Journal: Cell death & disease

Article Title: Endoplasmic reticulum stress related super-enhancers suppress cuproptosis via glycolysis reprogramming in lung adenocarcinoma.

doi: 10.1038/s41419-025-07613-0

Figure Lengend Snippet: Fig. 3 XBP1s protects LUAD cells from cell death induced by ES treatment. A WB analysis confirmed a significant upregulation of XBP1s expression in LUAD cell lines A549 and PC9 following ES treatment. B Box plot illustrates the distribution of H-scores for XBP1s expression in tumor tissue microarray from low- (n = 38) and high-grade (n = 39) LUAD. The H-scores in the high-grade group are significantly higher than those in the low-grade group. C Box plot illustrates the paired H-scores of XBP1s expression in tumor and adjacent normal tissues from a LUAD tissue microarray (n = 77), with statistical analysis performed using a paired t-test. D H-scores of XBP1s expression in tumor and adjacent normal tissues from the tissue microarray, stratified into low- (n = 38) and high-grade (n = 39) groups. Statistical analysis was performed using a paired t-test within each group. E Representative IHC images of XBP1s staining in low- and high-grade tissues. F After treating A549 cells with 40 nM and 100 nM ES for 72 hours, cell viability was measured. In the XBP1-overexpression (OE) group, viability was significantly higher compared to the empty vector (EV) control. Similarly, in the knockdown experiments, the negative control (NC) group exhibited significantly higher viability compared to the XBP1-knockdown (sh) group (n = 3). G Fold Viability of A549 cells after treatment with various concentration of ES. The OE group was able to maintain cell viability at higher ES concentrations, whereas the sh group exhibited increased sensitivity to ES in a dose-dependent manner. H Bar plot showing EdU positive ratio of A549 cells in four groups (OE, EV, sh, and NC) after treatment with 40 nM and 100 nM ES for 72 h (n = 3). I Representative fluorescence images of EdU staining. The OE group maintained significantly higher proliferation levels under ES treatment. J Representative images of tumor tissues from xenograft mice with OE or EV group under ES treatment or control conditions. Adjacent graph shows tumor growth curves and volume bar chart, highlighting the effect of XBP1s overexpression on tumor progression and cuproptosis resistance. (n = 6 per group). H High-resolution Transmission Electron Microscopy (TEM) images (×8.0k and ×25.0 k magnifications) were acquired using a Hitachi HT7700 microscope. The EV group exhibited a higher number of mitochondria compared to the OE group, but with abnormal morphology, such as loss of mitochondrial cristae and swelling. Despite these observations, no significant differences were detected in the statistical analysis (n = 3). The data are presented as the mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: Antibodies used in this study include: XBP1s (Cell Signaling Technology, #40435, 1:1000 for WB, 1:300 for IF, 1:50 for CUT&Tag); XBP1s (Merck, #MABC521, 1:400 for IHC); DLAT (Proteintech, # 68303-1-Ig, 1:400 for IHC and 1:1000 for WB); FDX1 (Proteintech, # 12592-1-AP, 1:400 for IHC); P300 (Cell Signaling Technology, #D2X6N, 1:300 for IF, 1:50 for CUT&Tag); MED1 (Thermo Fisher, #PA5-36114, 1:300 for IF, 1:50 for CUT&Tag); BRD4 (Cell Signaling Cell Death and Disease (2025) 16:316 Technology, #13440, 1:300 for IF, 1:50 for CUT&Tag); RNA Pol II-S2P (Millipore, cat# 04-1571, 1:300 for IF); RNA Pol II-S5P (Millipore, cat# 04-1572, 1:300 for IF); H3K4me3 (Cell Signaling Technology, cat# 9751, 1:300 for IF); H3K27ac (PTMbio, #PTM-160, 1:300 for IF, 1:50 for CUT&Tag); H3K4me1 (PTMbio, #PTM-5158, 1:300 for IF); MGRN1 (Proteintech, #11285-1-AP, 1:2000 for WB, 1:200 for IHC); β-actin (Proteintech, # 81115-1-RR, 1:10000 for WB); LIPT1 (Thermo Fisher, #PA5-106991, 1:1000 for WB and 1:200 for IHC); anti-DYKDDDDK tag (Proteintech, # 80010-1-RR, 1:5000 for WB); anti-HA tag (Proteintech, # 81290-1-RR, 1:5000 for WB); anti-His tag (Proteintech, #66005-1-Ig, 1:5000 for WB); HRP-conjugated goat anti-rabbit (Proteintech, # SA00001-2, 1:10000 for WB); HRP-conjugated goat anti-mouse (Proteintech, # SA00001-1, 1:10000 for WB); CoraLite® Plus 488-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM002, 1:1000 for IF); CoraLite® Plus 594-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM004, 1:1000 for IF); CoraLite® Plus 647-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM005, 1:1000 for IF) Tissue copper (Cu) colorimetric assay and intracellular copper probe staining Cu levels in LUAD tissues were detected using a Cu colorimetric assay kit (# E-BC-K300-M; Elabscience, Wuhan, China).

