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RIPK1 protein, human

RIPK1, or receptor-interacting serine/threonine-protein kinase 1, is a key regulator of cell death pathways.
It plays a central role in the induction of apoptosis, necroptosis, and inflammatory signaling in response to various stimuli.
The RIPK1 protein is involeved in the activation of NF-κB and MAPK signaling cascades, and its dysregulation has been implicated in the pathogenesis of numerous diseases, including inflammatory disorders, neurodegeneration, and cancer.
Understanding the complex functions and regulation of RIPK1 is crucial for the development of targeted therapies.

Most cited protocols related to «RIPK1 protein, human»

Ripk1fl/fl, Ripk1D138N/D138N (ref. 29 ), Triffl/fl, Faddfl/fl (ref. 30 (link)), FADD-IRES-eGFPfl/fl (ref. 4 (link)) and R26-StopflIkk2ca22 (link) mice expressing a constitutively active Ikk2 (Ikk2ca) transgene were generated by gene targeting in C57BL/6 embryonic stem (ES) cells. FLPe-Deleter31 (link) mice were used to delete the FRT-flanked neo cassette and Cre-Deleter32 (link) mice were employed to delete the RIPK1 floxed sequences in the germ line and generate Ripk1−/− mice and MEFs. Villin-Cre33 (link), VillinCreERT2 (ref. 34 (link)), K14-Cre35 (link), Tnfr1−/− (ref. 36 (link)), Myd88−/− (ref. 37 (link)) and Ripk3−/− (ref. 38 (link)) mice were backcrossed for at least ten generations into the C57BL/6 genetic background. Mice were maintained at the SPF animal facilities of the Institute for Genetics, University of Cologne, and of the University of Massachusetts Medical School and kept under a 12 h light cycle, and given a regular chow diet (Harlan diet no. 2918 or Prolab Isopro RMH3000 5P76) ad libitum. Germ-free mice were produced at the gnotobiotic facilities of the University of Ulm and of the Hannover Medical School. All animal procedures were conducted in accordance with European, national and institutional guidelines and protocols were approved by local government authorities (Landesamt für Natur, Umwelt und Verbraucherschutz Nordrhein-Westfalen, Germany). Animals requiring medical attention were provided with appropriate care and excluded from the experiments described. No other exclusion criteria existed. For antibiotic treatment of RIPK1IEC-KO mice one of two different broad spectrum antibiotic mixtures was added to the drinking water starting from embryonic day (E)17.5: 1 g l−1 ampicillin (ICN Biomedicals), 1 g l−1 neomycin (Sigma), 0.5 g l−1 Meronem (AstraZeneca) and 0.5 g l−1 ciprofloxacin (Fluka) or 1 g l−1 ampicillin (ICN Biomedicals), 1 g l−1 metronidazole (Sigma), 0.5 g l−1 vancomycin (Eberth) and 0.2 g l−1 ciprofloxacin (Fluka). In VillinCreERT2/Ripk1fl/fl mice a modified version of the first protocol was employed, where Meronem was replaced by 0.5 g l−1 vancomycin (Eberth). As neither the overall physiological response nor tissue pathology differed between the two regimens, mouse groups receiving either antibiotic mixture were analysed together and data from individual experiments were pooled. VillinCreERT2 recombinase activity was induced by three daily intraperitoneal administrations of 1 mg tamoxifen. Littermates not carrying the Villin-Cre, Villin-CreERT2 and K14-Cre transgenes were used as controls in all experiments. Mice of the indicated genotype were assigned at random to groups. Mouse studies were performed in a blinded fashion. Unless otherwise indicated, mice were analysed at 3 weeks of age. Groups included male and female animals.
