Journal: bioRxiv

Article Title: Cardiovascular Disease-Associated Non-Coding Variants Disrupt GATA4-DNA Binding and Regulatory Functions

doi: 10.1101/2024.09.19.613959

Figure Lengend Snippet: Expression and purification of GATA4 ZF. A) SDS-PAGE (left) and western blot (right) of GATA4 ZF (18.8 kDa). Abbreviations: wash fractions (W), eluted fractions (E), concentrated fractions (C), un-induced protein sample (U), IPTG-induced protein sample (I), supernatant fraction (S), and pellet fraction (P). B) Western Blot of full-length GATA4 expressed in HeLA cells (48.6 kDa).

Article Snippet: HeLa Cervical Adenocarcinoma Human (HeLa) cells (ATCC–CCL-2) were grown in Eagle’s Minimum Essential Medium (EMEM) (ATCC - 30-2003) with 10% Fetal Bovine Serum (FBS) at 37 °C and 5% CO 2 .

Techniques: Expressing, Purification, SDS Page, Western Blot

Journal: bioRxiv

Article Title: Cardiovascular Disease-Associated Non-Coding Variants Disrupt GATA4-DNA Binding and Regulatory Functions

doi: 10.1101/2024.09.19.613959

Figure Lengend Snippet: CVD-associated SNPs alter gene expression and are in eQTL in cardiac tissue. A) Relative luciferase activity in HeLa cells transfected with reporter plasmids containing reference (blue with black circles) and alternate (orange with black squares) of variants rs1506537, rs56992000, rs2941506, and rs2301249. B) Cardiac tissue eQTL analysis of MRPL33 , TOP2B , PGAP3 , and CSK expressed in heart atrial appendage or left ventricle when rs1506537, rs56992000, rs2941506, and rs2301249 occur, respectively. C) University of California Santa Cruz (UCSC) Genome Browser tracks of variants rs2941506 and rs56992000 near TAD boundaries and regulatory elements.

Article Snippet: HeLa Cervical Adenocarcinoma Human (HeLa) cells (ATCC–CCL-2) were grown in Eagle’s Minimum Essential Medium (EMEM) (ATCC - 30-2003) with 10% Fetal Bovine Serum (FBS) at 37 °C and 5% CO 2 .

Techniques: Gene Expression, Luciferase, Activity Assay, Transfection

Journal: bioRxiv

Article Title: The p53 Protein is a Suppressor of Atox1 Copper Chaperon in Tumor Cells Under Genotoxic Effects

doi: 10.1101/2023.07.25.550476

Figure Lengend Snippet: Dependence of Atox1 and p53 level in HCT116 and A549 cell lines with different TP53 status: A - immunoblotting with antibodies to p53, p21, and Atox1; beta-actin was used as a normalization. A densitometric analysis of the obtained data is shown below. B – RT-qPCR analysis with primers for TP53, CDKN1A, and ATOX1 genes; GAPDH gene was used as a reference. C - immunofluorescence staining with primary antibodies to Atox1, secondary antibodies with AlexaFluor488. DAPI was used for nuclei staining. WT – wild type cells, TP53 -/- – cells without TP53. For all experiments: n = 3, mean +/− SEM, paired Student t-test, p < 0,05.

Article Snippet: Transformed human cell lines used: HCT116 (colon adenocarcinoma) with intact p53; HCT116p53 -/- with a deletion of both alleles of the TP53 genes, as well as the A549 line with wild (A549) and knockout p53 (A549p53 -/- ) by the CRISPR-Cas9 method, acquired at ATCC.

Techniques: Western Blot, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: bioRxiv

Article Title: The p53 Protein is a Suppressor of Atox1 Copper Chaperon in Tumor Cells Under Genotoxic Effects

doi: 10.1101/2023.07.25.550476

Figure Lengend Snippet: Influence of cytotoxic agents on the activity of Atox1 at different status (WT and KO) of the TP53 gene in A549 and HCT116 cell lines, 24h after drugs exposure. A - immunoblotting with antibodies to p53, p21, and Atox1; beta-actin was used as a normalization. A densitometric analysis of the obtained data is shown below. B – RT-qPCR analysis with primers for TP53, CDKN1A and ATOX1 genes; GAPDH gene was used as a reference, C - immunofluorescence staining with primary antibodies to Atox1, secondary antibodies with AlexaFluor488. DAPI was used for nuclei staining. DOX – doxorubicin (0,1μM), CIS – cisplatin (35μM), PMA – phorbol-12-myristate- 13-acetate (80nM), H 2 O 2 – hydrogen peroxide (450μM), BLE – bleomycin (10μM). WT – wild type cells, TP53 -/- – cells without TP53. For all experiments: n = 3, mean +/− SEM, two-way ANOVA, p < 0,05.

Article Snippet: Transformed human cell lines used: HCT116 (colon adenocarcinoma) with intact p53; HCT116p53 -/- with a deletion of both alleles of the TP53 genes, as well as the A549 line with wild (A549) and knockout p53 (A549p53 -/- ) by the CRISPR-Cas9 method, acquired at ATCC.

Techniques: Activity Assay, Western Blot, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: bioRxiv

Article Title: The p53 Protein is a Suppressor of Atox1 Copper Chaperon in Tumor Cells Under Genotoxic Effects

doi: 10.1101/2023.07.25.550476

Figure Lengend Snippet: Influence of ionizing radiation on the activity of Atox1 at different status (WT and KO) of the TP53 gene in A549 and HCT116 cell lines, 24h after ionizing irradiation (10Gy) exposure. A - immunoblotting with antibodies to p53, p21, and Atox1; beta-actin was used as a normalization. A densitometric analysis of the obtained data is shown below. B – RT-qPCR analysis with primers for TP53, CDKN1A and ATOX1 genes; GAPDH gene was used as a reference. The value of WT 0Gy (control) was taken as a 1.0 for all genes and is not shown in the graphs. For all experiments: n = 3, mean +/− SEM, two-way ANOVA, p < 0,05.

Article Snippet: Transformed human cell lines used: HCT116 (colon adenocarcinoma) with intact p53; HCT116p53 -/- with a deletion of both alleles of the TP53 genes, as well as the A549 line with wild (A549) and knockout p53 (A549p53 -/- ) by the CRISPR-Cas9 method, acquired at ATCC.

Techniques: Activity Assay, Irradiation, Western Blot, Quantitative RT-PCR, Control