p ire1α  (Novus Biologicals)

Journal: The Journal of Biological Chemistry

Article Title: Brucella Omp25 activates the unfolded protein response to promote intracellular proliferation and inflammation

doi: 10.1016/j.jbc.2026.111333

Figure Lengend Snippet: Brucella abortus Omp25 activates UPR signaling. A , screening of Brucella proteins for UPR activation. HEK293T cells (5 × 10 5 ) were transfected with empty vector or Brucella protein expression plasmids for 24 h before RT–qPCR analysis. B , effects of B. abortus Omp25 on transcription of UPR downstream genes. HEK293T cells (5 × 10 5 ) were transfected with increasing amounts of Omp25-FLAG plasmid (0, 75, 150, or 300 ng) for 24 h before RT–qPCR analysis. C , effects of Omp25 on XBP1 mRNA splicing. Total RNA was analyzed by RT–PCR, and XBP1s band intensities were quantified relative to GAPDH. D , effects of Omp25 on activation of UPR reporters. HEK293T cells (1 × 10 5 ) were cotransfected with Omp25-FLAG (50 ng), pRL-TK (20 ng), and luciferase reporters for ATF6 (20 ng), ATF4 (20 ng), or XBP1s (10 ng). Luciferase activity was measured 24 h post-transfection. E and F , Omp25 activates PERK, IRE1α, and ATF6 pathways. HEK293T cells (5 × 10 5 ) were transfected with increasing doses of Omp25-FLAG (0, 0.3, 0.9, and 2.7 μg) for 24 h ( E ) or with 2 μg Omp25-FLAG for 24, 36, and 48 h ( F ) before Western blotting with the indicated antibodies. Band intensities were quantified relative to β-actin. Data are presented as mean ± SD from n = 3 to 4 independent samples. Statistical significance was determined using one-way ANOVA ( A , B ) or unpaired t test ( D ). ∗ p < 0.05, ∗∗ p < 0.01. ATF, activating transcription factor; HEK293T, human embryonic kidney 293T cell line; IRE1α, inositol-requiring enzyme 1 alpha; Omp, outer membrane protein; PERK, PKR-like ER kinase; qPCR, quantitative PCR; UPR, unfolded protein response; XBP1, X-box binding protein 1; XBP1s, the spliced form of XBP1.

Article Snippet: Mouse monoclonal antibodies against HA (66006, Proteintech); rabbit monoclonal antibodies against HA (H6908, Sigma); mouse monoclonal antibodies against FLAG (F3165, Sigma); horseradish peroxidase (HRP)-FLAG (ZB15939, Servicebio); rabbit polyclonal antibodies against Myc (16286-1-AP, Proteintech); HRP-His (HRP-66005, Proteintech); IgG (I5381 and I5006, Sigma); BiP (11587-1-AP, Proteintech); phosphorylated PERK (29546-1-AP, Proteintech); p-IRE1α ( Ab124945 , Abcam); IRE1α (3294S, Cell Signaling Technology); PERK (20582-1-AP, Proteintech); phosphorylated eIF2α (3398S, Cell Signaling Technology); eIF2α (11170-1-AP, Proteintech); ATF6 (ab122897, Abcam); XBP1s (24868-1-AP, Proteintech); CHOP (15204-1-AP, Proteintech); p-IκBα (9246S, Cell Signaling Technology); IκBα (9242S, Cell Signaling Technology); GST (66001-2-Ig, Proteintech); β-actin (A2228, Sigma–Aldrich); anti-mouse IgG (H+L), F(ab')2 fragment (Alexa Fluor 594 Conjugate) (8890, Cell Signaling Technology); Alexa Fluor 555 goat anti-rabbit IgG (H+L) (A-21428, Invitrogen), and Alexa Fluor 488 goat anti-rabbit IgG (H+L) (A11008, Invitrogen) were purchased from the indicated manufacturers.

Techniques: Activation Assay, Transfection, Plasmid Preparation, Expressing, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction, Luciferase, Activity Assay, Western Blot, Membrane, Real-time Polymerase Chain Reaction, Binding Assay

Journal: The Journal of Biological Chemistry

Article Title: Brucella Omp25 activates the unfolded protein response to promote intracellular proliferation and inflammation

doi: 10.1016/j.jbc.2026.111333

Figure Lengend Snippet: Omp25 disrupts BiP interactions with PERK, IRE1α, and ATF6. A , effects of Omp25 on the association of BiP with PERK, IRE1α, and ATF6. HEK293T cells (1 × 10 7 ) were transfected with the indicated plasmids for 24 h, followed by co-IP and immunoblot analysis. B , Omp25 interacts with BiP but not with PERK, IRE1α, or ATF6. HEK293T cells (1 × 10 7 ) were transfected with the indicated plasmids for 24 h, followed by co-IP and immunoblot analysis. C , effects of Omp25 on the endogenous association of BiP with PERK, IRE1α, and ATF6. HEK293T cells (1 × 10 7 ) transfected with empty vector or Omp25-FLAG were analyzed by co-IP using anti-BiP antibody, followed by immunoblot analysis. D , effects of Omp25 on the association of BiP with PERK, IRE1α, and ATF6 in GST pull-down assays. Increasing amounts of recombinant His-Omp25 were incubated with GST-BiP and His-tagged luminal domains of PERK, ATF6, or IRE1α, followed by GST pull-down and immunoblot analysis. Band intensities were quantified by ImageJ. ATF6, activating transcription factor 6; BiP, binding immunoglobulin protein; Co-IP, coimmunoprecipitation; GST, glutathione S -transferase; HEK293T, human embryonic kidney 293T cell line; IRE1α, inositol-requiring enzyme 1 alpha; Omp25, outer membrane protein 25; PERK, PKR-like ER kinase.