Techniques: Expressing, Microarray, Staining, Over Expression, Plasmid Preparation, Control, Knockdown, Negative Control, Concentration Assay, Transmission Assay, Electron Microscopy, Microscopy

Journal: Cell death & disease

Article Title: Endoplasmic reticulum stress related super-enhancers suppress cuproptosis via glycolysis reprogramming in lung adenocarcinoma.

doi: 10.1038/s41419-025-07613-0

Figure Lengend Snippet: Fig. 4 XBP1s undergoes phase separation in vitro and form condensates in the nucleus. A Domain structure and graphs of XBP1s IDRs based on VSL2 and IUPred algorithms. Scores >0.5 indicate disorder. Yellow shade depicts the designated core IDR (cIDR). B Representative fluorescence images (488 nm) of XBP1s droplets at varying protein concentrations (5 µM–30 µM) in a buffer containing 150 nM NaCl and 20% PEG-8000 (default condition unless otherwise specified). Both the size and number of droplets increased as protein concentration rose. Quantification of droplet area is shown on the right. C Representative fluorescence images (488 nm) of XBP1s-droplets at different temperatures. The droplet number peaked at 37 °C. Quantification of droplet area is shown on the right. D, E Representative fluorescence images (488 nm) of XBP1s-droplets at different NaCl concentrations (25–1000 nm). At 150 mM NaCl, droplet numbers were the highest, while at 200 mM NaCl, droplets were larger but fewer, indicating a concentration-dependent phase separation behavior. F Fluorescence recovery after photobleaching (FRAP) analysis of EGFP-XBP1s droplets in vitro. The graph illustrates the recovery of fluorescence intensity within 0–60 s, indicating the liquid-like dynamics of the droplets. G Live-cell imaging of A549 and PC9 cells, EGFP-XBP1s puncta were observed under treatment with buffers, both in the presence and absence of 5% 1,6-hexanediol. Nuclei were distinctly visualized through Hoechst staining. H Live-cell imaging showed the fusion phenomenon of XBP1s droplets was observed through live-cell imaging in A549 and PC9 cells, indicating the liquid-like dynamics of the droplets. I FRAP recovery of EGFP-XBP1s puncta in live A549 and PC9 cells. White squares mark photo-bleached puncta. The fluorescence recovery occurred within 2–30 s in A549 cells and 2–38 s in PC9 cells, further confirming the dynamic and liquid-like properties of XBP1s condensates. Quantification of fluorescence recovery is displayed on the right.

