Journal: International Journal of Molecular Sciences

Article Title: Hepatocyte Nuclear Factor 4α (HNF4α) Plays a Controlling Role in Expression of the Retinoic Acid Receptor β ( RARβ ) Gene in Hepatocytes

doi: 10.3390/ijms24108608

Figure Lengend Snippet: HNF4α regulates the promoter of the RARβ gene in HepG2 cells. HepG2 cells in 24-well plates were co-transfected for 24 h with a fragment of the human RARβ promoter expanding from −1.7 kbp from transcription start site to +0.217 kpb, as the full length promoter construct, in p GL3-basic-luc vector together with p RLTK plasmid containing Renilla-luc, as the control, and with either human HNF4α, RARα/RXRα, or all three transcription factors, and then treated further either without ( A ) or with RA for 1 to 24 h, after which the cells were collected to assay for luciferase activity. HNF4α alone or with RARα/RXRα suppressed the promoter activity of the human RARβ gene in the cells treated without RA ( A ), but upregulated the promoter when the cells were treated with RA following transfection ( B ). HNF4α regulates the promoter of the mouse RARβ gene in HepG2 cells ( C ). It suppresses the promoter activity of the mouse gene (empty bars in C) in HepG2 cells treated with the vehicle, but it increases the promoter activity in the cells treated with RA (black bar in C).

Article Snippet: Construction of the plasmid vectors including p GL3-Basic-hCYP26A1-E4-luc (submitted to addgene.org), p GL3-Basic-hRARβp-luc (human RARβ2 promoter), p GL3-Basic-hCYP2C9p-luc, p cDNNA3.1-hRARα.hRXRα (submitted to addgene.org), and p cDNA3.1-hHNF4α were reported previously [ , , , , ].

Techniques: Transfection, Construct, Plasmid Preparation, Control, Luciferase, Activity Assay

Journal: International Journal of Molecular Sciences

Article Title: Hepatocyte Nuclear Factor 4α (HNF4α) Plays a Controlling Role in Expression of the Retinoic Acid Receptor β ( RARβ ) Gene in Hepatocytes

doi: 10.3390/ijms24108608

Figure Lengend Snippet: Mutation of the critical residues present in the ligand binding domain of human HNF4α suppresses transcription activation of the promoters of human APOC3 and CYP2C9 genes. HepG2 cells were co-transfected with either the p GL3-b-hApoC3 ( A – C ) or p GL3-b-hCYP2C9 ( D – F ) promoter construct together with either hHNF4α, RARα/RXRα, or all three transcription factors, and then treated with either vehicle or 1 µM RA for 24 h, after which the cells were assayed for their luciferase activity. The effects of the individual mutant residues in the ligand binding domain of the human HNF4α compared with WT HNF4α on the promoter activity of APOC3 ( B ) or CYP2C9 ( E ) in HepG2 cells treated with either vehicle or 1 μM RA for 24 h. The endogenous HNF4α transcriptional activity toward APOC3 ( C ) and CYP2C9 ( F ) promoters was assessed in HepG2 cells with or without the addition of human HNF4α mutants. Data from each bar represent the mean of n = 3 wells ± SD. HNF4α mutant #’s are as follows: (1) S190K, (2) 191MK, (3) R221G, (4) L228K, (5) L229K, (6) R235G, (7) I347K, and (8) I355K.

Article Snippet: Construction of the plasmid vectors including p GL3-Basic-hCYP26A1-E4-luc (submitted to addgene.org), p GL3-Basic-hRARβp-luc (human RARβ2 promoter), p GL3-Basic-hCYP2C9p-luc, p cDNNA3.1-hRARα.hRXRα (submitted to addgene.org), and p cDNA3.1-hHNF4α were reported previously [ , , , , ].