Publication 2014
Ampicillin Animals Antibiotics Attention Ciprofloxacin Diet Embryo Embryonic Stem Cells Europeans FADD protein, human Females Genetic Background Genotype Germ Line Injections, Intraperitoneal Internal Ribosome Entry Sites Males Metronidazole Mice, Laboratory N-fluoresceinylphosphatidylethanolamine Neomycin physiology Recombinase RIPK1 protein, human RIPK3 protein, human Tamoxifen Tissues TNFRSF1A protein, human Transgenes Treatment Protocols Vancomycin villin
Tnfrsf1a−/−, Tnfrsf1b−/−, casp8−/− MEFs were provided by David Vaux (Walter and Eliza Hall Institute Biotechnology Centre, Australia). Ripk1−/− and Ripk3−/− MEFs were obtained from Zheng-gang Liu (National Institutes of Health) and Francis Chan (University of Massachusetts).
Publication 2014
CASP8 protein, human RIPK1 protein, human RIPK3 protein, human TNFRSF1A protein, human
For the immunoprecipitation of NLRP3 and ZBP1, RIPK3, or RIPK1 in the overexpression system, HEK293T cells were seeded into six-well plates and transfected with 600 ng each of pcDNA3-N-FLAG-NLRP3 (Addgene, #75127), RIPK3-GFP (Addgene #41382), pCMV-mRIPK1, or pcDNA-mCherry-ZBP1 expression plasmids for 30 h. For the immunoprecipitation of the complex by RIPK3, HEK293T cells in a 10 cm dish were co-transfected with 600 ng each of pCMV6-mASC-turboGFP (Origene MG201872), pCDH-CMV-CASP8, pcDNA3-N-FLAG-NLRP3 (Addgene, #75127), RIPK3-GFP (Addgene #41382), pCMV-mRIPK1, and pcDNA-mCherry-ZBP1 expression plasmids for 30 h. The pan-caspase inhibitor zVAD-fmk was used to inhibit cell death due to the overexpression of these molecules. Subsequently, cells were lysed in NP-40 lysis buffer (1% NP-40, 150 mM NaCl, 50 mM HEPES, and protease inhibitor cocktail) for 20–30 min. Cell lysates were then centrifuged at 13,000 × g for 10 min. Supernatants were collected and incubated with 1–2 μg of the indicated antibody overnight at 4°C. Protein A/G-plus agarose beads (Santa Cruz sc-2003) were added into the lysates and incubated for 1–2 h at 4°C. After incubation, the beads were washed 4 times with lysis buffer and boiled in 2 × SDS loading buffer at 100°C for 5 min. Immunoprecipitates in sample buffer were subjected to immunoblotting analysis.
For immunoprecipitation in the endogenous system, fully differentiated primary wild-type C57BL/6 BMDMs were seeded 24 h prior to stimulations. BMDMs were stimulated with LPS (100 ng/mL) alone, LPS + TAK1-inhibitor (5Z-7-oxozeaenol [0.1 μM]) or LPS + TAK1-inhibior + zVAD (30 mM). After 4 h of stimulation, BMDMs were lysed in NP-40 lysis buffer. Whole cell lysates were harvested and incubated with 3 mg of indicated primary antibodies overnight at 4°C. Protein A/G plus agarose beads were added to these samples and incubated at 4°C for another 2 h. Agarose beads were centrifuged and washed in NP-40 lysis buffer three times, and immunoprecipitates were eluted by adding sample buffer. Immunoprecipitates in sample buffer were then subjected to immunoblotting analysis.
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Publication 2020
Antibodies benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone Buffers Cardiac Arrest CASP8 protein, human Caspase Inhibitors Cell Death Cells HEPES Hyperostosis, Diffuse Idiopathic Skeletal Immunoblotting Immunoglobulins Immunoprecipitation Nonidet P-40 Plasmids Protease Inhibitors RIPK1 protein, human RIPK3 protein, human Sepharose Sodium Chloride Staphylococcal Protein A

