Journal: The Journal of Biological Chemistry

Article Title: Ganglioside GM2 induces epithelial–mesenchymal transition (EMT) in cancer cells in a MEK/ERK/Egr1-dependent transcriptional program

doi: 10.1016/j.jbc.2026.111306

Figure Lengend Snippet: Ganglioside GM2 induces the expression of ERK-target genes. A , schematic representation of RNA sequencing in the presence or absence of exogenous GM2. B , heatmap analysis of differentially regulated genes ( p value ≤0.05) analyzed by RNA sequencing in GM2 treated versus nontreated HeLa cells for 1 h. The heatmap represents upregulation ( red ) and downregulation ( blue ) of gene expression in GM2 treated versus nontreated HeLa cells. ERK target gene expression is shown in the presence of GM2 and non-GM2 condition in the heatmap where red and blue color denotes the upregulation and downregulation respectively ( B , right side panel). C , volcano plot shows differentially expressed genes identified from RNA-sequencing. Significantly upregulated ERK target genes are marked in the volcano plot. Fold change values of the four upregulated genes are listed in right side box ( C , right panel). Quantitative real time PCR data reflect the upregulation of the ERK target genes ( i.e. , Egr1 , Nr4a1 , FosB , and Dusp5 ) in cervical cancer cell line, HeLa ( D ), breast cancer cell line, MCF7 ( E ) and kidney cancer cell line, SK-RC-45 ( F ) in the presence of exogenous GM2 (25 μM concentration) for 1 h. All mRNA quantification data are normalized against GAPDH and represented as relative fold changes with respect to untreated control. Data represent mean ± SEM of three independent experiments (Student's t test, two-tailed, paired); ∗∗ p < 0.01, ∗∗∗ p < 0.001. Western blot analysis shows GM2-induced upregulation of Egr1, FosB, and Nr4a1 protein expression in HeLa ( G ), MCF7 ( H ) and SK-RC-45 ( I ) cell lines. ERK, extracellular signal-regulated kinase; Egr1, early growth response protein 1; Nr4a1, nuclear receptor subfamily 4, group A, member 1; Dusp5, dual specificity phosphatase 5.

Article Snippet: FosB (#2251), Ctgf (#86641S), total ERK (#4695S), GAPDH (#97166) pMEK (#9154), N-cadherin (#14215), and Vimentin (#5741) were purchased from Cell Signaling Technology and Hsp90 (#BB-AB012), β-actin (#BB-AB0024) from BioBharati Life Science Pvt Ltd. Adam10 (#ab124695) primary antibody was purchased from Abcam.

Techniques: Expressing, RNA Sequencing, Gene Expression, Targeted Gene Expression, Real-time Polymerase Chain Reaction, Concentration Assay, Control, Two Tailed Test, Western Blot

Journal: The Journal of Biological Chemistry

Article Title: Ganglioside GM2 induces epithelial–mesenchymal transition (EMT) in cancer cells in a MEK/ERK/Egr1-dependent transcriptional program

doi: 10.1016/j.jbc.2026.111306

Figure Lengend Snippet: Time-dependent induction of ERK-target gene expression results in functional translocation of Egr1 to the nucleus. A , time dependent relative fold change values of Egr1 , Nr4a1 , FosB , and Dusp5 mRNA levels are estimated by RT-PCR at different time points in the presence of exogenous GM2 in HeLa cells. All mRNA quantification data are normalized against 7SL and represented as relative fold changes with respect to untreated control. Data represent mean ± SEM of at least two independent experiments [Student's t test (two-tailed; paired); ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001]. B , Time-dependent expression of Egr1, Nr4a1, and FosB protein levels are detected by western immunoblotting at different time points in the presence of GM2 in HeLa cells. C , densitometric quantification of Egr1, FosB, and NR4A1 proteins are represented as bar graphs on the right side. Band intensities are quantified using ImageJ software and normalized to β-actin (loading control). Data represent mean ± SEM of at least two independent experiments [Student's t test (one-tailed; paired); ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001]. D , confocal imaging shows GM2-mediated expression and nuclear localization of Egr1 ( red ) protein time dependently in the presence of exogenous GM2 in HeLa cells. Nucleus is stained with DAPI ( blue ). The scale bar represents 50 μm and applies uniformly to all fields shown. E , Confocal microscopy demonstrates the comparative expression of GM2 ( green ) in GM2 overexpressing murine kidney cancer cell line Renca V versus its parental cell line Renca, a low GM2 expressing cell line. Nucleus is stained with DAPI ( blue ). The scale bar represents 50 μm and applies uniformly to all fields shown. F , western blot data show higher expression of Egr1, Nr4a1, FosB, and Dusp5 protein in Renca V versus Renca cell line and G, bar graphs show the densitometric quantification data of Egr1, Nr4a1, FosB, and Dusp5 protein in Renca versus Renca V cell line. Band intensities are quantified using ImageJ software and normalized to β-actin (loading control). Data represent mean ± SEM of at least two independent experiments [Student's t test (one-tailed; unpaired); ∗ p < 0.05, ∗∗ p < 0.01. ERK, extracellular signal-regulated kinase; Egr1, early growth response protein 1; RT-PCR, real-time PCR; DAPI, 4′,6-diamidino-2-phenylindole; Nr4a1, nuclear receptor subfamily 4, group A, member 1.