Techniques: Mutagenesis, Ligand Binding Assay, Activation Assay, Transfection, Construct, Luciferase, Activity Assay

Journal: International Journal of Molecular Sciences

Article Title: Hepatocyte Nuclear Factor 4α (HNF4α) Plays a Controlling Role in Expression of the Retinoic Acid Receptor β ( RARβ ) Gene in Hepatocytes

doi: 10.3390/ijms24108608

Figure Lengend Snippet: Mutation of the critical amino acid residues present in the ligand binding domain of human HNF4α suppresses transcription activation of the promoters of human CYP26A1 and RARβ genes in HepG2 cells treated with RA. HepG2 cells grown in 24-well plates were co-transfected with either p GL3-b-hCYP26A1 ( A ) or p GL3-b-hRARβ ( B ) promoters, each with p RLTK as the control, together with either wildtype (WT) HNF4α or its individual mutants (Mutant # 1 to 8), and then treated with either vehicle or 1 µM RA for 24 h, after which the cells were assayed for luciferase activity. Data from each bar represent the mean of n = 3 wells ± SD. HNF4α mutant #’s are as follows: (1) S190K, (2) 191MK, (3) R221G, (4) L228K, (5) L229K, (6) R235G, (7) I347K, and (8) I355K.

Article Snippet: Construction of the plasmid vectors including p GL3-Basic-hCYP26A1-E4-luc (submitted to addgene.org), p GL3-Basic-hRARβp-luc (human RARβ2 promoter), p GL3-Basic-hCYP2C9p-luc, p cDNNA3.1-hRARα.hRXRα (submitted to addgene.org), and p cDNA3.1-hHNF4α were reported previously [ , , , , ].

Techniques: Mutagenesis, Ligand Binding Assay, Activation Assay, Transfection, Control, Luciferase, Activity Assay

Journal: Cells

Article Title: MYH9 Facilitates Cell Invasion and Radioresistance in Head and Neck Cancer via Modulation of Cellular ROS Levels by Activating the MAPK-Nrf2-GCLC Pathway

doi: 10.3390/cells11182855

Figure Lengend Snippet: MYH9 modulated intracellular ROS level leads to radioresistance in HNC cells. ( A ) MYH9 silencing increased ROS production in HNC cells. After transfection of MYH9-shRNA plasmids in theboth HNC cells (OECM-1 and Detroit 562), the ROS level was determined using the H2DCF-DA oxidation method and analyzed by flow cytometry. ( B ) MYH9 silencing reduced the Nrf2 transcriptional activity. Upper: A schematic representation of the pGL3 promoter-8xARE construct which was identified for measuring Nrf2 transcriptional activity. Lower: A luciferase report assay was used to compare the relative level of Nrf2 transcriptional activity in the HNC cells (OECM-1 and Detroit 562) transfected with MYH9-shRNA (sh-M) versus control transfectant (Vt). ( C , D ) MYH9 silencing inhibited the expression of ROS scavenger enzymes at the mRNA level of the OECM1 cells ( C ) and the protein ( D ) levels in two HNC cell lines. ( E ) Significantly higher expressions of ROS scavenger enzymes (GCLC, and GCLM) in HNC tumor samples compared to the normal samples. Correlative expressions between MYH9 and ROS scavenger enzymes-associated molecules GCLC and GCLM, in the HNC tumor samples. The gene expression data were retrieved from the TCGA-HNSC dataset. All the experiments were performed at least three times independently, and a typical result was shown. The error bars shown in the relevant figures indicate the standard deviation of the quantification results in all experiments. (*: p < 0.05, **: p < 0.01, ***: p < 0.001, t -test).

Article Snippet: The plasmid of the pGL3 promoter-8xARE luciferase reporter construct purchased from MDbio Inc. (Taipei, Taiwan) was used to assess Nrf2 transcriptional activity.

