p62/KEAP1/NRF2 Pathway Antibody Sampler Kit #48768
Product Information
Kit Usage Information
Protocols
- 7074: Western Blotting
- 8047: Western Blotting, Immunofluorescence
- 12721: Western Blotting, Immunoprecipitation (Agarose), Immunofluorescence, Flow, ChIP Magnetic, Chromatin IP-seq
- 12741: Western Blotting, Immunohistochemistry (Paraffin), Immunofluorescence*, Flow
- 16177: Western Blotting, Immunohistochemistry (Paraffin), Immunofluorescence, Flow
- 39749: Western Blotting, Immunoprecipitation (Agarose)
- 43966: Western Blotting, Immunoprecipitation (Agarose)
- 62262: Western Blotting, Immunofluorescence
Product Description
Specificity / Sensitivity
Each antibody in the p62/KEAP1/NRF2 Pathway Antibody Sampler Kit detects endogenous levels of its target protein. Phospho-SQSTM1/p62 (Ser349) (E7M1A) Rabbit mAb recognizes endogenous levels of SQSTM1/p62 protein only when phosphorylated at Ser349.
Source / Purification
Monoclonal antibody is produced by immunizing animals with synthetic peptides corresponding to residues surrounding Leu44 of human LC3B (conserved in LC3A), Ala275 of human NRF2, Leu228 of human NQO1, Leu118 of mouse HO-1, and near the carboxy termini of human KEAP1 and SQSTM1/p62 proteins. Phosphorylation-specific monoclonal antibodies are produced by immunizing rabbits with synthetic phosphopeptides corresponding to Ser349 of human SQSTM1/p62 protein.
Background
The cap ’n’ collar (CNC), leucine zipper (bZIP) transcription factor NRF2 (also called nuclear factor erythroid 2-related factor 2 (NFE2L2)) is the master regulator of the cellular antioxidant response, regulating the expression of over 200 genes that contain antioxidant response elements (AREs) in their regulatory regions by heterodimerizing with small MAF proteins (1). While NRF2 is expressed in all cell types, its basal protein levels are usually kept low during homeostatic conditions, mainly by KEAP1 (Kelch-like ECH-associated protein 1). Under normal conditions, KEAP1 binds to and targets NRF2 for ubiquitination-dependent proteasomal degradation. Upon oxidative stress, KEAP1 is modified on some sensor cysteines, affecting its conformation and thus interfering its binding to NRF2, allowing newly synthesized NRF2 to accumulate and translocate to the nucleus to activate its target genes, including HO-1 (heme oxygenase 1) and NQO1 (NAD(P)H:quinone oxidoreductase 1) (2,3). Another mode of NRF2 regulation involves the autophagy adapter protein p62 (or sequestosome 1 [SQSTM1]) in a KEAP1-dependent but cysteine-independent manner, the so called non-canonical pathway. Autophagy is a tightly regulated cellular quality control system that removes damaged proteins or organelles. Autophagy can also be activated to degrade macromolecules to provide nutrients under cellular starvation stress. p62, especially upon phosphorylation at Ser349 (Ser351 in mouse p62), can compete with NRF2 for binding KEAP1 and, as a result, p62 sequesters KEAP1 into the autophagosome and prevents KEAP1-mediated NRF2 degradation. This process may also require autophagy protein LC3. In addition, studies also found that KEAP1 is a p62-regulated substrate for autophagy-mediated degradation, indicating that p62 also plays a role in controlling KEAP1 turnover (4,5). Dysregulation of autophagy results in prolonged NRF2 activation and this may contribute to many diseases, including cancer and neurodegenerative diseases (6-9).
- Zhu, M. and Fahl, W.E. (2001) Biochem Biophys Res Commun 289, 212-9.
- Bellezza, I. et al. (2018) Biochim Biophys Acta Mol Cell Res 1865, 721-33.
- Yamamoto, M. et al. (2018) Physiol Rev 98, 1169-203.
- Jiang, T. et al. (2015) Free Radic Biol Med 88, 199-204.
- Nezis, I.P. and Stenmark, H. (2012) Antioxid Redox Signal 17, 786-93.
- Rojo de la Vega, M. et al. (2018) Cancer Cell 34, 21-43.
- Sánchez-Martín, P. et al. (2019) FEBS J 286, 8-23.
- Shah, S.Z.A. et al. (2018) Front Mol Neurosci 11, 310.
- Ichimura, Y. and Komatsu, M. (2018) Front Oncol 8, 210.
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