Article Snippet: Mouse monoclonal antibodies against HA (66006, Proteintech); rabbit monoclonal antibodies against HA (H6908, Sigma); mouse monoclonal antibodies against FLAG (F3165, Sigma); horseradish peroxidase (HRP)-FLAG (ZB15939, Servicebio); rabbit polyclonal antibodies against Myc (16286-1-AP, Proteintech); HRP-His (HRP-66005, Proteintech); IgG (I5381 and I5006, Sigma); BiP (11587-1-AP, Proteintech); phosphorylated PERK (29546-1-AP, Proteintech); p-IRE1α ( Ab124945 , Abcam); IRE1α (3294S, Cell Signaling Technology); PERK (20582-1-AP, Proteintech); phosphorylated eIF2α (3398S, Cell Signaling Technology); eIF2α (11170-1-AP, Proteintech); ATF6 (ab122897, Abcam); XBP1s (24868-1-AP, Proteintech); CHOP (15204-1-AP, Proteintech); p-IκBα (9246S, Cell Signaling Technology); IκBα (9242S, Cell Signaling Technology); GST (66001-2-Ig, Proteintech); β-actin (A2228, Sigma–Aldrich); anti-mouse IgG (H+L), F(ab')2 fragment (Alexa Fluor 594 Conjugate) (8890, Cell Signaling Technology); Alexa Fluor 555 goat anti-rabbit IgG (H+L) (A-21428, Invitrogen), and Alexa Fluor 488 goat anti-rabbit IgG (H+L) (A11008, Invitrogen) were purchased from the indicated manufacturers.

Techniques: Transfection, Co-Immunoprecipitation Assay, Western Blot, Plasmid Preparation, Recombinant, Incubation, Binding Assay, Membrane

Journal: Scientific Reports

Article Title: SHP2 improves ovarian morphology and steroidogenic function in a rat PCOS model by modulating IRE1α/XBP1/NLRP3-mediated granulosa cell pyroptosis

doi: 10.1038/s41598-026-43536-2

Figure Lengend Snippet: A . Boxplot for data standardization evaluation. B . Principal component analysis scatter plot. C . Volcano plot of differentially expressed genes. D . Heatmap of differentially expressed genes. Ea. GO cellular component (CC) enrichment bubble plot. Eb. GO molecular function (MF) enrichment bubble plot. Ec. GO biological process (BP) enrichment bubble plot. Fa. KEGG pathway enrichment bubble plot. Fb. KEGG pathway enrichment lollipop plot. G . Correlation pie chart of key regulatory factors (IRE1α, XBP-1, p38, SP1, ZEB1, PKP3, Rb, E2F1, Cyclin, SHP2, BRD4). H . Scatter plot of E2F1-CDK1 co-regulation. I. Scatter plot of NLRP3-GSDMD pyroptosis axis.

Article Snippet: Antibodies against p-SHP2, p-IRE1α, XBP-1s, p-SP1, ZEB1, PKP3, nuclear E2F1, CyclinB1, NLRP3, GSDMD, and GAPDH were purchased from Wuhan Sanying Biotechnology Co., Ltd. IgG secondary antibodies and ECL hypersensitive luminescent solution were purchased from Beijing Biosynthesis Biotechnology Co., Ltd.

Techniques:

Journal: Scientific Reports

Article Title: SHP2 improves ovarian morphology and steroidogenic function in a rat PCOS model by modulating IRE1α/XBP1/NLRP3-mediated granulosa cell pyroptosis

doi: 10.1038/s41598-026-43536-2

Figure Lengend Snippet: SHP2 Alleviates PCOS Phenotypes via IRE1α/XBP1/NLRP3 and ZEB1/PKP3-Mediated Regulation. (A) Western blot detection of SHP2-regulated IRE1α/XBP1/ZEB1/PKP3 protein pathways. (B) Quantitative analysis of protein expression levels.* p < 0.05, ** p < 0.01, ns indicates P > 0.05; data are represented as the mean ± SD.

Article Snippet: Antibodies against p-SHP2, p-IRE1α, XBP-1s, p-SP1, ZEB1, PKP3, nuclear E2F1, CyclinB1, NLRP3, GSDMD, and GAPDH were purchased from Wuhan Sanying Biotechnology Co., Ltd. IgG secondary antibodies and ECL hypersensitive luminescent solution were purchased from Beijing Biosynthesis Biotechnology Co., Ltd.

Techniques: Western Blot, Expressing

Journal: Scientific Reports

Article Title: SHP2 improves ovarian morphology and steroidogenic function in a rat PCOS model by modulating IRE1α/XBP1/NLRP3-mediated granulosa cell pyroptosis

doi: 10.1038/s41598-026-43536-2

Figure Lengend Snippet: SHP2 improves ovarian morphology and steroidogenic function in a rat PCOS model by regulating the IRE1α/XBP1/NLRP3 and ZEB1/PKP3 signaling pathways, thereby influencing granulosa cell pyroptosis and proliferation.

Article Snippet: Antibodies against p-SHP2, p-IRE1α, XBP-1s, p-SP1, ZEB1, PKP3, nuclear E2F1, CyclinB1, NLRP3, GSDMD, and GAPDH were purchased from Wuhan Sanying Biotechnology Co., Ltd. IgG secondary antibodies and ECL hypersensitive luminescent solution were purchased from Beijing Biosynthesis Biotechnology Co., Ltd.

Techniques: Protein-Protein interactions