Techniques: Transfection, shRNA, Flow Cytometry, Activity Assay, Construct, Luciferase, Control, Expressing, Gene Expression, Standard Deviation

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: (a) Representative Western blot analysis of DLC1 and α-tubulin (loading control) in total lysates of HUVECs cultured on fibronectin-coated 2 kPa or 50 kPa hydrogels, or plastic. Graphs show the average protein expression levels ±s.e.m. Signal corrected for background and relative to expression in HUVECs on plastic. Data from 4 independent experiments. *P<0.05, **P<0.01, ***P<0.001 (One-way ANOVA and Dunnett’s post-test). (b) Quantitative real time PCR analysis of DLC1 mRNA isolated from HUVECs cultured on fibronectin-coated 2 kPa or 50 kPa hydrogels, or plastic. Graph shows the average gene expression levels ±s.e.m. Values are normalized to housekeeping gene (RPLP0) expression levels and relative to mRNA expression levels in HUVECs on plastic. Data from 3 independent experiments. n.s.; not significant, *P<0.05, **P<0.01 (One-way ANOVA and Dunnett’s post-test). (c) Schematics of UCSC genome browser results at position chr8:13,074,715-13,142,890 of the human genome (GRCh38/hg38) displaying the genomic location of DLC1 transcript variants 1 (NP872584.2), 2 (NP.006085.2) and 5 (NP.001303597.1) and the presence of a TEAD motif at the transcriptional start site of DLC1 transcript variant 2. Plotted are the results from publicly available GEO data TEAD1 ChIP-Seq data from various cell types, the data show a binding peak of TEAD1 at the TSS of DLC1 isoform 2. See for more details including histone modification profiles and DNase hypersensitivity profiles of the promoter region in HUVECs. (d) Boxplot showing the mean (± upper and lower quartiles) relative promoter activity of DLC1 in lysates of HEK cells transfected with wild type human DLC1 promoter region (from -418 to +319 bp position of the transcriptional start site of DLC1 isoform 2) fused to a firefly luciferase reporter or a DLC1-promoter luciferase reporter in which the TEAD-binding motif was mutated from CATTCCA to AGACTAT. Firefly luciferase activities were normalized to co-transfected renilla luciferase activity. Data is from 3 independent experiments. Whiskers show min/max values. *P<0.05 (Two-tailed unpaired Student’s t-test). (e, f) Representative Western blot analysis of DLC1 in total lysates of HUVECs transduced with shRNA Control, shRNA YAP (e) or shRNA TAZ (f) . Graphs show the average protein expression levels ±s.e.m. Signal corrected for background and normalized to expression in shControl-transduced HUVECs. Data from 3 independent experiments. *P <0.05, **P <0.01, ***P<0.001 (Paired Student’s t-test). (g) Representative Western blot analysis of DLC1, pYAP Serine 127 and β-actin (loading control) in total cell lysate samples of non-stimulated HUVECs (-) or HUVECs stimulated with 1mg/ml VEGF for 2 hours. Graphs show the average protein expression levels ±s.e.m. Signal corrected for background and normalized to expression in non-transduced HUVECs. Data from 4 independent experiments. *P<0.05 (Paired Student’s t-test). (h) Representative Western blot analysis of DLC1, YAP1, Myc, CTGF and β-actin (loading control) in total lysates of non-transduced HUVECs (-) and HUVECs transduced with Myc-tagged human YAP-5SA. Graphs show the average protein expression levels ±s.e.m. Signal corrected for background and normalized to expression in non-transduced HUVECs. Data from 4 independent experiments. *P<0.05 (Paired Student’s t-test). (i) Representative Western blot analysis of DLC1, YAP1, Myc, CTGF and β-actin (loading control) in total lysates of non-transduced HUVECs (-) and HUVECs transduced with Myc-tagged human YAP-5SA cultured on 2 kPa hydrogels. Graphs show the average protein expression levels ±s.e.m. Signal corrected for background and normalized to expression in non-transduced HUVECs. Data from 3 independent experiments. *P<0.05 (Paired Student’s t-test). Scans of whole Western blots are depicted in .

