Anti-Smad3 (phospho S423 + S425) antibody (ab51451)

Overview

  • Product nameAnti-Smad3 (phospho S423 + S425) antibody
    See all Smad3 primary antibodies
  • Description
    Rabbit polyclonal to Smad3 (phospho S423 + S425)
  • Specificityab51451 is specific for the phosphorylated form of the protein although minimal reactivity with nonphosphorylated human Smad3 can be observed by ELISA and WB. ab51451 also reacts with phosphorylated Smad1 and Smad5 but negligible reactivity is seen against other phosphorylated Smad family members.
  • Tested applicationsIHC-P, WB, ELISA, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Chicken, Human
    Predicted to work with: Rat, Cow, Pig, Xenopus laevis, Zebrafish
  • Immunogen

    Synthetic peptide: PSIRCSSpVSp, corresponding to amino acids 417-425 of Human Smad3

Properties

Applications

Our Abpromise guarantee covers the use of ab51451 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
IHC-P Use at an assay dependent dilution. PubMed: 20519446
WB 1/2000 - 1/20000. Predicted molecular weight: 48 kDa. Stimulation with 2 ng/ml TGF-beta for 1 hour is suggested.
ELISA 1/25000 - 1/100000.
ICC/IF Use at an assay dependent dilution.

Target

  • FunctionReceptor-regulated SMAD (R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta (transforming growth factor) and activin type 1 receptor kinases. Binds the TRE element in the promoter region of many genes that are regulated by TGF-beta and, on formation of the SMAD3/SMAD4 complex, activates transcription. Also can form a SMAD3/SMAD4/JUN/FOS complex at the AP-1/SMAD site to regulate TGF-beta-mediated transcription. Has an inhibitory effect on wound healing probably by modulating both growth and migration of primary keratinocytes and by altering the TGF-mediated chemotaxis of monocytes. This effect on wound healing appears to be hormone-sensitive. Regulator of chondrogenesis and osteogenesis and inhibits early healing of bone fractures. Positively regulates PDPK1 kinase activity by stimulating its dissociation from the 14-3-3 protein YWHAQ which acts as a negative regulator.
  • Involvement in diseaseColorectal cancer
    Loeys-Dietz syndrome 3
  • Sequence similaritiesBelongs to the dwarfin/SMAD family.
    Contains 1 MH1 (MAD homology 1) domain.
    Contains 1 MH2 (MAD homology 2) domain.
  • DomainThe MH1 domain is required for DNA binding. Also binds zinc ions which are necessary for the DNA binding.
    The MH2 domain is required for both homomeric and heteromeric interactions and for transcriptional regulation. Sufficient for nuclear import.
    The linker region is required for the TGFbeta-mediated transcriptional activity and acts synergistically with the MH2 domain.
  • Post-translational
    modifications
    Phosphorylated on serine and threonine residues. Enhanced phosphorylation in the linker region on Thr-179, Ser-204 and Ser-208 on EGF and TGF-beta treatment. Ser-208 is the main site of MAPK-mediated phosphorylation. CDK-mediated phosphorylation occurs in a cell-cycle dependent manner and inhibits both the transcriptional activity and antiproliferative functions of SMAD3. This phosphorylation is inhibited by flavopiridol. Maximum phosphorylation at the G(1)/S junction. Also phosphorylated on serine residues in the C-terminal SXS motif by TGFBR1 and ACVR1. TGFBR1-mediated phosphorylation at these C-terminal sites is required for interaction with SMAD4, nuclear location and transactivational activity, and appears to be a prerequisite for the TGF-beta mediated phosphorylation in the linker region. Dephosphorylated in the C-terminal SXS motif by PPM1A. This dephosphorylation disrupts the interaction with SMAD4, promotes nuclear export and terminates TGF-beta-mediated signaling. Phosphorylation at Ser-418 by CSNK1G2/CK1 promotes ligand-dependent ubiquitination and subsequent proteasome degradation, thus inhibiting SMAD3-mediated TGF-beta responses. Phosphorylated by PDPK1.
    Acetylation in the nucleus by EP300 in the MH2 domain regulates positively its transcriptional activity and is enhanced by TGF-beta.
    Ubiquitinated. Monoubiquitinated, leading to prevent DNA-binding. Deubiquitination by USP15 alleviates inhibition and promotes activation of TGF-beta target genes.
  • Cellular localizationCytoplasm. Nucleus. Cytoplasmic and nuclear in the absence of TGF-beta. On TGF-beta stimulation, migrates to the nucleus when complexed with SMAD4. Through the action of the phosphatase PPM1A, released from the SMAD2/SMAD4 complex, and exported out of the nucleus by interaction with RANBP1. Co-localizes with LEMD3 at the nucleus inner membrane. MAPK-mediated phosphorylation appears to have no effect on nuclear import. PDPK1 prevents its nuclear translocation in response to TGF-beta.
  • Information by UniProt
  • Database links
  • Alternative names
    • DKFZP586N0721 antibody
    • DKFZp686J10186 antibody
    • hMAD 3 antibody
    • hMAD-3 antibody
    • hSMAD3 antibody
    • HSPC193 antibody
    • HST17436 antibody
    • JV15 2 antibody
    • JV15-2 antibody
    • JV152 antibody
    • LDS1C antibody
    • LDS3 antibody
    • MAD (mothers against decapentaplegic Drosophila) homolog 3 antibody
    • MAD homolog 3 antibody
    • Mad homolog JV15 2 antibody
    • Mad protein homolog antibody
    • MAD, mothers against decapentaplegic homolog 3 antibody
    • MAD3 antibody
    • MADH 3 antibody
    • MADH3 antibody
    • MGC60396 antibody
    • Mothers against decapentaplegic homolog 3 antibody
    • Mothers against DPP homolog 3 antibody
    • SMA and MAD related protein 3 antibody
    • SMAD 3 antibody
    • SMAD antibody
    • SMAD family member 3 antibody
    • SMAD, mothers against DPP homolog 3 antibody
    • SMAD3 antibody
    • SMAD3_HUMAN antibody
    see all

