Anti-SP1 (phospho T453) antibody (ab59257)

Rabbit polyclonal SP1 (phospho T453) antibody. Validated in WB, ELISA, IHC, ChIP, ICC/IF and tested in Mouse, Rat, Human. Cited in 18 publication(s). Independently reviewed in 6 review(s).

Overview

  • Product name

    Anti-SP1 (phospho T453) antibody
    See all SP1 primary antibodies
  • Description

    Rabbit polyclonal to SP1 (phospho T453)
  • Host species

    Rabbit
  • Specificity

    ab59257 detects endogenous levels of SP1 only when phosphorylated at threonine 453.
  • Tested applications

    Suitable for: ELISA, IHC-P, WB, ChIP, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic phosphopeptide derived from human SP1 around the phosphorylation site of threonine 453 (I-R-TP-P-T).

  • Positive control

    • IHC-P: Human brain tissue. WB: A549 cell extracts.

Properties

Applications

Our Abpromise guarantee covers the use of ab59257 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ELISA 1/5000.
IHC-P Use at an assay dependent concentration.
WB 1/500 - 1/1000. Detects a band of approximately 90 kDa (predicted molecular weight: 81 kDa).
ChIP Use at an assay dependent concentration. PubMed: 20375015
ICC/IF Use at an assay dependent concentration. PubMed: 21115498

Target

  • Function

    Transcription factor that can activate or repress transcription in response to physiological and pathological stimuli. Binds with high affinity to GC-rich motifs and regulates the expression of a large number of genes involved in a variety of processes such as cell growth, apoptosis, differentiation and immune responses. Highly regulated by post-translational modifications (phosphorylations, sumoylation, proteolytic cleavage, glycosylation and acetylation). Binds also the PDGFR-alpha G-box promoter. May have a role in modulating the cellular response to DNA damage. Implicated in chromatin remodeling. Plays a role in the recruitment of SMARCA4/BRG1 on the c-FOS promoter. Plays an essential role in the regulation of FE65 gene expression. In complex with ATF7IP, maintains telomerase activity in cancer cells by inducing TERT and TERC gene expression.
  • Tissue specificity

    Up-regulated in adenocarcinomas of the stomach (at protein level).
  • Sequence similarities

    Belongs to the Sp1 C2H2-type zinc-finger protein family.
    Contains 3 C2H2-type zinc fingers.
  • Post-translational
    modifications

    Phosphorylated on multiple serine and threonine residues. Phosphorylation is coupled to ubiquitination, sumoylation and proteolytic processing. Phosphorylation on Ser-59 enhances proteolytic cleavage. Phosphorylation on Ser-7 enhances ubiquitination and protein degradation. Hyperphosphorylation on Ser-101 in response to DNA damage has no effect on transcriptional activity. MAPK1/MAPK3-mediated phosphorylation on Thr-453 and Thr-739 enhances VEGF transcription but, represses FGF2-triggered PDGFR-alpha transcription. Also implicated in the repression of RECK by ERBB2. Hyperphosphorylated on Thr-278 and Thr-739 during mitosis by MAPK8 shielding SP1 from degradation by the ubiquitin-dependent pathway. Phosphorylated in the zinc-finger domain by calmodulin-activated PKCzeta. Phosphorylation on Ser-641 by PKCzeta is critical for TSA-activated LHR gene expression through release of its repressor, p107. Phosphorylation on Thr-668, Ser-670 and Thr-681 is stimulated by angiotensin II via the AT1 receptor inducing increased binding to the PDGF-D promoter. This phosphorylation is increased in injured artey wall. Ser-59 and Thr-681 can both be dephosphorylated by PP2A during cell-cycle interphase. Dephosphorylation on Ser-59 leads to increased chromatin association during interphase and increases the transcriptional activity. On insulin stimulation, sequentially glycosylated and phosphorylated on several C-terminal serine and threonine residues.
    Acetylated. Acetylation/deacetylation events affect transcriptional activity. Deacetylation leads to an increase in the expression the 12(s)-lipooxygenase gene though recruitment of p300 to the promoter.
    Ubiquitinated. Ubiquitination occurs on the C-terminal proteolytically-cleaved peptide and is triggered by phosphorylation.
    Sumoylated by SUMO1. Sumoylation modulates proteolytic cleavage of the N-terminal repressor domain. Sumoylation levels are attenuated during tumorigenesis. Phosphorylation mediates SP1 desumoylation.
    Proteolytic cleavage in the N-terminal repressor domain is prevented by sumoylation. The C-terminal cleaved product is susceptible to degradation.
    O-glycosylated; contains at least 8 N-acetylglucosamine side chains. Levels are controlled by insulin and the SP1 phosphorylation states. Insulin-mediated O-glycosylation locates SP1 to the nucleus, where it is sequentially deglycosylated and phosphorylated. O-glycosylation affects transcriptional activity through disrupting the interaction with a number of transcription factors including ELF1 and NFYA. Also inhibits interaction with the HIV1 promoter. Inhibited by peroxisomome proliferator receptor gamma (PPARgamma).
  • Cellular localization

