| 货号 | 24876T |
| 目标/特异性 | Each antibody in the Sequestosome Signaling Antibody Sampler Kit detects endogenous levels of its target protein. K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb detects polyubiquitin chains formed by Lys63 residue linkage. It does not react with monoubiquitin or polyubiquitin chains formed by linkage to a different lysine residue. TRAF6 (D21G3) Rabbit mAb is not predicted to cross-react with other TRAF family members. TrkA (12G8) Rabbit mAb does not cross-react with TrkB. |
| 供应商 | CST |
| 背景 | Sequestosome 1 (SQSTM1, p62) is a ubiquitin binding protein involved in cell signaling, oxidative stress, and autophagy (1-4). It was first identified as a protein that binds to the SH2 domain of p56Lck (5) and independently found to interact with PKCζ (6,7). SQSTM1 was subsequently found to interact with ubiquitin, providing a scaffold for several signaling proteins and triggering degradation of proteins through the proteasome or lysosome (8). Interaction between SQSTM1 and TRAF6 leads to the K63-linked polyubiquitination of TRAF6 and subsequent activation of the NF-κB pathway (9). Protein aggregates formed by SQSTM1 can be degraded by the autophagosome (4,10,11). SQSTM1 binds autophagosomal membrane protein LC3/Atg8, bringing SQSTM1-containing protein aggregates to the autophagosome (12). Lysosomal degradation of autophagosomes leads to a decrease in SQSTM1 levels during autophagy; conversely, autophagy inhibitors stabilize SQSTM1 levels. SQSTM1 also interacts with KEAP1, which is a cytoplasmic inhibitor of NRF2, a key transcription factor involved in cellular responses to oxidative stress (3). Under basal conditions, KEAP1 binds and retains NRF2 in the cytoplasm where it can be targeted for ubiquitin-mediated degradation (13). Small amounts of constitutive nuclear NRF2 maintain cellular homeostasis through regulation of basal expression of antioxidant response genes. Following oxidative or electrophilic stress, KEAP1 releases NRF2, thereby allowing the activator to translocate to the nucleus and bind to ARE-containing genes (14). The coordinated action of NRF2 and other transcription factors mediates the response to oxidative stress (15). Thus, accumulation of SQSTM1 can lead to an increase in NRF2 activity (3). KEAP1 also targets the down regulation of NF-κB activity by targeting IKKβ degradation (16). TrkA is a member of Trk receptor tyrosine kinases and is activated by NGF, which stimulates TrkA polyubiquitination (17,18). TrkA regulates proliferation and is important for development and maturation of the nervous system (19). SQSTM1 interaction with TRAF6 controls synthesis of K63 polyubiquititination on TrkA (18, 20). TrkA polyubiquitination is essential for neurotrophin-dependent receptor internalization, cell differentiation, and signaling (18). |
| 运输条件 | 0.75 |
| 存放说明 | -20C |
| 参考文献 | 1 . Geetha, T. et al. (2005) Mol Cell 20, 301-12. 2 . Kirkin, V. et al. (2009) Mol Cell 34, 259-69. 3 . Cullinan, S.B. et al. (2004) Mol Cell Biol 24, 8477-86. 4 . Seibenhener, M.L. et al. (2007) FEBS Lett 581, 175-9. 5 . Nguyen, T. et al. (2005) J Biol Chem 280, 32485-92. 6 . Segal, R.A. and Greenberg, M.E. (1996) Annu Rev Neurosci 19, 463-89. 7 . Komatsu, M. et al. (2010) Nat Cell Biol 12, 213-23. 8 . Jaiswal, A.K. (2004) Free Radic Biol Med 36, 1199-207. 9 . Bjørkøy, G. et al. (2006) Autophagy 2, 138-9. 10 . Joung, I. et al. (1996) Proc Natl Acad Sci USA 93, 5991-5. 11 . Sanchez, P. et al. (1998) Mol Cell Biol 18, 3069-80. 12 . Puls, A. et al. (1997) Proc Natl Acad Sci USA 94, 6191-6. 13 . Vadlamudi, R.K. et al. (1996) J Biol Chem 271, 20235-7. 14 . Wooten, M.W. et al. (2005) J Biol Chem 280, 35625-9. 15 . Wooten, M.W. et al. (2005) J Biol Chem 280, 35625-9. 16 . Bjørkøy, G. et al. (2005) J Cell Biol 171, 603-14. 17 . Komatsu, M. et al. (2007) Cell 131, 1149-63. 18 . Lee, D.F. et al. (2009) Mol Cell 36, 131-40. 19 . Pankiv, S. et al. (2007) J Biol Chem 282, 24131-45. 20 . Huang, E.J. and Reichardt, L.F. (2003) Annu Rev Biochem 72, 609-42. |
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition. | |
Western blot analysis comparing the titration of recombinant monoubiquitin, K48-linked polyubiquitin and K63-linked polyubiquitin using K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb (upper) and Ubiquitin Antibody #3933 (lower). | |
Western blot analysis of seven distinct recombinant polyubiquitin chains (300 ng each) using K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb (upper) and Ubiquitin Antibody #3933 (lower). | |
Western blot analysis of extracts from HeLa cells, untreated or treated with the proteasome inhibitor MG132 (10 µM for 6 hours), using K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb and Ubiquitin Antibody #3933 (lower). | |
Western blot analysis of various cell lines using K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb. | |
Western blot analysis of extracts from HeLa cells using K63-linkage Specific Polyubiquitin (D7A11) Rabbit mAb, untreated or following antibody pre-incubation with either K63 ubiquitinylated branched peptide to block the signal or a linear peptide surrounding K63 of ubiquitin that cannot block the signal. | |
Western blot analysis of extracts from various cell lines using TRAF6 (D21G3) Rabbit mAb. | |
Western blot analysis of extracts from 293T cells, either mock transfected (-) or transfected with a cDNA expression construct encoding full-length human TRAF6 (+), using TRAF6 (D21G3) Rabbit mAb. | |
Confocal immunofluorescent analysis of OVCAR8 cells using KEAP1 (D6B12) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). | |
Western blot analysis of extracts from various cell lines using KEAP1 (D6B12) Rabbit mAb. | |
Western blot analysis of extracts from OVCAR8 cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence®KEAP1 siRNA I #5285 (+) or SignalSilence® KEAP1 siRNA II #5289 (+), using KEAP1 (D6B12) Rabbit mAb (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The KEAP1 (D6B12) Rabbit mAb confirms silencing of KEAP1 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control. | |
Western blot analysis of extracts from various cell lines using SQSTM1/p62 (D5E2) Rabbit mAb. | |
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a tagged human SQSTM1/p62 construct (+), using SQSTM1/p62 (D5E2) Rabbit mAb. | |
Western blot analysis of extracts from SK-MEL-2 cells, untreated (-) or starved overnight in Earles Balanced Salt Solution (EBSS) (+), using SQSTM1/p62 (D5E2) Rabbit mAb. | |
Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence®SQSTM1/p62 siRNA I #6394 (+) or SignalSilence® SQSTM1/p62 siRNA II #6399 (+), using SQSTM1/p62 (D5E2) Rabbit mAb (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The SQSTM1/p62 (D5E2) Rabbit mAb confirms silencing of SQSTM1/p62 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control. |
