The 5-year survival rate for colorectal cancer is approximately 55 % because of its invasion and metastasis. The epithelial–mesenchymal transition (EMT) is one of the well-defined processes during the invasion and distant metastasis of primary epithelial tumors. miR-429, a member of the miR-200 family of microRNAs, was previously shown to inhibit the expression of transcriptional repressors ZEB1/delta EF1 and SIP1/ZEB2, and regulate EMT. In this study, we showed that miR-429 was significantly downregulated in colorectal carcinoma (CRC) tissues and cell lines. We found that miR-429 inhibited the proliferation and growth of CRC cells in vitro and in vivo, suggesting that miR-429 could play a role in CRC tumorigenesis. We also showed that downregulation of miR-429 may contribute to carcinogenesis and the initiation of EMT of CRC by targeting Onecut2. Further researches indicated that miR-429 inhibited the cells migration and invasion and reversed TGF-β-induced EMT changes in SW620 and SW480 cells. miR-429 could reverse the change of EMT-related markers genes induced by TGF-β1, such as E-cadherin, CTNNA1, CTNNB1, TFN, CD44, MMP2, Vimentin, Slug, Snail, and ZEB2 by targeting Onecut2. Taken together, our data showed that transcript factor Onecut2 is involved in the EMT, migration and invasion of CRC cells; miR-429 inhibits the initiation of EMT and regulated expression of EMT-related markers by targeting Onecut2; and miR-429 or Onecut2 is the important therapy target for CRC. Electronic supplementary materialThe online version of this article (doi:10.1007/s11010-013-1950-x) contains supplementary material, which is available to authorized users.

We report here the detection of the release of cytochrome c from mitochondria into the cytosol during menadione-induced apoptosis in tobacco protoplasts. Western blot analysis indicated that the caspase specific inhibitors AC-DEVD-CHO (Ac-Asp-Glu-Val-Asp-aldehyde) and AC-YVAD-CHO (N-acetyl-Try-Val-Ala-aspartinal) inhibited the degradation of a caspase 3 specific substrate PARP (poly(ADP-ribose) polymerase), and they had no effect on the release of cytochrome c. Further study showed that menadione could not induce apoptosis of mouse liver nuclei in tobacco cytosol extract containing no mitochondria. However, when cytochrome c or mitochondria was added into the cytosol extract, apoptosis of mouse liver nuclei and the degradation of PARP could both be detected. The results provide strong evidence that menadione can induce apoptosis in tobacco protoplasts via the release of cytochrome c from mitochondria into the cytosol.z 1999 Federation of European Biochemical Societies.

The adenomatous polyposis coli (APC) is a frequently mutated tumour suppressor gene in cancers. However, whether APC is regulated at the epitranscriptomic level remains elusive. In this study, we analysed TCGA data and separated 200 paired oesophageal squamous cell carcinoma (ESCC) specimens and their adjacent normal tissues and demonstrated that methyltransferase-like 3 (METTL3) is highly expressed in tumour tissues. m6A-RNA immunoprecipitation sequencing revealed that METTL3 upregulates the m6A modification of APC, which recruits YTHDF for APC mRNA degradation. Reduced APC expression increases the expression of β-catenin and β-catenin-mediated cyclin D1, c-Myc, and PKM2 expression, thereby leading to enhanced aerobic glycolysis, ESCC cell proliferation, and tumour formation in mice. In addition, downregulated APC expression correlates with upregulated METTL3 expression in human ESCC specimens and poor prognosis in ESCC patients. Our findings reveal a mechanism by which the Wnt/β-catenin pathway is upregulated in ESCC via METTL3/YTHDF-coupled epitranscriptomal downregulation of APC.

