phospho irak1  (Proteintech)

Journal: Cell reports

Article Title: Nanoparticle-based itaconate treatment recapitulates low-cholesterol/low-fat diet-induced atherosclerotic plaque resolution

doi: 10.1016/j.celrep.2024.114911

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Article Snippet: anti-mouse phospho-IRAK1 (Thr387) Polyclonal Antibody , Bioss Antibodies , Cat#: bs-3194R; RRID:AB_10857144.

Techniques: Purification, Plasmid Preparation, Produced, Recombinant, Concentration Assay, Saline, Labeling, Membrane, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Phospholipid Assay, BIA-KA, RNAscope, HD Assay, Polymer, Sequencing, Expressing, Software, Microscopy

Journal: International immunopharmacology

Article Title: Artemisinin attenuates perinatal inflammation and consequent oxidative stress in oligodendrocyte precursor cells by inhibiting IRAK-4 and IRAK-1.

doi: 10.1016/j.intimp.2024.113117

Figure Lengend Snippet: Fig. 3. Docking analysis and CETSA results showing that IRAK-4 and IRAK-1 are both potential target proteins for ART. PyMOL and AutoDock were used for molecular modelling (Homo sapiens: irak4, PDB ID: 6O94; irak1, PDB ID: 6BFN) (Rattus norvegicus: irak4, UniProt: D4A7K4; irak1, UniProt: B2RYH5), and analysis of properties and binding sites (ART and IRAK-4; ART and IRAK-1) revealed that ART mainly forms hydrogen bonds and binds to sites near phosphorylation sites. b, c, d, e A CETSA was used to assess the binding of ART to its targets over a predefined temperature gradient (gradient: 45.0, 46.8, 49.8, 54.4, 59.9, 64.8, 68.0, and 70.0 ◦C). Western blot analysis of differences in protein expression (IRAK-4, IRAK-1, and β-actin) between the Ctrl and ART (2.5 μM) groups. Quantification of Band intensity. The data are expressed as the mean ± SEM (n = 6/group). *p < 0.05 and **p < 0.01 versus the indicated groups.

Article Snippet: For immunofluorescence, antigen retrieval was performed using Quick Antigen Retrieval Solution for Frozen Sections (P0090, Beyond, Shanghai, China), and the tissues were incubated with 0.3 % Triton X100 for 10 min, blocked and incubated with primary antibodies against phospho-IRAK4 (p-IRAK-4) (Thr345/Ser346) (1:100, DF7567, Affinity, Jiangsu, China), phospho-IRAK1 (p-IRAK-4) (Thr387) (1:100, AF8009, Affinity), NF-kB p65 (1:100, AF5006, Affinity), and Nrf2 (1:100, AF0639, Affinity) for 16 h. The samples were incubated with a goat antirat IgG antibody (1:200, SA00003-11, Proteintech) labelled with FITC, a goat anti-mouse IgG antibody (1:100, SA00009-1, Proteintech) labelled with Cy3, and a goat anti-rabbit IgG antibody (1:200, SA00003-2, Proteintech) labelled with FITC at 37 ◦C in the dark for 1.5 h. For immunohistochemistry, the tissues were incubated with enzyme-conjugated goat anti-mouse immunoglobulin G secondary antibody (SP-900; ZSGBBio, Beijing, China) solution for 15 min at 37 ◦C.

Techniques: Binding Assay, Phospho-proteomics, Western Blot, Expressing

Journal: Cell death & disease

Article Title: Selective targeting of IRAK1 attenuates low molecular weight hyaluronic acid-induced stemness and non-canonical STAT3 activation in epithelial ovarian cancer.

doi: 10.1038/s41419-024-06717-3

Figure Lengend Snippet: Fig. 2 IRAK1 is upregulated in HGSOC. A Heatmap of mRNA expression of TIR signaling genes from CBioPortal TCGA, Firehose Legacy serous cystadenocarcinoma dataset. B Copy number alterations of TIR signaling genes from CBioPortal TCGA, Firehose Legacy serous cystadenocarcinoma dataset. C Oncoprint of TCGA, Firehose Legacy serous cystadenocarcinoma dataset. D Boxplot of IRAK1 mRNA expression in normal tissue (NT), PT, and RT from TCGA ovarian cancer dataset. E Kaplan–Meier survival curve of TCGA ovarian cancer dataset (high, n = 201, low, n = 172, p = 0.037). F Diagnosis Age versus IRAK1 mRNA expression of TCGA, Firehose Legacy ovarian serous cystadenocarcinoma dataset. Spearman correlation = −0.23, p < 0.0001). G Representative immunohistochemistry for IRAK1 staining of the normal fallopian tube and PT from two patients from HGSOC TMA. H The histological staining score of IRAK1 in TMA, containing matched normal fallopian tube (ctrl), PT, and Met from 100 patients with stage 3/4 HGSOC. ns not significant, ***p < 0.001, ****p < 0.0001.