Article Snippet: Antibodies used in this study include: XBP1s (Cell Signaling Technology, #40435, 1:1000 for WB, 1:300 for IF, 1:50 for CUT&Tag); XBP1s (Merck, #MABC521, 1:400 for IHC); DLAT (Proteintech, # 68303-1-Ig, 1:400 for IHC and 1:1000 for WB); FDX1 (Proteintech, # 12592-1-AP, 1:400 for IHC); P300 (Cell Signaling Technology, #D2X6N, 1:300 for IF, 1:50 for CUT&Tag); MED1 (Thermo Fisher, #PA5-36114, 1:300 for IF, 1:50 for CUT&Tag); BRD4 (Cell Signaling Cell Death and Disease (2025) 16:316 Technology, #13440, 1:300 for IF, 1:50 for CUT&Tag); RNA Pol II-S2P (Millipore, cat# 04-1571, 1:300 for IF); RNA Pol II-S5P (Millipore, cat# 04-1572, 1:300 for IF); H3K4me3 (Cell Signaling Technology, cat# 9751, 1:300 for IF); H3K27ac (PTMbio, #PTM-160, 1:300 for IF, 1:50 for CUT&Tag); H3K4me1 (PTMbio, #PTM-5158, 1:300 for IF); MGRN1 (Proteintech, #11285-1-AP, 1:2000 for WB, 1:200 for IHC); β-actin (Proteintech, # 81115-1-RR, 1:10000 for WB); LIPT1 (Thermo Fisher, #PA5-106991, 1:1000 for WB and 1:200 for IHC); anti-DYKDDDDK tag (Proteintech, # 80010-1-RR, 1:5000 for WB); anti-HA tag (Proteintech, # 81290-1-RR, 1:5000 for WB); anti-His tag (Proteintech, #66005-1-Ig, 1:5000 for WB); HRP-conjugated goat anti-rabbit (Proteintech, # SA00001-2, 1:10000 for WB); HRP-conjugated goat anti-mouse (Proteintech, # SA00001-1, 1:10000 for WB); CoraLite® Plus 488-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM002, 1:1000 for IF); CoraLite® Plus 594-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM004, 1:1000 for IF); CoraLite® Plus 647-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM005, 1:1000 for IF) Tissue copper (Cu) colorimetric assay and intracellular copper probe staining Cu levels in LUAD tissues were detected using a Cu colorimetric assay kit (# E-BC-K300-M; Elabscience, Wuhan, China).

Techniques: In Vitro, Protein Concentration, Concentration Assay, Fluorescence, Live Cell Imaging, Staining

Journal: Cell death & disease

Article Title: Endoplasmic reticulum stress related super-enhancers suppress cuproptosis via glycolysis reprogramming in lung adenocarcinoma.

doi: 10.1038/s41419-025-07613-0

Figure Lengend Snippet: Fig. 5 XBP1s compartmentalizes SEs-related coactivators to nuclear puncta. Colocalization of transcriptional coactivators P300 (A), BRD4 (B), CDK9 (C), MED1 (D) with Flag-XBP1s (green) in nuclear puncta in A549 cells. Line scans of the colocalization images are depicted by white arrows with quantification (2D intensity histograms) shown to the right, highlighting Pearson’s correlation coefficients (R-values). Areas of colocalization are indicated in yellow. E Representative images of droplet formation of EGFP-XBP1s (488 nm) with mCherry-P300-IDR, mCherry- MED1-IDR, mCherry-BRD4-IDR or mCherry -CDK9-IDR (594 nm). The co-localization of EGFP-XBP1s (green) and mCherry-labeled IDR proteins (red) demonstrates their phase separation compatibility and mutual incorporation into condensates. Localization of active histone markers H3K27ac (F), H3K4me1 (G) and H3K4me3 (H), repressive histone markers H3K9me3 (I) and active RNA POI II S2P (J) and S5P (K) (red)with Flag- XBP1s (green) in A549 cells, which suggested that the condensates formed by XBP1s are associated with the transcriptional activation of target genes. Line scans of the colocalization images are depicted by white arrows with quantification (2D intensity histograms) shown to the right and below, highlighting Pearson’s correlation coefficients (R-values). Areas of colocalization are indicated in yellow.