BackgroundDespite the overwhelming number of investigations on AKT, little is known about lncRNA on AKT regulation, especially in GBM cells.MethodsRNA-binding protein immunoprecipitation assay (RIP) and RNA pulldown were used to confirm the binding of LINC00470 and fused in sarcoma (FUS). Confocal imaging, co-immunoprecipitation (Co-IP) and GST pulldown assays were used to detect the interaction between FUS and AKT. EdU assay, CCK-8 assay, and intracranial xenograft assays were performed to demonstrate the effect of LINC00470 on the malignant phenotype of GBM cells. RT-qPCR and Western blotting were performed to test the effect of LINC00470 on AKT and pAKT.ResultsIn this study, we demonstrated that LINC00470 was a positive regulator for AKT activation in GBM. LINC00470 bound to FUS and AKT to form a ternary complex, anchoring FUS in the cytoplasm to increase AKT activity. Higher pAKT activated by LINC00470 inhibited ubiquitination of HK1, which affected glycolysis, and inhibited cell autophagy. Furthermore, higher LINC00470 expression was associated with GBM tumorigenesis and poor patient prognosis.ConclusionsOur findings revealed a noncanonical AKT activation signaling pathway, i.e., LINC00470 directly interacts with FUS, serving as an AKT activator to promote GBM progression. LINC00470 has an important referential significance to evaluate the prognosis of patients.Electronic supplementary materialThe online version of this article (10.1186/s13045-018-0619-z) contains supplementary material, which is available to authorized users.

We developed and characterized a novel picoliter droplet-in-oil array generated by a double-inkjet printing method on a uniform hydrophobic silicon chip specifically designed for quantitative polymerase chain reaction (qPCR) analysis. Double-inkjet printing was proposed to efficiently address the evaporation issues of picoliter droplets during array generation on a planar substrate without the assistance of a humidifier or glycerol. The method utilizes piezoelectric inkjet printing equipment to precisely eject a reagent droplet into an oil droplet, which had first been dispensed on a hydrophobic and oleophobic substrate. No evaporation, random movement, or cross-contamination was observed during array fabrication and thermal cycling. We demonstrated the feasibility and effectiveness of this novel double-inkjet method for real-time PCR analysis. This method can readily produce multivolume droplet-in-oil arrays with volume variations ranging from picoliters to nanoliters. This feature would be useful for simultaneous multivolume PCR experiments aimed at wide and tunable dynamic ranges. These double-inkjet-based picoliter droplet arrays may have potential for multiplexed applications that require isolated containers for single-cell cultures, single molecular enzymatic assays, or digital PCR and provide an alternative option for generating droplet arrays on planar substrates without chemical patterning.

We report here the apoptosis of mouse liver nuclei induced in the cytosol of carrot cells by cytochrome c. Several typical characteristics of apoptosis, such as chromatin condensation, margination and apoptotic bodies, were detected. The result of DNA gel electrophoresis showed that DNA was degraded into nucleosomal fragments. The terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labelling procedure was also performed to detect the breakage of 3P-OH ends of a DNA strand. Furthermore, we found that nuclear lamins were degraded from 88 kDa and 66 kDa to 37 kDa and 47 kDa fragments. The DNA fragmentation could be inhibited by AC-DEVD-CHO and AC-YVAD-CHO. The results indicate that the apoptosis in plant cells may share some similar pathways to apoptosis in animal cells.z 1999 Federation of European Biochemical Societies.

Tumor-associated macrophages (TAMs) constitute a major component of inflammatory cells in the glioblastoma multiforme (GBM) tumor microenvironment. TAMs have been implicated in GBM angiogenesis, invasion, local tumor recurrence, and immunosuppression. Coagulation factor X (FX) is a vitamin K-dependent plasma protein that plays a role in the regulation of blood coagulation. In this study, we first found that FX was highly expressed and positively correlated with TAM density in human GBM. FX exhibited a potent chemotactic capacity to recruit macrophages and promoted macrophages toward M2 subtype polarization, accelerating GBM growth. FX bound to extracellular signal-related kinase (ERK)1/2 and inhibited p-ERK1/2 in GBM cells. FX was secreted in the tumor microenvironment and increased the phosphorylation and activation of ERK1/2 and AKT in macrophages, which may have been responsible for the M2 subtype macrophage polarization. Moreover, although the lncRNA CASC2c has been verified to function as a miR-101 competing endogenous RNA (ceRNA) to promote miR-101 target genes in GBM cells, we first confirmed that CASC2c did not function as a miR-338-3p ceRNA to promote FX expression, and that FX was a target gene of miR-338-3p. CASC2c interacted with and reciprocally repressed miR-338-3p. Both CASC2c and miR-388-3p bound to FX and commonly inhibited its expression and secretion. CASC2c repressed M2 subtype macrophage polarization. Taken together, our findings revealed a novel mechanism highlighting CASC2c and FX as potential therapeutic targets to improve GBM patients by altering the GBM microenvironment.

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