SQSTM1/p62 Antibody - #AF5384

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Product: SQSTM1/p62 Antibody
Catalog: AF5384
Description: Rabbit polyclonal antibody to SQSTM1/p62
Application: WB IHC IF/ICC
Reactivity: Human, Mouse, Rat, Monkey
Prediction: Pig, Zebrafish, Bovine, Horse, Sheep, Rabbit, Chicken
Mol.Wt.: 62 kDa; 48kD(Calculated).
Uniprot: Q13501
RRID: AB_2837869

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Data Sheet
COA
Protocols
WB Handbook
IHC Protocol
IF/ICC Protocol

Product Info

Source:
Rabbit
Application:
WB 1:500-1:2000, IF/ICC 1:100-1:500, IHC 1:50-1:200
*The optimal dilutions should be determined by the end user.
*Tips:

WB: For western blot detection of denatured protein samples. IHC: For immunohistochemical detection of paraffin sections (IHC-p) or frozen sections (IHC-f) of tissue samples. IF/ICC: For immunofluorescence detection of cell samples. ELISA(peptide): For ELISA detection of antigenic peptide.

Reactivity:
Human,Mouse,Rat,Monkey
Prediction:
Pig(100%), Zebrafish(92%), Bovine(100%), Horse(100%), Sheep(100%), Rabbit(100%), Chicken(85%)
Clonality:
Polyclonal
Specificity:
SQSTM1/p62 Antibody detects endogenous levels of total SQSTM1/p62.
RRID:
AB_2837869
Cite Format: Affinity Biosciences Cat# AF5384, RRID:AB_2837869.
Conjugate:
Unconjugated.
Purification:
The antiserum was purified by peptide affinity chromatography using SulfoLink™ Coupling Resin (Thermo Fisher Scientific).
Storage:
Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol. Store at -20 °C. Stable for 12 months from date of receipt.
Alias:

Fold/Unfold

A170; DMRV; EBI 3 associated protein of 60 kDa; EBI 3 associated protein p60; EBI3 associated protein of 60 kDa; EBI3 associated protein p60; EBI3-associated protein of 60 kDa; EBIAP; FTDALS3; MGC127197; ORCA; OSF-6; Osi; OSIL; Oxidative stress induced like; p60; p62; p62B; Paget disease of bone 3; PDB 3; PDB3; Phosphotyrosine independent ligand for the Lck SH2 domain of 62 kDa; Phosphotyrosine independent ligand for the Lck SH2 domain p62; Phosphotyrosine-independent ligand for the Lck SH2 domain of 62 kDa; PKC-zeta-interacting protein; Protein kinase C-zeta-interacting protein; Sequestosome 1; Sequestosome-1; SQSTM 1; SQSTM_HUMAN; Sqstm1; STAP; STONE14; Ubiquitin binding protein p62; Ubiquitin-binding protein p62; ZIP 3; ZIP; ZIP3;

Immunogens

Immunogen:
Uniprot:

Q13501(SQSTM_HUMAN) >>Visit HPA database.

Gene(ID):
SQSTM1(8878)
Expression:
Q13501 SQSTM_HUMAN:

Ubiquitously expressed.

Description:
Adapter protein which binds ubiquitin and may regulate the activation of NFKB1 by TNF-alpha, nerve growth factor (NGF) and interleukin-1. May play a role in titin/TTN downstream signaling in muscle cells. May regulate signaling cascades through ubiquitination. Adapter that mediates the interaction between TRAF6 and CYLD (By similarity). May be involved in cell differentiation, apoptosis, immune response and regulation of K(+) channels.
Sequence:
MASLTVKAYLLGKEDAAREIRRFSFCCSPEPEAEAEAAAGPGPCERLLSRVAALFPALRPGGFQAHYRDEDGDLVAFSSDEELTMAMSYVKDDIFRIYIKEKKECRRDHRPPCAQEAPRNMVHPNVICDGCNGPVVGTRYKCSVCPDYDLCSVCEGKGLHRGHTKLAFPSPFGHLSEGFSHSRWLRKVKHGHFGWPGWEMGPPGNWSPRPPRAGEARPGPTAESASGPSEDPSVNFLKNVGESVAAALSPLGIEVDIDVEHGGKRSRLTPVSPESSSTEEKSSSQPSSCCSDPSKPGGNVEGATQSLAEQMRKIALESEGRPEEQMESDNCSGGDDDWTHLSSKEVDPSTGELQSLQMPESEGPSSLDPSQEGPTGLKEAALYPHLPPEADPRLIESLSQMLSMGFSDEGGWLTRLLQTKNYDIGAALDTIQYSKHPPPL