Article Snippet: Antibodies used in this study include: XBP1s (Cell Signaling Technology, #40435, 1:1000 for WB, 1:300 for IF, 1:50 for CUT&Tag); XBP1s (Merck, #MABC521, 1:400 for IHC); DLAT (Proteintech, # 68303-1-Ig, 1:400 for IHC and 1:1000 for WB); FDX1 (Proteintech, # 12592-1-AP, 1:400 for IHC); P300 (Cell Signaling Technology, #D2X6N, 1:300 for IF, 1:50 for CUT&Tag); MED1 (Thermo Fisher, #PA5-36114, 1:300 for IF, 1:50 for CUT&Tag); BRD4 (Cell Signaling Cell Death and Disease (2025) 16:316 Technology, #13440, 1:300 for IF, 1:50 for CUT&Tag); RNA Pol II-S2P (Millipore, cat# 04-1571, 1:300 for IF); RNA Pol II-S5P (Millipore, cat# 04-1572, 1:300 for IF); H3K4me3 (Cell Signaling Technology, cat# 9751, 1:300 for IF); H3K27ac (PTMbio, #PTM-160, 1:300 for IF, 1:50 for CUT&Tag); H3K4me1 (PTMbio, #PTM-5158, 1:300 for IF); MGRN1 (Proteintech, #11285-1-AP, 1:2000 for WB, 1:200 for IHC); β-actin (Proteintech, # 81115-1-RR, 1:10000 for WB); LIPT1 (Thermo Fisher, #PA5-106991, 1:1000 for WB and 1:200 for IHC); anti-DYKDDDDK tag (Proteintech, # 80010-1-RR, 1:5000 for WB); anti-HA tag (Proteintech, # 81290-1-RR, 1:5000 for WB); anti-His tag (Proteintech, #66005-1-Ig, 1:5000 for WB); HRP-conjugated goat anti-rabbit (Proteintech, # SA00001-2, 1:10000 for WB); HRP-conjugated goat anti-mouse (Proteintech, # SA00001-1, 1:10000 for WB); CoraLite® Plus 488-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM002, 1:1000 for IF); CoraLite® Plus 594-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM004, 1:1000 for IF); CoraLite® Plus 647-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM005, 1:1000 for IF) Tissue copper (Cu) colorimetric assay and intracellular copper probe staining Cu levels in LUAD tissues were detected using a Cu colorimetric assay kit (# E-BC-K300-M; Elabscience, Wuhan, China).

Techniques: Labeling, Activation Assay

Journal: Cell death & disease

Article Title: Endoplasmic reticulum stress related super-enhancers suppress cuproptosis via glycolysis reprogramming in lung adenocarcinoma.

doi: 10.1038/s41419-025-07613-0

Figure Lengend Snippet: Fig. 7 XBP1s promotes LIPT1 ubiquitination and alters glycolysis and mitochondrial respiration. A Volcano plot showing differential protein expression profiles in A549 cells overexpressing XBP1s (XBP1s-OE) compared to the expression vector (EV) control after treatment with 40 nM ES. Red and green dots represent significantly upregulated and downregulated proteins, respectively (log2FC > 1.5, -log10 p- value > 1.3). Notably, LIPT1 is significantly downregulated. B KEGG pathway enrichment analysis of significantly altered proteins, highlighting activated glycolysis, ubiquitin-mediated proteolysis, and pyruvate metabolism pathways. C Western blot (WB) and quantitative PCR (qPCR) analyses of LIPT1 and XBP1s expression in LUAD cells with XBP1s overexpression (OE) compared to EV. LIPT1 protein levels are reduced in XBP1s-OE cells, while mRNA levels remain unchanged. β-actin was used as a loading control. D Immunoprecipitation (IP) assay demonstrating enhanced ubiquitination of LIPT1 in the presence of XBP1s in LUAD cells. Cells were treated with MG132 to inhibit proteasomal degradation. Flag-LIPT1, Myc-XBP1s, and HA-Ub were co-expressed, and ubiquitinated LIPT1 was detected using anti-HA antibodies. E Lactic acid and pyruvic acid levels in LUAD cells overexpressing LIPT1. Overexpression of LIPT1 reduces both metabolites (n = 3). F Lactic acid and pyruvic acid levels in LUAD cells with XBP1s overexpression (OE) or rescue of LIPT1 expression (Rescue). XBP1s-OE increases glycolysis metabolites, while LIPT1 rescue reduces their levels (n = 3). Data are shown as mean ± SD, *P < 0.05, **P < 0.01, ***P < 0.001, ***P < 0.0001, ns not significant. G Extracellular acidification rate (ECAR) measurements in LUAD cells. XBP1s-OE enhances glycolysis, while LIPT1 rescue reverses the effect. Shaded areas indicate glycolytic function after oligomycin addition. Data are presented as mean ± SD. H Oxygen consumption rate (OCR) analysis of mitochondrial respiration in LUAD cells. XBP1s-OE suppresses mitochondrial respiration, while LIPT1 rescue restores respiration levels. OCR values after sequential addition of oligomycin, FCCP, and antimycin A/rotenone are shown. Data are presented as mean ± SD.

