Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: ZNF518B is overexpressed in CRC patients. The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples. The plots correspond to the comparison of whole gene and its major isoforms in normal versus all tumour tissues or stage-classified tumour tissues. Results were compared with the Kruskal-Wallis test. (*p < 0.05).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Expressing

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Overexpression of the ZNF518B canonical isoform 1 in a prospective cohort of CRC patients from our University Hospital (70 tumour and 69 non-tumour adjacent samples). The figure shows box plots with whiskers as in Fig. , including the position of medians. Results were compared with the Mann-Whitney test. (***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Over Expression, MANN-WHITNEY

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Effects of ZNF518B knocking-down on the proliferation of DLD1 and HCT116 cells. ( a ) Efficiency of knocking-down for isoforms 1 and 2 in DLD1 and HCT116 cells after 48 h of transfection. ( b ) Influence of knocking-down on cell proliferation as measured by MTT assays; continuous lines, control cells; broken lines, cells transfected with ZNF518B siRNA. ( c ) Effects of ZNF518B knocking-down in a representative colony formation assay. ( d ) Quantification of the different cell colony formation assays. Statistical analysis of data in a and d was carried out with the Mann-Whitney test, whereas two-way ANOVA was used for data in panel b. (***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Transfection, Colony Assay, MANN-WHITNEY

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Effects of ZNF518B knocking-down on the migration and invasiveness of DLD1 and HCT116 cells. ( a , b ) Representative images and quantitative analysis showing the suppression of migration of DLD1 and HCT116 cells by knocking-down the ZNF518B gene. The assay was done in transwell chambers. ( c ) Wound-healing assay showing the diminution of cell migration after silencing the ZNF518B gene. ( d , e ) Representative images and quantitative analysis showing the suppression of invasiveness of DLD1 and HCT116 cells by knocking-down the ZNF518B gene. The assay was carried out in Matrigel-coated transwell chambers. ( f ) Quantification of cell-adhesion assays to collagen type I-coated dishes, showing the loss of adhesiveness after silencing the ZNF518B gene. Statistical analysis was carried out with the Mann-Whitney test. (***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Migration, Wound Healing Assay, MANN-WHITNEY

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Effects of ZNF518B expression on the level of EMT markers. DLD1 cells were treated with mixed ZNF518B siRNAs or with scrambled siRNA (control) for 48 and 72 h and the EMT markers were analysed. The levels of the SNAI1 ( a ) and CHD1 ( b ) genes were determined by RT-qPCR. ( c ) Western blots showing the level of ZNF518B (left) and N-cadherin and SNAIL (right) after 72 h of silencing ZNF518B . Histone deacetylase 1 (HDAC) or glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were used as loading controls. Due to the different size of ZNF518B and of the EMT markers, the electrophoreses were carried out separately in 6% and 12% polyacrylamide gels, respectively. Ponceau-stained membranes were cropped according to the molecular weight of the corresponding proteins and the resulting strips were developed with the antibodies described under Materials and Methods. ( d ) ImageJ analysis of the western blots of panel c . Four grey values relative to the loading controls were measured in every case and averaged. Statistical analysis was carried out with the Mann-Whitney test. (*p < 0.05; ***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Histone Deacetylase Assay, Staining, Molecular Weight, MANN-WHITNEY

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Organisation of chromatin at the promoter and proximal transcribed region of ZNF518B gene. The experiments were carried out with DLD1, SW48 and D-Mut1 cells. ( a ) Micrococcal nuclease protection assay. The protection, determined as described in the text, is plotted against the position of the centre of the tiled amplicons used, whose location is given below. The coordinates are given in bp relative to TSS. ( b ) Model for nucleosome positioning. ( c ) Detail of the nuclease protection assay in the region from −200 to +200. ( d ) Prediction of the nucleosome occupancy score ( red ) and of the probability of starting nucleosomes ( blue ), as determined by the NuPoP program (12).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques:

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Epigenetic modification of histones in the promoter and proximal coding region of the ZNF518B gene. The experiments were carried out at single-nucleosome level by using the Nuc-ChIP approach. ( A ) Acetylation marks in nucleosomes N − 3, N − 2 and N − 1. ( B ) Methylation marks at amplicon 75, which covers the 5′ part of nucleosome N + 1 in DLD1 and D-Mut1 cells and the 3′ part of the nucleosome covering the TSS in SW48 cells. Results were compared with the Kruskal-Wallis test. (**p < 0.005; ***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Modification, Methylation, Amplification

Journal: Scientific Reports

Article Title: ZNF518B gene up-regulation promotes dissemination of tumour cells and is governed by epigenetic mechanisms in colorectal cancer

doi: 10.1038/s41598-019-45411-9

Figure Lengend Snippet: Effects of the inhibition of histone deacetylases on the expression of ZNF518B gene and in the migration of SW48 cells. ( a ) A culture of cells was treated either with trichostatin A (TSA) or with the solvent alone (DMSO) for 24 or 48 h and the level of ZNF518B expression relative to ACTB was determined by RT-qPCR as described in the text. ( b ) Representative images of SW48 cells migration determined by transwell analysis in the absence or presence of TSA for 24 h. ( c ) Quantification of cell migration. Two areas of three independent wells were counted and the six values averaged. Data were analysed with Student’s t-test. (**p < 0.005; ***p < 0.001).

Article Snippet: The figure shows box plots with whiskers with maximum 1.5 IQR of the ZNF518B expression measured by qPCR in an OriGene cDNA array of normal and tumour samples.

Techniques: Inhibition, Expressing, Migration, Quantitative RT-PCR