williams syndrome transcription factor (WSTF) is a transcription factor and tyrosine kinase. WSTF overexpression promotes migration and proliferation of various cancers, and Ser158 (WSTFS158) phosphorylation plays an important role in this process. However, the role of the other posttranslational modifications of WSTF is unknown. Here, we report that lysine (K) 426 on WSTF is acetylated by MOF and deacetylated by SIRT1. Mechanistically, male-specific lethal (MSL) 1v1 interaction with WSTF facilitates its interaction with MOF for WSTF acetylation, which in turn promotes WSTFS158 phosphorylation. The kinase and transcriptional regulatory activity of WSTF were enhanced by acetylation. WSTFK426ac levels positively and significantly correlated with tumor size, histological grade, and age. Moreover, we demonstrated that acetylated WSTF promotes cancer cell proliferation, migration, invasion, and tumor formation. In conclusion, we identified the enzymes regulating WSTF K426 acetylation, and demonstrated an acetylation-dependent mechanism that modulates the activities of WSTF and contributes to tumorigenesis. Our findings provide new clues to study WSTF-mediated normal development and disease.

miR-639 Expression Is Silenced by DNMT3A-Mediated Hypermethylation and Functions as a Tumor Suppressor in Liver Cancer Cells.

Emerging evidence has indicated that abnormal methylation of DNA contributes to hepatocarcinogenesis. However, the regulatory mechanisms are not well known. Here, we revealed that microRNA-639 (miR-639) expression is downregulated in liver cancer tissues and cells. The repression of miR-639 expression was attributed to hypermethylation in its promoter region, and DNA methyltransferase (DNMT3A) was found to mediate this hypermethylation. Repression of miR-639 expression promoted cell growth and migration/invasion in vitro and the growth of tumors in xenograft mouse models. Furthermore, miR-639 bound to the 3' UTR of both MYST2 and ZEB1 and suppressed their expression. MYST2 promoted the growth of liver cancer cells and ZEB1 facilitated the migration/invasion of liver cancer cells. Ectopic expression of MYST2 and ZEB1 counteracted the repression of malignancy induced by miR-639, which coincided with the reciprocal correlation between miR-639 and MYST2 and ZEB1 expression in clinical hepatocellular carcinoma (HCC) tissues. Thus, DNMT3A-mediated hypermethylation suppressed miR-639 expression, derepressing the expression of MSYT2 and ZEB1, which promoted tumorigenesis of liver cancer. These findings may shed light on the mechanism of abnormal expression of miRNAs involved in the malignancy of liver cancer and provide new biomarkers for liver cancer.