Article Snippet: Antibodies used in this study include: XBP1s (Cell Signaling Technology, #40435, 1:1000 for WB, 1:300 for IF, 1:50 for CUT&Tag); XBP1s (Merck, #MABC521, 1:400 for IHC); DLAT (Proteintech, # 68303-1-Ig, 1:400 for IHC and 1:1000 for WB); FDX1 (Proteintech, # 12592-1-AP, 1:400 for IHC); P300 (Cell Signaling Technology, #D2X6N, 1:300 for IF, 1:50 for CUT&Tag); MED1 (Thermo Fisher, #PA5-36114, 1:300 for IF, 1:50 for CUT&Tag); BRD4 (Cell Signaling Cell Death and Disease (2025) 16:316 Technology, #13440, 1:300 for IF, 1:50 for CUT&Tag); RNA Pol II-S2P (Millipore, cat# 04-1571, 1:300 for IF); RNA Pol II-S5P (Millipore, cat# 04-1572, 1:300 for IF); H3K4me3 (Cell Signaling Technology, cat# 9751, 1:300 for IF); H3K27ac (PTMbio, #PTM-160, 1:300 for IF, 1:50 for CUT&Tag); H3K4me1 (PTMbio, #PTM-5158, 1:300 for IF); MGRN1 (Proteintech, #11285-1-AP, 1:2000 for WB, 1:200 for IHC); β-actin (Proteintech, # 81115-1-RR, 1:10000 for WB); LIPT1 (Thermo Fisher, #PA5-106991, 1:1000 for WB and 1:200 for IHC); anti-DYKDDDDK tag (Proteintech, # 80010-1-RR, 1:5000 for WB); anti-HA tag (Proteintech, # 81290-1-RR, 1:5000 for WB); anti-His tag (Proteintech, #66005-1-Ig, 1:5000 for WB); HRP-conjugated goat anti-rabbit (Proteintech, # SA00001-2, 1:10000 for WB); HRP-conjugated goat anti-mouse (Proteintech, # SA00001-1, 1:10000 for WB); CoraLite® Plus 488-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM002, 1:1000 for IF); CoraLite® Plus 594-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM004, 1:1000 for IF); CoraLite® Plus 647-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM005, 1:1000 for IF) Tissue copper (Cu) colorimetric assay and intracellular copper probe staining Cu levels in LUAD tissues were detected using a Cu colorimetric assay kit (# E-BC-K300-M; Elabscience, Wuhan, China).

Techniques: Ubiquitin Proteomics, Expressing, Plasmid Preparation, Control, Western Blot, Real-time Polymerase Chain Reaction, Over Expression, Immunoprecipitation

Journal: Cell death & disease

Article Title: Endoplasmic reticulum stress related super-enhancers suppress cuproptosis via glycolysis reprogramming in lung adenocarcinoma.

doi: 10.1038/s41419-025-07613-0

Figure Lengend Snippet: Fig. 8 Combination with SE inhibitors significantly enhanced the tumor-suppressing effect of ES in xenograft model. A Representative images of tumor samples from xenograft mice under different treatment conditions (Control, MGRN1-KD, ES+Control, ES + MGRN1-KD, ES +Control and ES + JQ1). B, C Quantitative analysis of tumor volume and weight. Tumor volume was measured every three days, and tumor weight was recorded at the endpoint. Data are presented as mean ± SD, with significant differences denoted by *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. D Immunohistochemical staining of XBP1s, MGRN1, and LIPT1 in tumor tissues, using previously described tumors (from Fig. 3J) and newly generated tumors in this experiment. These results validate the regulatory relationship among XBP1s, MGRN1, and LIPT1. E Ki67 staining of xenograft tumor tissues to evaluate the impact of XBP1s-related SEs on cell proliferation.