    Nucleus. Cytoplasm. Nuclear location is governed by glycosylated/phosphorylated states. Insulin promotes nuclear location, while glucagon favors cytoplasmic location.
  • Information by UniProt
  • Database links

  • Alternative names

    • SP 1 antibody
    • SP1 antibody
    • Sp1 transcription factor antibody
    • SP1_HUMAN antibody
    • Specificity protein 1 antibody
    • Transcription factor Sp1 antibody
    • TSFP 1 antibody
    • TSFP1 antibody
    see all

Images

  • All lanes : Anti-SP1 (phospho T453) antibody (ab59257) at 1/500 dilution

    Lane 1 : A549 cell extracts
    Lane 2 : A549 cell extracts with immunising phospho peptide

    Predicted band size: 81 kDa
    Observed band size: 90 kDa
    why is the actual band size different from the predicted?

  • Immunofluorescence analysis of HeLa cells, using ab59257 Antibody. The picture on the right is treated with the synthesized peptide.

  • Wild type MEF cells (2 × 107 cells) were cross-linked with formaldehyde, quenched with glycine, resuspended in SDS lysis buffer (1% SDS, 10 mm EDTA, 50 mm Tris-HCl, pH 8.0, with protease inhibitors and phosphatase inhibitors), sonicated on ice, and centrifuged at 4 °C. Supernatant (400 μl) were diluted to a final volume of 4 ml in a mixture of 9 parts dilution buffer (1% Triton X-100, 150 mm NaCl, 2 mm EDTA, 20 mm Tris-HCl, with protease inhibitors, pH 8.0) and 1 part lysis buffer.

    Mixtures were incubated with 4 μg of anti-p-Sp1 (ab 59257) or anti-Sp1 antibodies with rotating at 4 °C overnight followed with incubation with 100 μl of protein A beads with rotating at 4 °C for 4 h. After gentle centrifugation (2000 rpm), beads were resuspended in 1 ml of wash buffer (1% Triton X-100, 0.1% SDS, 150 mm NaCl, 2 mm EDTA, 20 mm Tris-HCl, with protease inhibitors, pH 8.0) and washed with wash buffer 3 times followed by one wash with a final wash buffer (1% Triton X-100, 0.1% SDS, 500 mm NaCl, 2 mm EDTA, 20 mm Tris-HCl, pH 8.0, with protease inhibitors). The immune complexes were eluted with elution buffer (1% SDS, 100 mm NaHCO3) followed by incubation with proteinase K and RNase A (500 μg/ml each) at 37 °C for 30 min. Reverse cross-links were performed by placing the tubes at 65 °C overnight. Immunoprecipitated DNA was extracted and dissolved in sterile water and Q-PCR was performed.

  • ab59257, at 1/50 dilution, staining SP1 in paraffin embedded human brain tissue by Immunohistochemistry in the absence or presence of the immunising peptide.
  • ab59257 (1:1000) Antibody detects endogenous levels of SP1 only when phosphorylated at Thr453. 

References

This product has been referenced in:

  • Román AC  et al. Histone H4 acetylation regulates behavioral inter-individual variability in zebrafish. Genome Biol 19:55 (2018). Read more (PubMed: 29695303) »
  • Lv W  et al. KIAA0101 inhibition suppresses cell proliferation and cell cycle progression by promoting the interaction between p53 and Sp1 in breast cancer. Biochem Biophys Res Commun 503:600-606 (2018). Read more (PubMed: 29902451) »
See all 18 Publications for this product

Customer reviews and Q&As

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1-6 of 6 Abreviews

Application
Immunocytochemistry
Sample
Human Cultured Cells (HEK293)
Permeabilization
Yes - Triton x-100, 0.01%
Specification
HEK293
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Dec 12 2016

Application
Western blot
Sample
Human Cell lysate - whole cell (HEK293)
Gel Running Conditions
Reduced Denaturing
Loading amount
25 µg
Specification
HEK293
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Nov 11 2016

Application
Western blot
Sample
Mouse Cell lysate - whole cell (Cardiomyocytes)
Gel Running Conditions
Reduced Denaturing
Loading amount
25 µg
Specification
Cardiomyocytes
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Sep 21 2016

Application
Western blot
Loading amount
25 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (Hep G2)
Specification
Hep G2
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Dec 29 2014

Application
Western blot
Sample
Mouse Tissue lysate - whole (Melanoma cancer)
Loading amount
30 µg
Specification
Melanoma cancer
Treatment
5gy Irradiation
Gel Running Conditions
Non-reduced Denaturing (8%)
Blocking step
Milk as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 3% · Temperature: 25°C

Ms. Seontae Kim

Verified customer

Submitted Jan 14 2013

Application
Western blot
Sample
Rat Cell lysate - nuclear (Intestine)
Loading amount
50 µg
Specification
Intestine
Treatment
DFO
Gel Running Conditions
Non-reduced Denaturing (7.5% gel SDS)
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C

Liwei Xie

Verified customer

Submitted Jun 17 2011

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