Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Chlamydia trachomatis induces the transcriptional activity of host YAP in a Hippo-independent fashion

doi: 10.3389/fcimb.2023.1098420

Figure Lengend Snippet: Chlamydia infection induces expression of a subset of YAP target genes. (A) Volcano plot of gene expression in bulk RNA-sequencing of End1/E6E7 immortalized epithelial cells (End1s) during infection with Chlamydia trachomatis ( Ct ) serovar L2 compared to mock infection, at 24 hours post-infection (hpi). n = 3, with a minimum of 3x10 7 unstranded single reads per replicate with a mean length of 150 bp. All fold changes are relative to the mock-infected control; red dots: false discovery rate p-value (FDRP) ≤ 0.05, labels: top 20 genes whose expression differed most significantly (lowest FDRP) from the mock infection. (B) Table of selected transcription factors identified as potential targets of infection-associated modulation via cross-referencing of differentially expressed genes identified in (A) with the ChIP Enrichment Analysis (ChEA) database of transcription factor target genes. See also Supplementary Data S2. (C) Volcano plot of gene expression in bulk RNA-sequencing of primary human endocervical epithelial cells (HCECs) during infection with Ct serovar L2 compared to mock infection, at 24 hours post-infection (hpi). n = 3, with a minimum of 3x10 7 unstranded single reads per replicate with a mean length of 150 bp. All fold changes are relative to the mock-infected control; red dots: false discovery rate p-value (FDRP) ≤ 0.05, labels: top 20 genes whose expression differed most significantly (lowest FDRP) from the mock infection. (D) Venn diagram of differentially expressed genes identified in (A, C) cross-referenced with the ChEA database of YAP target genes. (E) Scatter plot of gene expression of YAP-responsive (ChEA), differentially expressed genes in either Ct serovar L2-infected End1s (x-axis) or HCECs (y-axis). All fold changes are relative to each cell type’s respective mock-infected control; blue line: linear regression model of correlation; grey shading: 95% confidence interval. R 2 and p-values calculated using Pearson’s correlation. (F) Heatmap of YAP target gene expression in Ct serovar L2-infected End1s (left columns) and HCECs (right columns). All fold changes are relative to each cell type’s respective mock-infected control; only genes differentially expressed (FDRP ≤ 0.05) in both cell types are shown. (G) Dot plot of GO biological process term enrichment in the set of YAP target genes differentially expressed in Ct serovar L2 infection of End1s or HCECs (top 25 most significantly enriched terms shown). Dot size: number of term-associated genes found in set, dot color: adjusted p-value.

Article Snippet: Primary human cervical epithelial cells (HCECs, ATCC PCS-0480-011, Lot 80306190) were cultured at 37° C with 5% atmospheric CO 2 in Cervical Epithelial Cell Basal Medium (CECBM, ATCC PCS-480-032) supplemented with all contents of a Cervical Epithelial Growth Kit (ATCC PCS-080-042).

Techniques: Infection, Expressing, Gene Expression, RNA Sequencing, Control, Targeted Gene Expression

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Chlamydia trachomatis induces the transcriptional activity of host YAP in a Hippo-independent fashion