Article Snippet: Antibodies used in this study include: XBP1s (Cell Signaling Technology, #40435, 1:1000 for WB, 1:300 for IF, 1:50 for CUT&Tag); XBP1s (Merck, #MABC521, 1:400 for IHC); DLAT (Proteintech, # 68303-1-Ig, 1:400 for IHC and 1:1000 for WB); FDX1 (Proteintech, # 12592-1-AP, 1:400 for IHC); P300 (Cell Signaling Technology, #D2X6N, 1:300 for IF, 1:50 for CUT&Tag); MED1 (Thermo Fisher, #PA5-36114, 1:300 for IF, 1:50 for CUT&Tag); BRD4 (Cell Signaling Cell Death and Disease (2025) 16:316 Technology, #13440, 1:300 for IF, 1:50 for CUT&Tag); RNA Pol II-S2P (Millipore, cat# 04-1571, 1:300 for IF); RNA Pol II-S5P (Millipore, cat# 04-1572, 1:300 for IF); H3K4me3 (Cell Signaling Technology, cat# 9751, 1:300 for IF); H3K27ac (PTMbio, #PTM-160, 1:300 for IF, 1:50 for CUT&Tag); H3K4me1 (PTMbio, #PTM-5158, 1:300 for IF); MGRN1 (Proteintech, #11285-1-AP, 1:2000 for WB, 1:200 for IHC); β-actin (Proteintech, # 81115-1-RR, 1:10000 for WB); LIPT1 (Thermo Fisher, #PA5-106991, 1:1000 for WB and 1:200 for IHC); anti-DYKDDDDK tag (Proteintech, # 80010-1-RR, 1:5000 for WB); anti-HA tag (Proteintech, # 81290-1-RR, 1:5000 for WB); anti-His tag (Proteintech, #66005-1-Ig, 1:5000 for WB); HRP-conjugated goat anti-rabbit (Proteintech, # SA00001-2, 1:10000 for WB); HRP-conjugated goat anti-mouse (Proteintech, # SA00001-1, 1:10000 for WB); CoraLite® Plus 488-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM002, 1:1000 for IF); CoraLite® Plus 594-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM004, 1:1000 for IF); CoraLite® Plus 647-Goat Anti-Mouse Recombinant Secondary Antibody (H+ L) (Proteintech, # RGAM005, 1:1000 for IF) Tissue copper (Cu) colorimetric assay and intracellular copper probe staining Cu levels in LUAD tissues were detected using a Cu colorimetric assay kit (# E-BC-K300-M; Elabscience, Wuhan, China).

Techniques: Control, Immunohistochemical staining, Staining, Generated

Journal: Journal of Virology

Article Title: Pharmacologic Activation of Lytic Epstein-Barr Virus Gene Expression without Virion Production

doi: 10.1128/JVI.00998-19

Figure Lengend Snippet: Clofoctol induces the UPR in a Burkitt lymphoma cell line. (A) Schematic representation of UPR signaling pathways. ATF6(N), an N-terminal fragment of ATF6. (B) qRT-PCR assays of the UPR markers. BX1-Akata cells were treated with 20 μM clofoctol, RNA was isolated at the indicated time points, and reverse-transcribed cDNA was used for CHOP, BiP, and XBP1s RNA quantification. (C) Immunoblots of the UPR markers. BX1-Akata cells were treated with 20 μM clofoctol, proteins were extracted at the indicated time points, and immunoblotting was performed with antibodies against ATF4, XBP1s, ATF6, and actin. (D) Immunoblot of an assay with two different antibodies showing expression of ubiquitinated (Ub) proteins 6 h after the treatment of BX1-Akata cells with clofoctol and bortezomib.