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Western Blot, Cell Culture, Expressing, Real-time Polymerase Chain Reaction, Isolation, Variant Assay, ChIP-sequencing, Binding Assay, Modification, Activity Assay, Transfection, Luciferase, Two Tailed Test, Transduction, shRNA

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: Full detailed schematics of UCSC genome browser results at position chr8:13,074,715-13,142,890 of the human genome (GRCh38/hg38) showing the genomic location of DLC1 transcript variants 1 (NP_872584.2), 2 (NP_006085.2), 5 (NP_001303597.1) and 4 (NP_001157743.1) and the presence of a TEAD motif at the transcriptional start site (TSS) of DLC1 transcript variant 2. Plotted are the results from publicly available GEO data TEAD1 ChIP-Seq data from various cell types and corresponding histone modification profiles in HUVECs in ENCODE. The data show a binding peak of TEAD1 at the TSS of DLC1 transcript variant 2. Histone modification profiles indicate that there is an open conformation of chromatin and an active promoter region around the TEAD binding motif, defined as the bimodal presence of both histone H3 trimethylation at lysine 4 (H3K4me3) and histone H3 acetylation at lysine 27 (H3K27ac), combined with increased DNase hypersensitivity.

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Variant Assay, ChIP-sequencing, Modification, Binding Assay

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: (a) Representative Western blot analysis of DLC1 and β-actin (loading control) in total lysates of HUVECs transduced with different shDLC1 clones (#1067, #1066, #1065, #1064, #1063). (b) Widefield immunofluorescence (IF) images of HUVECs transduced with shControl or shDLC1 (pool of #1063/1064) and stained for VE-cadherin (green) and F-actin (red). (c) Line graph shows the average ± stdev transendothelial impedance measured at 4000 Hz across barrier forming endothelial cells transduced with shControl or shDLC1 (pool of #1063/1064) plated on fibronectin-coated 8W10E ECIS arrays. Representative data is from 2 independent experiments and an average of 6 wells per condition. (d) Boxplot showing quantification of RhoA activity in G-LISA assays in lysates of shControl and shDLC1 transduced HUVECs, with or without thrombin stimulation. Data is from 4 independent experiments. Whiskers show min/max values. n.s.; not significant (Two-tailed unpaired Student’s t-test). (e) Representative widefield IF images of HUVECs transduced with GFP-DLC1 stained for Paxillin pY118 (red) and F-actin (blue). (f) Widefield IF images of shControl and shDLC1 transduced HUVECs and stained for Paxillin pY118 (green), F-actin (blue), VE-cadherin (red). Boxplot showing quantification of manually counted number of focal adhesions per µm2 of shControl and shDLC1 transduced HUVECs. Whiskers show min/max values. Data is from 2 independent experiments, shControl (42 cells from 12 images) and shDLC1 (46 cells from 19 images). **P<0.01 (Two-tailed unpaired Student’s t-test). (g) Left images are stills from time lapse TIRF microscopy at t=0 of HUVECs transduced with shControl and shDLC1 and paxillin-mCherry. Heat map shows the corresponding focal adhesion dynamics over 30 minutes in a unique color per time frame. Note the stability of focal adhesions in shDLC1 HUVECs as indicated by the white colored FAs in the right panels. See corresponding Supplemental Movie 1 for the ∼2.5 hours time-lapse recording. (h) Bar graph showing quantification of focal adhesion lifetime, assembly and disassembly rates based on TIRF time lapse experiments with paxillin-mCherry expressing HUVECs. Error bars are s.e.m. Data is from 2 independent experiments, shControl (6 movies) and shDLC1 (8 movies). Focal adhesion tracking was performed using the focal adhesion analysis webserver . **P<0.01 (Two-tailed unpaired Student’s t-test). (i) DIC images and cell-substrate traction force maps of HUVECs transduced with shControl or shDLC1. Boxplot showing the mean (± upper and lower quartiles) of measured RMS traction forces of shControl (12 image fields) and shDLC1 (18 image fields) endothelial cells. Data is from 3 independent experiments. Whiskers show min/max values. **P<0.01 (Two-tailed unpaired Student’s t-test).