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Publication 2020
Actins Antibodies Buffers Caspase Caspase-7 Caspase-8 Caspase 1 Caspase 3 Caspase 6 Cells Electrophoresis inhibitors Interleukin-1 beta LMNA protein, human Milk, Cow's Mus Nonidet P-40 Phosphoric Monoester Hydrolases polyacrylamide gels polyvinylidene fluoride Protease Inhibitors Proteins Rabbits Radioimmunoprecipitation Assay RIPK1 protein, human RIPK3 protein, human STAT1 protein, human Tissue, Membrane

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Publication 2014
Animals CASP8 protein, human FADD protein, human Institutional Animal Care and Use Committees Mice, House RIPK1 protein, human RIPK3 protein, human TNFRSF1A protein, human

Most recents protocols related to «RIPK1 protein, human»

RNA sequencing data and clinical information on 33 tumors and normal tissues (Supplementary Table S1) were retrieved from The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) using USCS Xena (https://xenabrowser.net). The "RMA" package13 (link) was used to remove not available and duplicates in the expression data; the data were then log2 (transcripts per kilobase million (TPM) + 1) converted for further analysis. Ten genes including NLRP3, CASP1, CASP8, TNFAIP3, RIPK1, RIPK3, NR2C2, RBCK1, PSTPIP2, and PYCARD (Supplementary Table S2) were identified as components of the PANoptosome, according to a previous study10 (link).
The data of immune cell infiltration were downloaded from the Tumor Immune Estimation Resource (TIMER) (http://timer.cistrome.org) and Immune Cell Abundance Identifier (ImmuCellAI) databases (http://bioinfo.life.hust.edu.cn/ImmuCellAI).
The dataset GSE35640 (65 metastatic melanoma patients treated with MAGE-A3 immunotherapy)14 (link), GSE 91061 (65 melanoma patients treated with Anti-CTLA4 and ant-PD1)15 (link), and GSE135222 (27 advanced non-small cell lung carcinoma patients treated with anti-PD-1/PD-L1)16 (link) were downloaded from the Gene Expression Omnibus database (https://www.ncbi.nlm.nih.gov) to explore the prediction ability of immunotherapy response based on PANoptosis scores.
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Publication 2023
CASP8 protein, human CD274 protein, human Cells CTLA4 protein, human Genes Genome Genotype Immunotherapy MAGEA3 protein, human Malignant Neoplasms Melanoma Neoplasms Non-Small Cell Lung Carcinoma NR2C2 protein, human Patients RIPK1 protein, human RIPK3 protein, human Tissues TNFAIP3 protein, human
Cellular extracts of 1x106 cells or 0.5 g of tissue were homogenized in RIPA lysis buffer (1% Triton X-100, 2% SDS, 150 mM NaCl, 10 mM HEPES, 2 mM EDTA) containing protease and phosphatase inhibitor cocktail (Roche, pH 8.0). After centrifugation at 13 000 × g for 5 min, cell lysates were prepared under reducing and denaturing conditions and subjected to SDS−PAGE. Equal concentrations of proteins were fractionated by electrophoresis on 10% acrylamide gels. The proteins were transferred onto a nitrocellulose membrane (Millipore, Billerica, MA, USA), followed by blocking of nonspecific binding sites in 5% nonfat milk in TBST (50 mM Tris-HCl - pH 7.4, 150 mM NaCl, 0.05% Tween 20) for 1 h at room temperature and blotted with primary antibodies in TBST overnight at 4°C. The following antibodies were used: anti-phospho-RIPK1 (Ser166) (Cell Signaling # 31122S), anti-RIPK3 (D8J3 L) (Cell Signaling # 15828S), anti-MLKL (D2I6N) (Cell Signaling #14993S), anti-phospho-MLKL (Cell Signaling #91689S) and anti-β-actin (Sigma, #A1978). Proteins of interest were identified by incubating the membrane with IRDye® LICOR secondary antibodies in TBST, followed by fluorescence imaging detection using the Odyssey® system (CLx Imaging System). Protein bands were quantified by densitometric image analysis using ImageJ software. All the data were normalized by β-actin expression quantification.
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Publication 2023
Acrylamide Actins Antibodies Binding Sites Buffers Cardiac Arrest Cell Extracts Cells Centrifugation Densitometry Edetic Acid Electrophoresis Gels HEPES L Cells Milk, Cow's Nitrocellulose Peptide Hydrolases Phosphoric Monoester Hydrolases Proteins Radioimmunoprecipitation Assay RIPK1 protein, human RIPK3 protein, human SDS-PAGE Sodium Chloride Tissue, Membrane Tissues Triton X-100 Tromethamine Tween 20