Anti-Smad3 (phospho S423 + S425) antibody images

  • All lanes : Anti-Smad3 (phospho S423 + S425) antibody (ab51451) at 1/2000 dilution

    Lane 1 : MEF lysate (control)
    Lane 2 : MEF lysate, after stimulation with 2 ng/ml of TGF-ß for 1 hour
    Lane 3 : MEF-/-Smad3 lysate (control)
    Lane 4 : MEF-/-Smad3 lysate, after stimulation with 2 ng/ml of TGF-ß for one hour
    Lane 5 : Hep3B lysate (control)
    Lane 6 : Hep3B lysate, after stimulation with 2 ng/ml of TGF-ß for one hour
    Lane 7 : HEK 293 lysate (control)


    Predicted band size : 48 kDa
    Image courtesy of Ying Zhang, NIH, CCR, Bethesda, MD.
  • ab51451 at 2.5 µg/ml staining epidermal keratinocytes at 40X.  This image shows strong nuclear staining with localization of the antibody as the precipated red siganl and a hematoxylin purple nuclear counterstain.

  • ab51451 staining Smad3 (phospho S423 + S425) in PC3 and LnCAP cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 70% ethanol and blocked for 1 hour at 26°C. Samples were incubated with primary antibody (1/150) for 16 hours at 4°C. An IRDye® 800CW-conjugated goat anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody. NC-PC3 - negative control PC3 cells, NC-LnCAP - negative control LnCAP cells.

    See Abreview

References for Anti-Smad3 (phospho S423 + S425) antibody (ab51451)

This product has been referenced in:
  • Ying Y  et al. Targeted Deletion of p53 in the Proximal Tubule Prevents Ischemic Renal Injury. J Am Soc Nephrol N/A:N/A (2014). Mouse . Read more (PubMed: 24854277) »
  • Kosla J  et al. Effective myofibroblast dedifferentiation by concomitant inhibition of TGF-ß signaling and perturbation of MAPK signaling. Eur J Cell Biol N/A:N/A (2013). Read more (PubMed: 24315689) »

See all 7 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Blocking step Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 26°C
Sample Human Cell (PC3 and LnCAP)
Specification PC3 and LnCAP
Permeabilization Yes - 70% Ethanol
Fixative Paraformaldehyde
Username

Mr. FRANCESCO ELIA MARINO

Verified customer

Submitted May 02 2014

Application Western blot
Loading amount 60 µg
Gel Running Conditions Reduced Non-Denaturing (Native) (12)
Sample Mouse Tissue lysate - whole (Mouse gastrocnemius)
Specification Mouse gastrocnemius
Blocking step Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 26°C
Username

Mr. FRANCESCO ELIA MARINO

Verified customer

Submitted Mar 26 2014

Thank you for calling Abcam earlier today.

As we talked about on the telephone, I am sending you ab52903 as a free of charge replacement. Your new order/reference number is********.

Please let me know if there is anything else I can...

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Thank you for your reply.

The only other phospho-smad3 (S423+S425) we have are:

ab11825 - http://www.abcam.com/Smad3-phospho-S423-S425-antibody-ab118825.html

ab52903 - http://www.abcam.com/Smad3-phospho-S423-S425-antibody...

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Thank you very much for contacting Abcam.

The isotype control that I would recommend for your flow cytometry experiments is:

http://www.abcam.com/Rabbit-IgG-ChIP-Grade-ab37415.html



Below is the infromation about...

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