Article Snippet: Clofoctol, anti-IgG, an XBP1s inhibitor (4u), a PERK inhibitor (GSK2606414), and an EIF2AK3 activator (CCT020312) were purchased from MilliporeSigma; bortezomib was from Millennium Pharmaceuticals.

Techniques: Quantitative RT-PCR, Isolation, Western Blot, Expressing

Journal: Journal of Virology

Article Title: Pharmacologic Activation of Lytic Epstein-Barr Virus Gene Expression without Virion Production

doi: 10.1128/JVI.00998-19

Figure Lengend Snippet: PERK and IRE1-XBP1s pathways mediate clofoctol-induced lytic activation. (A and B) BX1-Akata cells were transduced with a pool of lentiviral particles containing 3 different shRNA constructs targeting PERK or a scrambled control lentiviral particle and selected with puromycin. (A) qRT-PCR was performed to confirm PERK knockdown and to measure Trb3 and Zta RNA levels after 24 h of clofoctol treatment. (B) Fluorescence microscopy was used to detect GFP-positive cells after 24 h of clofoctol treatment, and the cell counts were compared to those after no treatment (control [ctrl]). (C and D) BX1-Akata cells were pretreated with a PERK inhibitor (Pi), with an XBP1s inhibitor (4u), or with both (Pi + 4u), which was followed by clofoctol (CLF) treatment. (C) qRT-PCR for Zta, BMRF1, and gp350 RNA quantification was performed 24 h after treatment. (D) Fluorescence microscopy was used to detect GFP-positive cells at 24 h after treatment, and cell counts were compared to those for the control.

Article Snippet: Clofoctol, anti-IgG, an XBP1s inhibitor (4u), a PERK inhibitor (GSK2606414), and an EIF2AK3 activator (CCT020312) were purchased from MilliporeSigma; bortezomib was from Millennium Pharmaceuticals.

Techniques: Activation Assay, Transduction, shRNA, Construct, Quantitative RT-PCR, Fluorescence, Microscopy

Journal: Cell Reports Medicine

Article Title: Ablation of ERO1A induces lethal endoplasmic reticulum stress responses and immunogenic cell death to activate anti-tumor immunity

doi: 10.1016/j.xcrm.2023.101206

Figure Lengend Snippet:

Article Snippet: Primary antibodies used included: anti-ERO1L (Abcam, Cat# ab177156), anti-IRE1α (Abcam, Cat# ab37073), anti-pIRE1α (Abcam, Cat# ab48187), anti-XBP1s (Cell Signaling Technology, Cat# 12782), anti-PERK (Abcam, Cat# ab229912), anti-pPERK (Cell Signaling Technology, Cat# 3179), anti-eIF2α (Cell Signaling Technology, Cat# 9722), anti-peIF2α (Cell Signaling Technology, Cat# 9721), anti-ATF4 (Cell Signaling Technology, Cat# 11815), anti-ATF6α (Santa Cruz, Cat# sc-166659), anti-CHOP (Santa Cruz, Cat# sc-7351), and anti-β-Actin.

Techniques: Control, Purification, Virus, Recombinant, CCK-8 Assay, Transfection, Protease Inhibitor, SYBR Green Assay, Saline, Lysis, Multiplex Assay, Cytotoxicity Assay, Cell Isolation, Enzyme-linked Immunosorbent Assay, Bicinchoninic Acid Protein Assay, Software

Journal: iScience

Article Title: Cetylpyridinium chloride triggers paraptosis to suppress pancreatic tumor growth via the ERN1-MAP3K5-p38 pathway

doi: 10.1016/j.isci.2024.110598

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-XBP1s , Proteintech Biotechnology , Cat# 24868-1-AP; RRID: AB_2879766.

Techniques: Recombinant, Lysis, Modification, Polyacrylamide Gel Electrophoresis, Protease Inhibitor, CCK-8 Assay, Sequencing, Software