Article Snippet: FosB (#2251), Ctgf (#86641S), total ERK (#4695S), GAPDH (#97166) pMEK (#9154), N-cadherin (#14215), and Vimentin (#5741) were purchased from Cell Signaling Technology and Hsp90 (#BB-AB012), β-actin (#BB-AB0024) from BioBharati Life Science Pvt Ltd. Adam10 (#ab124695) primary antibody was purchased from Abcam.

Techniques: Targeted Gene Expression, Functional Assay, Translocation Assay, Reverse Transcription Polymerase Chain Reaction, Control, Two Tailed Test, Expressing, Western Blot, Software, One-tailed Test, Imaging, Staining, Confocal Microscopy, Real-time Polymerase Chain Reaction

Journal: The Journal of Biological Chemistry

Article Title: Ganglioside GM2 induces epithelial–mesenchymal transition (EMT) in cancer cells in a MEK/ERK/Egr1-dependent transcriptional program

doi: 10.1016/j.jbc.2026.111306

Figure Lengend Snippet: GM2-mediated ERK-target gene expression is MEK-dependent. A , schematic representation depicting the strategy to inhibit GM2-mediated ERK-target gene expression using the MEK inhibitor U0126. Western data depict the protein levels of Egr1 and FosB in ( B ) HeLa and Egr1, FosB, and Nr4a1 in ( C ) MCF7 cell line using MEK inhibitor U0126 (pretreated for 2 h with 10 μM concentration) in the presence or absence of exogenous GM2 (1 h treatment with 25 μM concentration). Quantitative RT-PCR data reflect GM2-induced expression of Egr1 , Nr4a1 , FosB , and Dusp5 in the presence of the MEK inhibitor U0126 at the mRNA level in ( D ) HeLa and ( E ) MCF7 cell lines. All mRNA quantification data are normalized against 7SL and represented as relative fold changes with respect to untreated control. Data represent mean ± SEM of three independent experiments [Student's t test, two-tailed; paired); ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns = nonsignificant]. F , imaging by confocal microscopy of GM2-mediated Egr1 protein (red ) expression and nuclear localization in the presence of U0126. Nucleus is stained with DAPI ( blue ). The scale bar represents 50 μm and applies uniformly to all fields shown. SFM, serum-free media; ERK, extracellular signal-regulated kinase; Egr1, early growth response protein 1; RT-PCR, real-time PCR; DAPI, 4′,6-diamidino-2-phenylindole; Nr4a1, nuclear receptor subfamily 4, group A, member 1.

Article Snippet: FosB (#2251), Ctgf (#86641S), total ERK (#4695S), GAPDH (#97166) pMEK (#9154), N-cadherin (#14215), and Vimentin (#5741) were purchased from Cell Signaling Technology and Hsp90 (#BB-AB012), β-actin (#BB-AB0024) from BioBharati Life Science Pvt Ltd. Adam10 (#ab124695) primary antibody was purchased from Abcam.

Techniques: Targeted Gene Expression, Western Blot, Concentration Assay, Quantitative RT-PCR, Expressing, Control, Two Tailed Test, Imaging, Confocal Microscopy, Staining, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction

Journal: The Journal of Clinical Investigation

Article Title: Chloride homeostasis dysfunction drives hyperactivation of corticotropin-releasing factor-expressing neurons in the amygdala in stress-induced hypertension

doi: 10.1172/JCI195536

Figure Lengend Snippet: ( A and B ) Changes in MAP ( A ) and HR ( B ) measured by radiotelemetry before, during and after CUMS in WKY rats, BHRs and age-matched unstressed WKY rats and BHRs ( n = 6 rats in each group). Data are expressed as means ± SEM. * P < 0.05 for CUMS BHR versus unstressed BHRs, # P < 0.05 for CUMS BHR versus CUMS WKY, ^ P < 0.05 for CUMS WKY versus unstressed WKY. Repeated measures of 2-way ANOVA with Tukey’s post hoc test. ( C ) Representative immunofluorescence images showing CRF immunopositive neurons (green), delta-FosB immunopositive neurons (red), and CRF and delta-FosB double-positive neurons in the CeA of WKY, CUMS WKY, BHR, and CUMS BHR group of rats ( n = 5 rats in each group). ( D and E ) Summary data showing the numbers of CRF-positive neurons ( D ) and delta-FosB positive neurons ( E ) in the CeA of these 4 groups. ( F ) The percentage of CRF and delta-FosB double-positive neurons in the CeA of these 4 groups ( n = 5 rats in each group). Data are expressed as means ± SEM. ** P < 0.01, *** P < 0.001. One-way ANOVA with Tukey’s multiple comparison tests.

Article Snippet: Subsequently, the sections were incubated with a mouse anti-delta-FosB antibody (1:50, #sc-398595), a rabbit anti-CRF antibody (1:100, #A1122, Abclonal) or a rabbit anti-NKCC1 antibody (1:100, #13884, Proteintech) overnight at 4°C.

Techniques: Immunofluorescence, Comparison