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Western Blot, Transduction, Clone Assay, Immunofluorescence, Staining, Activity Assay, Two Tailed Test, Microscopy, Expressing

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: (a) Representative Phase-contrast images of HUVECs transduced with shControl, shDLC1 3’UTR or shDLC1 #1063, in a scratch-wound assay (t=0, t=4, t=8 and t=12 hours after scratch). Yellow line highlights the unclosed wound area. See corresponding Supplemental Movie 2 for the ∼16 hours timelapse recording. (b) Graphs showing the percentage of wound closure of HUVECs transduced with shControl, shDLC1 3’UTR or shDLC1 #1063 at three time points during scratch-wound assay. Data is from 4 independent experiments, shControl (22 movies), shDLC1 3’UTR (14 movies) and shDLC1 #1063 (18 movies). *P<0.05, ***P<0.001 (Two-way ANOVA and Tukey’s multiple comparison test). (c) Representative Western blot analysis of DLC1 and α-tubulin (loading control) in total lysates of HUVECs transduced with shControl, shDLC1 3’UTR or shDLC1 #1063. Scans of whole Western blots are depicted in . (d) Boxplot showing the mean (± upper and lower quartiles) of proliferation ratio of the shControl, shDLC1 3’UTR, shDLC1 #1063 HUVECs corresponding to the scratch assays in Fig. 3b. Proliferation ratio was manually determined by counting cell numbers at ROIs within the endothelial monolayers before and after 12 hrs scratch wound migration. n.s.; not significant (One-way ANOVA with Dunnett’s multiple comparison test). (e) Phase-contrast images of scratch wound assays overlayed with single cell tracking analysis of 8 hours using the Chemotaxis Tool in ImageJ. Boxplot showing the mean (± upper and lower quartiles) velocity and directionality of shControl, shDLC1 3’UTR or shDLC1 #1063 transduced HUVECs. Whiskers show min/max values. Data is from 4 independent experiments, shControl (429 cells from 22 movies), shDLC1 3’UTR (236 cells from 12 movies), shDLC1 #1063 (303 cells from 18 movies). n.s.; not significant, ***P<0.001 (One-way ANOVA and Dunnett’s multiple comparison test). (f) Widefield IF images of HUVECs transduced with shControl, shDLC1 3’UTR or shDLC1 #1063 6 hours after initiation of scratch wound assays stained for DAPI (blue), GM130 (red; Golgi) and VE-cadherin (green). The dotted lines indicate migration front and the asterisks indicate cells containing an oriented Golgi in the direction of scratch wound migration. Boxplot showing the mean (± upper and lower quartiles) of cells with an oriented Golgi. Data is from 3 independent experiments, shControl (34 images), shDLC1 3’UTR (30 images), shDLC1 #1063 (35 images). ***P<0.001 (One-way ANOVA with Dunnett’s multiple comparison test). (g) Widefield IF images of HUVECs transduced with shControl, shDLC1 3’UTR or shDLC1 #1063 6 hours after initiation of scratch wound assays stained for Vinculin (red), F-actin (blue) and VE-cadherin (green). Boxplots show the mean (± upper and lower quartiles) Focal Adhesion Alignment Index and Focal Adhesion dominant angle as determined by image analysis of the vinculin channels using the Focal Adhesion Analysis Server . Data is from 3 independent experiments, shControl (34 images), shDLC1 3’UTR (15 images), shDLC1 #1063 (36 images). Whiskers show min/max value. n.s.; not significant, *P< 0.05, **P<0.01 (One-way ANOVA with Dunnett’s multiple comparison test).

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Transduction, Scratch Wound Assay Assay, Western Blot, Migration, Single Cell Tracking, Chemotaxis Assay, Staining