Predictions

Predictions:

Score>80(red) has high confidence and is suggested to be used for WB detection. *The prediction model is mainly based on the alignment of immunogen sequences, the results are for reference only, not as the basis of quality assurance.

Species
Results
Score
Pig
100
Horse
100
Bovine
100
Sheep
100
Rabbit
100
Zebrafish
92
Chicken
85
Xenopus
67
Dog
0
Model Confidence:
High(score>80) Medium(80>score>50) Low(score<50) No confidence

PTMs - Q13501 As Substrate

Site PTM Type Enzyme
A2 Acetylation
K7 Ubiquitination
Y9 Phosphorylation
K13 Acetylation
K13 Ubiquitination
S24 Phosphorylation
S28 Phosphorylation
S49 Phosphorylation
Y67 Phosphorylation
K91 Ubiquitination
T138 Phosphorylation
K141 Ubiquitination
S143 Phosphorylation
Y148 Phosphorylation
S152 Phosphorylation
K157 Ubiquitination
K165 Ubiquitination
S170 Phosphorylation
S176 Phosphorylation
S182 Phosphorylation
R183 Methylation
K187 Ubiquitination
K189 Ubiquitination
S207 Phosphorylation
T221 Phosphorylation
S224 Phosphorylation
S226 Phosphorylation
S233 Phosphorylation
S249 Phosphorylation
K264 Ubiquitination
S266 Phosphorylation
T269 Phosphorylation P06493 (CDK1) , O15264 (MAPK13)
S272 Phosphorylation P06493 (CDK1) , O15264 (MAPK13)
S275 Phosphorylation
S276 Phosphorylation
S277 Phosphorylation
T278 Phosphorylation
K281 Ubiquitination
S282 Phosphorylation
S284 Phosphorylation
S287 Phosphorylation
S288 Phosphorylation
S291 Phosphorylation
S294 Phosphorylation
K295 Ubiquitination
S306 Phosphorylation
K313 Ubiquitination
S318 Phosphorylation
S328 Phosphorylation
S332 Phosphorylation
T339 Phosphorylation
S342 Phosphorylation
S343 Phosphorylation
K344 Acetylation
K344 Ubiquitination
S349 Phosphorylation
T350 Phosphorylation
S355 Phosphorylation
S361 Phosphorylation
S365 Phosphorylation
S366 Phosphorylation
S370 Phosphorylation
T375 Phosphorylation
K378 Acetylation
K378 Ubiquitination
Y383 Phosphorylation
S403 Phosphorylation Q9UHD2 (TBK1)
S407 Phosphorylation
K420 Ubiquitination
T430 Phosphorylation
Y433 Phosphorylation
S434 Phosphorylation
K435 Ubiquitination

Research Backgrounds

Function:

Autophagy receptor required for selective macroautophagy (aggrephagy). Functions as a bridge between polyubiquitinated cargo and autophagosomes. Interacts directly with both the cargo to become degraded and an autophagy modifier of the MAP1 LC3 family. Along with WDFY3, involved in the formation and autophagic degradation of cytoplasmic ubiquitin-containing inclusions (p62 bodies, ALIS/aggresome-like induced structures). Along with WDFY3, required to recruit ubiquitinated proteins to PML bodies in the nucleus. May regulate the activation of NFKB1 by TNF-alpha, nerve growth factor (NGF) and interleukin-1. May play a role in titin/TTN downstream signaling in muscle cells. May regulate signaling cascades through ubiquitination. Adapter that mediates the interaction between TRAF6 and CYLD (By similarity). May be involved in cell differentiation, apoptosis, immune response and regulation of K(+) channels. Involved in endosome organization by retaining vesicles in the perinuclear cloud: following ubiquitination by RNF26, attracts specific vesicle-associated adapters, forming a molecular bridge that restrains cognate vesicles in the perinuclear region and organizes the endosomal pathway for efficient cargo transport. Promotes relocalization of 'Lys-63'-linked ubiquitinated STING1 to autophagosomes. Acts as an activator of the NFE2L2/NRF2 pathway via interaction with KEAP1: interaction inactivates the BCR(KEAP1) complex, promoting nuclear accumulation of NFE2L2/NRF2 and subsequent expression of cytoprotective genes.

PTMs:

Phosphorylated. May be phosphorylated by PRKCZ (By similarity). Phosphorylated in vitro by TTN. Phosphorylation at Ser-403 by ULK1 is stimulated by SESN2. Phosphorylated at Ser-403 by TBK1, leading to promote relocalization of 'Lys-63'-linked ubiquitinated STING1 to autophagosomes. Phosphorylation at Ser-349 by MTOR promotes interaction with KEAP1 and inactivation of the BCR(KEAP1) complex, promoting NFE2L2/NRF2 nuclear accumulation and expression of phase II detoxifying enzymes (By similarity).

Ubiquitinated by RNF26: ubiquitinated SQSTM1 attracts specific vesicle-associated adapters, forming a molecular bridge that restrains cognate vesicles in the perinuclear region and organizes the endosomal pathway for efficient cargo transport. Deubiquitination by USP15 releases target vesicles for fast transport into the cell periphery. Ubiquitinated by the BCR(KEAP1) complex at Lys-420, increasing SQSTM1 sequestering activity and promoting its degradation. Ubiquitinated via 'Lys-29' and 'Lys-33'-linked polyubiquitination leading to xenophagic targeting of bacteria and inhibition of their replication.

Subcellular Location:

Cytoplasm>Cytosol. Late endosome. Lysosome. Cytoplasmic vesicle>Autophagosome. Nucleus. Endoplasmic reticulum. Nucleus>PML body. Cytoplasm>Myofibril>Sarcomere.
Note: In cardiac muscle, localizes to the sarcomeric band (By similarity). Commonly found in inclusion bodies containing polyubiquitinated protein aggregates. In neurodegenerative diseases, detected in Lewy bodies in Parkinson disease, neurofibrillary tangles in Alzheimer disease, and HTT aggregates in Huntington disease. In protein aggregate diseases of the liver, found in large amounts in Mallory bodies of alcoholic and nonalcoholic steatohepatitis, hyaline bodies in hepatocellular carcinoma, and in SERPINA1 aggregates. Enriched in Rosenthal fibers of pilocytic astrocytoma. In the cytoplasm, observed in both membrane-free ubiquitin-containing protein aggregates (sequestosomes) and membrane-surrounded autophagosomes. Colocalizes with TRIM13 in the perinuclear endoplasmic reticulum. Co-localizes with TRIM5 in cytoplasmic bodies. When nuclear export is blocked by treatment with leptomycin B, accumulates in PML bodies.

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionSubcellular location
Tissue Specificity:

Ubiquitously expressed.

Subunit Structure:

Homooligomer or heterooligomer; may form homotypic arrays. Dimerization interferes with ubiquitin binding. Interacts directly with PRKCI and PRKCZ (Probable). Forms ternary complexes with PRKCZ and KCNAB2 or PRKCZ and GABBR3. Also interacts with KCNAB1, GABRR1, GABRR2 and GABRR3. Forms an NGF-induced complex with IKBKB, PRKCI and TRAF6 (By similarity). Interacts with EBI3, LCK, RASA1, PRKCZ, PRKCI, NR2F2, NTRK1, NTRK2, NTRK3, NBR1, MAP2K5, TRIM13, TRIM55 and MAPKAPK5. Interacts with the proteasome subunits PSMD4 and PSMC2. Interacts with K63-polyubiquitinated MAPT/TAU. Interacts with IKBKB through PRKCZ and PRKCI. Interacts with NGFR through TRAF6 and bridges that complex to NTRK1. Forms a complex with MAP2K5 and PRKCZ or PRKCI. Component of a ternary complex with PAWR and PRKCZ. Upon TNF-alpha stimulation, interacts with RIPK1 probably bridging IKBKB to the TNF-R1 complex composed of TNF-R1/TNFRSF1A, TRADD and RIPK1. Forms a complex with JUB/Ajuba, PRKCZ and TRAF6. Interacts with TRAF6 and CYLD. Identified in a complex with TRAF6 and CYLD (By similarity). Identified in a heterotrimeric complex with ubiquitin and ZFAND5, where ZFAND5 and SQSTM1 both interact with the same ubiquitin molecule. Directly interacts with MAP1LC3A and MAP1LC3B, as well as with other MAP1 LC3 family members, including GABARAP, GABARAPL1 and GABARAPL2; these interactions are necessary for the recruitment MAP1 LC3 family members to inclusion bodies containing polyubiquitinated protein aggregates and for their degradation by autophagy. Interacts with FHOD3. Interacts with TRMI5. Interacts with SESN1. Interacts with SESN2. Interacts with ULK1. Interacts with UBD. Interacts with WDR81; the interaction is direct and regulates the interaction of SQSTM1 with ubiquitinated proteins. Interacts with WDFY3; this interaction is required to recruit WDFY3 to cytoplasmic bodies and to PML bodies. Interacts with TRIM23. Interacts with LRRC25. Interacts with TRIM50. Interacts with TRIM16. Interacts with STING1; leading to relocalization of STING1 to autophagosomes. Interacts (when phosphorylated at Ser-349) with KEAP1; the interaction is direct and inactivates the BCR(KEAP1) complex by sequestering KEAP1 in inclusion bodies, promoting its degradation.

Family&Domains:

The UBA domain binds specifically 'Lys-63'-linked polyubiquitin chains of polyubiquitinated substrates. Mediates the interaction with TRIM55. Both the UBA and PB1 domains are necessary and sufficient for the localization into the ubiquitin-containing inclusion bodies.

The PB1 domain mediates homooligomerization and interactions with FHOD3, MAP2K5, NBR1, PRKCI, PRKCZ and WDR81. Both the PB1 and UBA domains are necessary and sufficient for the localization into the ubiquitin-containing inclusion bodies.

The ZZ-type zinc finger mediates the interaction with RIPK1.

The LIR (LC3-interacting region) motif mediates the interaction with ATG8 family proteins.

References

1). Sirt3-mediated mitophagy regulates AGEs-induced BMSCs senescence and senile osteoporosis. Redox Biology (PubMed: 33662874) [IF=11.4]

Application: WB    Species: mice    Sample: bone marrow mesenchymal stem (BMSCs)

Fig. 2. Effects of different concentrations of AGEs on mitochondrial function and mitophagy of BMSCs. The BMSCs were treated with AGEs (50–200 μg/mL) or BSA for 24–72 h. (A) Representative fluorescence images with DCF (green) staining in BMSCs stimulated with AGEs. Scale bar: 50 μm. (B) Representative fluorescence images with Mito-SOX (red) and Mito-Tracker (green) double-staining in BMSCs stimulated with AGEs. Scale bar: 50 μm. (C) The MMP was detected through JC-1 staining in BMSCs stimulated with AGEs. Scale bar: 50 μm. (D) Representative fluorescence images with Mtphagy Dye (red) and Mito-Tracker (green) double-staining in BMSCs stimulated with AGEs. Scale bar: 50 μm. (E) Representative fluorescence images with LC3B (red) and Mito-Tracker (green) double-staining in BMSCs stimulated with AGEs. Scale bar: 50 μm. (F) Representative Western blotting assay and quantitation of the level of LC3B, P62, Parkin, Sirt3. **p < 0.01 versus BSA. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)
2). Polysaccharide from Strongylocentrotus nudus eggs regulates intestinal epithelial autophagy through CD36/PI3K-Akt pathway to ameliorate inflammatory bowel disease. International Journal of Biological Macromolecules (PubMed: 37327932) [IF=8.2]
3). Patchouli alcohol protects against chronic unpredictable mild stress-induced depressant-like behavior through inhibiting excessive autophagy via activation of mTOR signaling pathway. BIOMEDICINE & PHARMACOTHERAPY (PubMed: 32244196) [IF=7.5]