Article Snippet: IRAK1 (4504S), pSTAT3 (9145S), STAT3 (9139S), MYC (9402S), KLF4 (12173S), p-p65 (13346S), p65 (8242S), p-p38 (9216S), p38 (8690S), Notch-1 (3608S), and Notch-3(5276S) were purchased from Cell Signaling Technologies (Danvers, Massachusetts).

Techniques: Expressing, Biomarker Discovery, Immunohistochemistry, Staining

Journal: Cell death & disease

Article Title: Selective targeting of IRAK1 attenuates low molecular weight hyaluronic acid-induced stemness and non-canonical STAT3 activation in epithelial ovarian cancer.

doi: 10.1038/s41419-024-06717-3

Figure Lengend Snippet: Fig. 3 LMW HA activates non-canonical IRAK1 signaling and stemness. A Western blot for IRAK1 across EOC cell lines. B Western blot time course for pIRAK1 (T209) and total IRAK1 in A1847 and OVCAR8 cells following stimulation with LMW HA (200 ng/mL). GAPDH served as an internal control. C Representative images of spheroid formation assay in A1847 and OVCAR8 cells either unstimulated or stimulated with LMW HA for 14 days. D Quantification of (C). Data is represented as mean from three independent experiments ± SEM. E (Top) Representative image of low and high exposures of immunoassay-based kinase array following stimulation with or without LMW HA for 15 min. 1. pSTAT5; 2. pp38; 3. β-Catenin; and 4. pERK. (Bottom) Quantification of pixel density of relative to untreated control. F Western blot of A1847 and OVCAR8 time course assessing phosphorylated and total expression of p38 and STAT3 following stimulation with or without LMW HA. The STAT3 target gene, MYC, was also assessed. *p < 0.05, ****p < 0.0001.

Article Snippet: IRAK1 (4504S), pSTAT3 (9145S), STAT3 (9139S), MYC (9402S), KLF4 (12173S), p-p65 (13346S), p65 (8242S), p-p38 (9216S), p38 (8690S), Notch-1 (3608S), and Notch-3(5276S) were purchased from Cell Signaling Technologies (Danvers, Massachusetts).

Techniques: Western Blot, Control, Tube Formation Assay, Expressing

Journal: Cell death & disease

Article Title: Selective targeting of IRAK1 attenuates low molecular weight hyaluronic acid-induced stemness and non-canonical STAT3 activation in epithelial ovarian cancer.

doi: 10.1038/s41419-024-06717-3

Figure Lengend Snippet: Fig. 5 TCS2210 is a selective inhibitor of IRAK1. A Chemical structure of TCS2210. B In silico docking of TCS2210 with IRAK1: (i) space-filled model (ii) zoomed-in space-filled model with predicted hydrogen bonding interactions. C Kinome tree interaction map of Eurofins ScanMAX analysis with TCS2210. D Western blot of CETSA assay for IRAK1 following incubation with TCS2210 or DMSO vehicle control. E SPR analysis of TCS2210 with active recombinant IRAK1 enzyme. Data are shown for 1:1 kinetic model and 1:1 affinity model. F Western blot of A2780 cells for pIRAK1 (T209) and total IRAK1 following preincubation with or without TCS2210, and stimulation with or without LMW HA.

Article Snippet: IRAK1 (4504S), pSTAT3 (9145S), STAT3 (9139S), MYC (9402S), KLF4 (12173S), p-p65 (13346S), p65 (8242S), p-p38 (9216S), p38 (8690S), Notch-1 (3608S), and Notch-3(5276S) were purchased from Cell Signaling Technologies (Danvers, Massachusetts).

Techniques: In Silico, Western Blot, Incubation, Control, Recombinant