doi: 10.3389/fcimb.2023.1098420

Figure Lengend Snippet: Chlamydia infection promotes YAP nuclear translocation. (A) Expression of CTGF and CYR61 at 24 hpi in mock- and Ct L2-infected End1 cells, as measured by RT-qPCR. n = 3 biological replicates; fold changes are relative to mean expression of the mock-infected and untreated control. Whiskers: minimum to maximum; n.s.: not significant (p-values > 0.05), asterisks: p-values ≤ 0.05, using pairwise Student’s t-tests and Bonferroni’s correction for multiple comparisons. (B) Expression of CTGF at 24 hpi in mock- and Ct L2-infected End1 cells transfected with non-targeting (NT) or YAP1-targeting siRNA (10 nM for 24 h prior to infection), as measured by RT-qPCR. n = 3 biological replicates; fold changes are relative to mean expression of the mock-infected and untreated control. Whiskers: minimum to maximum; asterisks: p-values ≤ 0.05, using pairwise Student’s t-tests and Bonferroni’s correction for multiple comparisons. (C) Expression of CTGF, INHBA, and BMP2 at 24 hpi in mock- and Ct L2-infected End1 cells treated with the YAP-TEAD inhibitor verteporfin (Vpf, 5 μM for 16 h starting at 8 hpi), as measured by RT-qPCR. n = 5 biological replicates; fold changes are relative to mean expression of the mock-infected and untreated control. Whiskers: minimum to maximum; asterisks: p-values ≤ 0.05, using pairwise Student’s t-tests and Bonferroni’s correction for multiple comparisons. (D) Representative micrographs of YAP (green) translocation into the nuclei (blue) of confluent mock- and Ct L2-infected End1 cells at 24 hpi. Asterisks: chlamydial inclusions, scale bar: 20 μm. (E) Quantification of YAP nuclear translocation in (D) as a ratio of nuclear to cytosolic YAP fluorescence. n = 5 biological replicates, 50 cells measured per sample. Black bars: group means; asterisks: p-values ≤ 0.05, using pairwise Wilcoxon rank sum tests and Bonferroni’s correction for multiple comparisons. (F) Representative micrographs of YAP nuclear translocation of confluent mock- and Ct L2-infected primary human cervical epithelial cells at 24 hpi. Asterisks, chlamydial inclusions, scale bar: 20 μm. (G) Quantification of YAP nuclear translocation in (F) as a ratio of nuclear to cytosolic YAP fluorescence. n = 5 biological replicates, 50 cells measured per sample. Black bars: group means; asterisks: p-values ≤ 0.05, using pairwise Wilcoxon rank sum tests and Bonferroni’s correction for multiple comparisons. (H) Quantification of YAP nuclear translocation at 2, 4, 8, 12, 18, and 24 hpi in confluent mock- and Ct L2-infected End1 cells as a ratio of nuclear to cytosolic YAP fluorescence. n = 5 biological replicates, 50 cells measured per sample. Blue dots: mock-infected cells, red dots: Ct L2-infected cells, black bars: group means, asterisks: p-value ≤ 0.05, using pairwise Wilcoxon rank-sum tests and Bonferroni’s correction for multiple comparisons. (I) Representative micrographs of YAP nuclear translocation at 18 hpi in confluent mock- and Ct L2-infected End1 cells treated with chloramphenicol (Cm, 50 μg/mL for 1 h at 17 hpi) or DMSO. Asterisks: chlamydial inclusions; scale bar: 20 μm. (J) Quantification of YAP nuclear translocation in (I) . n = 3 biological replicates, 50 cells measured per sample. Black bars: group means; asterisks: p-values ≤ 0.05, using pairwise Wilcoxon rank-sum tests and Bonferroni’s correction for multiple comparisons.

Article Snippet: Primary human cervical epithelial cells (HCECs, ATCC PCS-0480-011, Lot 80306190) were cultured at 37° C with 5% atmospheric CO 2 in Cervical Epithelial Cell Basal Medium (CECBM, ATCC PCS-480-032) supplemented with all contents of a Cervical Epithelial Growth Kit (ATCC PCS-080-042).

Techniques: Infection, Translocation Assay, Expressing, Quantitative RT-PCR, Control, Transfection, Fluorescence

Journal: BMJ Open

Article Title: Detection of Chlamydia trachomatis and Neisseria gonorrhoeae in an STI population: performances of the Presto CT-NG assay, the Lightmix Kit 480 HT CT/NG and the COBAS Amplicor with urine specimens and urethral/cervicovaginal samples

doi: 10.1136/bmjopen-2013-003607

Figure Lengend Snippet: Sensitivity, specificity, PPV and NPV for the three assays for Chlamydia trachomatis detection

Article Snippet: Newly developed dual detection systems for C trachomatis and N gonorrhoeae are implemented in Europe in the past 2 years including the Presto CT-NG assay (Goffin Molecular Technologies, Houten, the Netherlands) and the Lightmix Kit 480 HT CT/NG (TIB MOLBIOL, Berlin, Germany).

Techniques:

Journal: Cancer Genomics & Proteomics

Article Title: RFC3 Knockdown Decreases Cervical Cancer Cell Proliferation, Migration and Invasion

doi: 10.21873/cgp.20493

Figure Lengend Snippet: Differential expression of RFC3 in cervical cancer tissues and cell lines. (A) RFC3 mRNA expression in cervical cancer tissues (n=306) and normal tissues (n=13), in addition to overall survival and disease-free survival were analyzed using the GEPIA2 web tool. (B) RFC3 mRNA expression in cervical cancer cell lines and human primary cervical epithelial cell was assessed using RT-qPCR analysis. The experiment was performed three times, and significant differences were determined by mean±SD. **p<0.01, ***p<0.001. (C) RFC3 protein expression was detected by western blotting in the same cell lines. Cell lysates were immunoblotted with an anti-RFC3 antibody, and an anti-β-actin antibody was used as a loading control.

Article Snippet: Human primary cervical epithelial cells were cultured in cervical epithelial cell basal medium added the contents of cervical epithelial cell growth kit (ATCC).

Techniques: Quantitative Proteomics, Expressing, Quantitative RT-PCR, Western Blot, Control