Primary antibodies used in this study were as follows: anti-ZBP1 (Adipogen, Zippy-1, AG-20B-0010, 1:1000), anti-mouse MLKL (phospho S345, Abcam, ab196436, 1:1000), anti-human MLKL (phospho S358, Abcam, ab187091, 1:1000), anti-MLKL (Proteintech, 66675-1-Ig, 1:1000), anti-RIP (phospho S166, CST, 31122, 1:1000), anti-RIPK1 (CST, 3493, 1:1000), anti-RIPK3 (phosphor T231 + 232, Abcam, ab222320, 1:1000), anti-RIPK3 (Santa Cruz, sc-374639, 1:1000), COL1A1 (Abcam, Ab21286, 1:2000), α-SMA (Sigma Aldrich, A2547, 1:1000), and the appropriate IRDyeTM 800-conjugated secondary antibody (LI-COR, 926-32210, 926-32211, 1:10,000). Signals were detected using the Odyssey Imaging System (LI-COR Biosciences, Lincoln, NE, USA) and analyzed with Odyssey software. Results were normalized relative to GAPDH (CST, 2118 S, 1:1000) or α-tubulin (Proteintech, 11224-1-AP, 1:1000) expression.
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Publication 2023
alpha-Tubulin Antibodies GAPDH protein, human Homo sapiens Immunoglobulins Mus Phosphorus RIPK1 protein, human RIPK3 protein, human
The genes involved in the PANoptosis process were collected from previous studies, including ADAR, MEFV, AIM2, NLRP1, NLRC4, NLRP9, NLRP3, ZBP1, TNFRSF1A, PYCARD, FADD, CASP10, CASP1, CASP2, CASP12, CASP4, CASP3, CASP6, CASP5, CASP7, DFNA5, CASP8, MLKL, GSDMD, RIPK3, RIPK1, NAIP, TNF, NLRP6, GSDMA, GSDMC, GSDMB, APAF1, BAK, BAX, DIABLO, DCN, CASP9, and FAS (Wang and Kanneganti, 2021 (link)).
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Publication 2023
APAF1 protein, human CASP2 protein, human CASP4 protein, human CASP6 protein, human CASP7 protein, human CASP8 protein, human CASP9 protein, human CASP10 protein, human Caspase 3 Deafness, Autosomal Dominant 5 FADD protein, human Genes NAIP protein, human RIPK1 protein, human RIPK3 protein, human TNFRSF1A protein, human
For RNA, primers were designed to amplify desired genomic regions that correspond to peaks of boundary RNA (GRO-seq) at the B1 and B2 CTCF sites of the INK4a/ARF TAD boundary. The amplified products were cloned into the pcDNA3 BoxB plasmid (a gift from Howard Chang, Addgene 29729). All clones were confirmed by Sanger sequencing and subsequently used for RNA synthesis using T7 RNA polymerase (Promega P207e) and biotin RNA labeling mix (Roche 11685597910). Primer sequences are listed in Supplemental Table S1.
RNA immunoprecipitation was adopted from Jayani et al. (2017) (link), with some modifications. HeLa cells were grown to 90%–95% confluency and were harvested by scraping and washed twice with ice-cold 1× PBS to gently resuspend in 2 mL of PBS. A volume of 2 mL of nuclear isolation buffer (NIB; 40 mM Tris–HCl at pH 7.5, 20 mM MgCl2, 1.28 M sucrose, 4% Triton X-100, 1 mM PMSF, protease inhibitors, and 20 U/mL SUPERase inhibitor; Thermo Fisher Scientific AM2694) was added, and the pellet was gently resuspended. Then, 6 mL of distilled water was added and kept on ice for 20 min with intermittent gentle shaking. Nuclei were pelleted by centrifugation at 2500g for 5 min at 4°C. The pellet was resuspended in 1 mL of RIP buffer (25 mM Tri–HCl at pH 7.4, 150 mM KCl, 0.5 mM DTT, 0.5% NP-40, 1 mM PMSF, protease inhibitors, and 20 U/mL SUPERase inhibitor) and incubated on ice for 5 min. Nuclei were sheared by 10 cycles of sonication (30 sec on and 30 sec off) in a Bioruptor (Diagenode) followed by centrifugation at 12,000 rpm for 10 min at 4°C. One microgram of biotinylated RNA was incubated with 20 μL of RNA structure buffer (RSB; 10 mM Tris–HCl at pH 7.0, 100 mM KCl, 10 mM MgCl2, 1 mM PMSF, protease inhibitors, and 20 U/mL SUPERase inhibitor) for 5 min at room temperature. Folded RNA was mixed with 1 mg of nuclear extract in 500 µL of RIP buffer and rotated for 1 h at 4°C. Fifteen microliters of Dynabeads MyOne streptavidin beads were added, and rotation continued for one more hour. Samples were washed three times with RIP buffer, and beads with proteins were boiled in 2×SDS for 10 min. Immunoblotting for CTCF was performed using CTCF antibody (Cell Signaling Technology 3418).
Publication 2023
Anabolism bacteriophage T7 RNA polymerase Biotin Buffers CDKN2A Gene Cell Nucleus Centrifugation Clone Cells Cold Temperature CTCF protein, human Genome HeLa Cells Immunoglobulins Immunoprecipitation isolation Magnesium Chloride Nonidet P-40 Oligonucleotide Primers Plasmids Promega Protease Inhibitors Proteins RIPK1 protein, human Streptavidin Sucrose Triton X-100 Tromethamine