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: (a) Representative phase-contrast images of sprouting spheroids 16 hours after VEGF stimulation of HUVECs lentivirally transduced with shControl, shDLC1 (#1064/1065). Boxplot shows the mean (± upper and lower quartiles) cumulative sprout length and the number of sprouts in the spheroid-based sprouting angiogenesis assays. Whiskers show min/max values. Data is from 3 independent experiments, shControl (63 spheroids) and shDLC1 (47 spheroids). **P<0.01 (Two-tailed unpaired Student’s t-test). (b) Representative phase-contrast images of sprouting spheroids 16 hours after VEGF stimulation of HUVECs lentivirally transduced with GFP or GFP-DLC1. Boxplot shows the mean (± upper and lower quartiles) cumulative sprout length and the number of sprouts in the spheroid-based sprouting angiogenesis assays. Whiskers show min/max values. Data is from 3 independent experiments, GFP (25 spheroids), GFP-DLC1 (27 spheroids). ***P<0.001 (Two-tailed unpaired Student’s t-test). (c) Representative phasecontrast images of sprouting spheroids 16 hours after VEGF stimulation of HUVECs transduced with shControl or shDLC1 3’UTR and rescued with GFP, GFP-DLC1 or GFP-DLC1-R677E. Boxplots show the mean (± upper and lower quartiles) cumulative sprout length and number of sprouts per spheroid. Whiskers present min/max values. Data is from 3 independent experiments, shControl (33 spheroids), shDLC1 3’UTR (29 spheroids), rescue GFP (39 spheroids), rescue GFP-DLC1 (37 spheroids), rescue GFP-DLC1 R667E (28 spheroids). n.s. non-significant, *P<0.05, ***P<0.001 (One-way ANOVA with Dunnett’s multiple comparisons test). (d) Representative Western blot analysis of DLC1, GFP and α-tubulin (loading control) in lysates of HUVECs transduced with shControl or shDLC1 3’UTR and rescued with GFP or GFP-DLC1. Note that the GFP-tagged DLC1 has a higher molecular weight than endogenous DLC1. Scans of whole Western blots are depicted in .

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Transduction, Two Tailed Test, Western Blot, Molecular Weight

Journal: bioRxiv

Article Title: DLC1 is a direct target of activated YAP/TAZ that drives collective migration and sprouting angiogenesis

doi: 10.1101/755389

Figure Lengend Snippet: Phase-contrast images of HUVECs transduced with shControl or shYAP1 and rescued with GFP or GFP-DLC1, in a scratch-wound assay (t=0, t=4, t=8 and t=12 hours after scratch). Yellow line highlights the unclosed wound area. Graphs showing the percentage of wound closure of HUVECs transduced with shControl or shYAP and rescued with GFP or GFP-DLC1 at three time-points during scratch-wound assay. Data is from 3 independent experiments, shControl (14 movies), shYAP (14 movies), rescue GFP (11 movies), rescue GFP-DLC1 (7 movies). n.s. nonsignificant, *P<0.05, ***P<0.001 (One-way ANOVA with Dunnett’s multiple comparisons test). (c) Western blot analysis of DLC1, YAP, GFP and □-actin in lysates of HUVECs transduced with shControl, shYAP and rescued with GFP or GFP-DLC1. Scans of whole Western blots are depicted in . (d) Representative widefield IF images of HUVECs transduced with shControl or shYAP, or shYAP-transduced HUVECs expressing GFP or GFP-DLC1, 6 hours after initiation of scratch wound assay. Stained for Vinculin (red) and F-actin (green). The asterisks indicate the GFP positive cells (not shown). (e) Representative phasecontrast images of sprouting spheroids 16 hours after VEGF stimulation of HUVECs transduced with shControl, shYAP and rescued with GFP or GFP-DLC1. Boxplots show the mean (± upper and lower quartiles) cumulative sprout length and number of sprouts per spheroid. Whiskers present min/max values. Data is from 3 independent experiments, shControl (27 spheroids), shYAP (18 spheroids), rescue GFP (21 spheroids), rescue GFP-DLC1 (28 spheroids). n.s.; not significant, *P<0.05, ***P<0.001 (One-way ANOVA with Dunnett’s multiple comparison test).

Article Snippet: Cells were transfected with the pGL3-DLC1-promotor luciferase reporter plasmids using PEI (Polysciences). pRL-TK Renilla reporter plasmid was co-transfected (1/50) as a control for transfection efficiency.

Techniques: Transduction, Scratch Wound Assay Assay, Western Blot, Expressing, Staining