Application: WB    Species: rat    Sample: hippocampus

Fig. 6. |The effect of PA on the levels of ex-pression of LC3-II and p62 protein in the hippocampus. (A) The effect of PA on LC3-II and p62 protein levels in hippocampus were in-vestigated by western blot analysis.
4). Huaier polysaccharides suppress triple-negative breast cancer metastasis and epithelial-mesenchymal transition by inducing autophagic degradation of Snail. Cell and Bioscience (PubMed: 34481526) [IF=7.5]

Application: WB    Species: human    Sample: MDA-MB-231 cells

Fig. 2 |PS-T reverses EMT by inducing autophagy. a MDA-MB-231 cells are treated with 5 μg/mL PS-T for 24 h. The levels of each autophagy marker in MDA-MB-231 cells are quantifed using the NIH ImageJ software. (mean ± SD, *P < 0.05 and ***P < 0.001).

Application: WB    Species: Human    Sample: MDA-MB-231 cells

Fig. 2 PS-T reverses EMT by inducing autophagy. a MDA-MB-231 cells are treated with 5 μg/mL PS-T for 24 h. The levels of each autophagy marker in MDA-MB-231 cells are quantified using the NIH ImageJ software. (mean  ±  SD, *P  <  0.05 and ***P  <  0.001). b Confocal microscopy images of cells treated with or without PS-T (5 μg/mL) for 24 h after transfection with mRFP-GFP-LC3 plasmid (scale bars: 10 μm). Quantification of LC3-GFP and LC3-RFP puncta/cells from three independent experiments (***P  <  0.001). c MDA-MB-231 cells are treated with PS-T (5 μg/mL) for 24 h with or without LY294002 (10 µM). The levels of each EMT marker in MDA-MB-231cells were quantified using NIH ImageJ software. (mean  ±  SD, *P  <  0.05 and **P  <  0.01)
5). Hydroxysafflor yellow A promotes apoptosis via blocking autophagic flux in liver cancer. Biomedicine & Pharmacotherapy (PubMed: 33485070) [IF=7.5]

Application: WB    Species: Human    Sample: HepG2 and Huh7 cells

Fig. 3. HSYA blocks autophagic flux in liver cancer cells. (A) Western blot assay for p62 expression of HepG2 and Huh7 cells treated with HSYA for 24 h; β-actin served as control. (B) HepG2 and Huh7 cells expressing mRFP-GFP-LC3 followed by treatment with 160 μmol/L HSYA for 24 h. The yellow dots (autophagosomes) and red dots (autolysosome) were captured by laser confocal microscopy; the GFP/RFP fluorescence intensity ratio was assessed by ImageJ. Scale bar: 20 μm, n = 6 images/group. Experiments were repeated three times, and all data are presented as mean ± S.D. *p < 0.05, **p < 0.01, ***p < 0.001 compared with the control group.
6). Geniposide ameliorates glucocorticoid-induced osteoblast apoptosis by activating autophagy. Biomedicine & Pharmacotherapy (PubMed: 36271582) [IF=7.5]

Application: IHC    Species: Rat    Sample: MC3T3-E1 cells

Fig. 3. The effects of GEN on autophagy in DEX-treated osteoblasts in vivo and in vitro. The immunohistochemical changes (× 40 magnification) of P62 (A, B), LC3-II (A, C), and Beclin-1 (A, D) were analyzed. The proteins expression of P62 (E, F), LC3-II (E, G), and Beclin-1 (E, H) in DEX-treated MC3T3-E1 cells were determined by western blot. * *P˂0.01. NC, negative control group; DEX, model rat group treated with DEX (5 mg/kg).