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RIPK1 is a protein kinase that plays a key role in regulating cell death and inflammatory signaling pathways. It is a central component of the death receptor complex and can initiate both apoptotic and necroptotic cell death. RIPK1 is also involved in the activation of NF-κB and MAPK signaling cascades, which can lead to the expression of pro-inflammatory genes.
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RIPK1 is a protein that plays a key role in cellular signaling pathways. It is involved in the regulation of cell death and inflammation. RIPK1 is a widely used research tool in the study of these cellular processes.
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Anti-RIPK1 is a laboratory antibody product that targets RIPK1 (receptor-interacting serine/threonine-protein kinase 1), a key protein involved in cellular signaling pathways. This antibody can be used for the detection and analysis of RIPK1 in various research applications.
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Anti-RIPK1 is a laboratory tool used for the detection and quantification of RIPK1 (Receptor-Interacting Serine/Threonine-Protein Kinase 1) in biological samples. It functions as a research reagent to assist in the study of RIPK1-related cellular processes and signaling pathways.
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PVDF membranes are a type of laboratory equipment used for a variety of applications. They are made from polyvinylidene fluoride (PVDF), a durable and chemically resistant material. PVDF membranes are known for their high mechanical strength, thermal stability, and resistance to a wide range of chemicals. They are commonly used in various filtration, separation, and analysis processes in scientific and research settings.
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β-actin is a protein that is found in all eukaryotic cells and is involved in the structure and function of the cytoskeleton. It is a key component of the actin filaments that make up the cytoskeleton and plays a critical role in cell motility, cell division, and other cellular processes.
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Caspase-3 is a key enzyme involved in the execution phase of cell apoptosis (programmed cell death). It plays a central role in the apoptotic pathway by cleaving various cellular substrates, leading to the characteristic morphological and biochemical changes associated with apoptosis.
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RIPK3 is a recombinant protein that functions as a serine/threonine-protein kinase involved in regulating necroptosis, a form of programmed cell death. It plays a central role in the RIP1-RIP3 complex, which is a key mediator of necroptosis.
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Necrostatin-1 is a laboratory compound that functions as a selective inhibitor of necroptosis, a form of programmed cell death. It acts by blocking the receptor-interacting serine/threonine-protein kinase 1 (RIPK1), a key regulator of necroptosis. Necrostatin-1 is commonly used in scientific research to study cell death pathways.
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Cleaved caspase-3 is an antibody that detects the activated form of caspase-3 protein. Caspase-3 is a key enzyme involved in the execution phase of apoptosis, or programmed cell death. The cleaved caspase-3 antibody specifically recognizes the active, cleaved form of the enzyme and can be used to monitor and quantify apoptosis in experimental systems.

More about "RIPK1 protein, human"

Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) is a crucial regulator of various cell death pathways, including apoptosis, necroptosis, and inflammatory signaling.
This multifunctional protein is involved in the activation of important signaling cascades like NF-κB and MAPK, making it a key player in the pathogenesis of numerous diseases, such as inflammatory disorders, neurodegeneration, and cancer.
Understanding the complex roles and regulation of RIPK1 is crucial for the development of targeted therapies.
RIPK1 can be inhibited by small-molecule compounds like Necrostatin-1, which has been shown to have neuroprotective effects.
Additionally, RIPK3, another member of the RIPK family, is closely linked to RIPK1 and plays a role in necroptosis.
Researchers often utilize RIPK1-specific antibodies, such as Anti-RIPK1, to study the expression and activation of this protein.
Western blotting techniques using PVDF membranes and the detection of cleaved Caspase-3, a marker of apoptosis, can provide insights into RIPK1-mediated cell death pathways.
By leveraging the latest scientific advancements and AI-driven platforms like PubCompare.ai, researchers can enhance the reproducibility and accuracy of their RIPK1-related studies, leading to a better understanding of this crucial regulator and the development of effective therapies.