Application: WB    Species: Rat    Sample: MC3T3-E1 cells

Fig. 3. The effects of GEN on autophagy in DEX-treated osteoblasts in vivo and in vitro. The immunohistochemical changes (× 40 magnification) of P62 (A, B), LC3-II (A, C), and Beclin-1 (A, D) were analyzed. The proteins expression of P62 (E, F), LC3-II (E, G), and Beclin-1 (E, H) in DEX-treated MC3T3-E1 cells were determined by western blot. * *P˂0.01. NC, negative control group; DEX, model rat group treated with DEX (5 mg/kg).
7). Exosomes derived from miR-26a-modified MSCs promote axonal regeneration via the PTEN/AKT/mTOR pathway following spinal cord injury. Stem Cell Research & Therapy (PubMed: 33820561) [IF=7.5]

Application: WB    Species: rat    Sample: PC12 cells

FIGURE S2 | miR-26a-overexpressing exosomes inhibited autophagic activity and promoted axonal generation in PC12 cells. (a) The ability of Exos-26a to generate neurofilament (red fluorescent dye) in PC12 cells, which could be reversed by rapamycin. (b, c) Representative images of western blots used to determine the expression levels of NF, mTOR, p-mTOR, AMPK, p-AMPK, S6K, p-S6K, ULK1, p-ULK1, and p62 and semiquantification of the data. RAP indicates miR-26a exosome and rapamycin (100 nM) treatment for 48 h before lysis. *P < 0.05, **P < 0.01, ***P < 0.001 compared with the control group by t test or ANOVA. #P < 0.05 and ##P < 0.01 compared with the RAP group by t test. n = 3 for each group.
8). Activation of aldehyde dehydrogenase-2 improves ischemic random skin flap survival in rats. Frontiers in Immunology (PubMed: 37441072) [IF=7.3]

Application: IF/ICC    Species: Mouse    Sample:

Figure 10 Immunofluorescence images of Beclin-1 (A), p62 (B), and LC3(C) were captured under fluorescence microscopy, and the relative fluorescence intensity was calculated. All images were obtained at identical magnification, ×200, scale bar = 50 μm. Data are represented as mean ± SEM (n = 3). **P
9). mTOR deletion ameliorates CD4+ T cell apoptosis during sepsis by improving autophagosome-lysosome fusion. APOPTOSIS (PubMed: 35435531) [IF=7.2]

Application: WB    Species: mouse    Sample: CD4+T cells

Fig. 1 | Mammalian target of rapamycin (mTOR) activity and autophagic fux in CD4+T cells. (A–E) Protein levels of phospho (p)-mTOR,p-P70 S6 kinase (S6K), LC3II/I and p62/sequestome 1 (SQSTM1) in CD4+T cells were quantifed by western blotting in wild-type (WT) mice, mTOR knockout via Cre expression under the control of lymphocyte-specifc protein tyrosine kinase (Lck) (Lck-mTOR) cecal ligation and puncture (CLP) mice, and tuberous sclerosis complex 1(TSC1) knockout (Lck-TSC1)+CLP mice. The amount of each protein level was normalized to that of β-actin. Means±standard deviations (SDs) of four mice per group are shown. **P<0.01, ***P<0.001,****P<0.0001.
10). A Potent Protective Effect of Baicalein on Liver Injury by Regulating Mitochondria-Related Apoptosis. APOPTOSIS (PubMed: 32409930) [IF=7.2]

Application: WB    Species: human    Sample: L02 cells

Fig. 2| Knockdown of MARCH5 sensitizes L02 cells to hepatotoxicity induced by oxidative stress. d The expression levels of apoptotic and autophagy related proteins were detected by western blotting. e The data analysis of blot densitometry. All of the data was expressed as the mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001
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