Overview

  • Product name

    Anti-Smad3 antibody - ChIP Grade
    See all Smad3 primary antibodies
  • Description

    Rabbit polyclonal to Smad3 - ChIP Grade
  • Host species

    Rabbit
  • Tested applications

    Suitable for: IHC-P, WB, ICC/IF, IP, ChIPmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
    Predicted to work with: Chicken, Cow, Dog, Pig, Xenopus laevis, Rhesus monkey
  • Immunogen

    Recombinant fragment within Human Smad3 (internal sequence). The exact sequence is proprietary.
    Database link: P84022

  • Positive control

    • WB: Jurkat whole cell lysate; Mouse and rat brain tissue extracts. IP: HeLa whole cell lysate. IHC-P: Human gastric cancer tissue. ICC/IF: HeLa cells. ChIP: Chromatin from HeLa whole cell lysate.

Properties

Applications

Our Abpromise guarantee covers the use of ab227223 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 1/100 - 1/1000.
WB 1/1000 - 1/10000. Predicted molecular weight: 48 kDa.
ICC/IF 1/100 - 1/1000.
IP 1/100 - 1/500.
ChIP Use at an assay dependent concentration.

Target

  • Function

    Receptor-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 disease

    Colorectal cancer
    Loeys-Dietz syndrome 3
  • Sequence similarities

    Belongs to the dwarfin/SMAD family.
    Contains 1 MH1 (MAD homology 1) domain.
    Contains 1 MH2 (MAD homology 2) domain.
  • Domain

    The 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.
    Poly-ADP-ribosylated by PARP1 and PARP2. ADP-ribosylation negatively regulates SMAD3 transcriptional responses during the course of TGF-beta signaling.
  • Cellular localization

    Cytoplasm. Nucleus. Cytoplasmic and nuclear in the absence of TGF-beta. On TGF-beta stimulation, migrates to the nucleus when complexed with SMAD4 (PubMed:15799969). Through the action of the phosphatase PPM1A, released from the SMAD2/SMAD4 complex, and exported out of the nucleus by interaction with RANBP1 (PubMed:16751101, PubMed:19289081). Co-localizes with LEMD3 at the nucleus inner membrane (PubMed:15601644). MAPK-mediated phosphorylation appears to have no effect on nuclear import (PubMed:19218245). PDPK1 prevents its nuclear translocation in response to TGF-beta (PubMed:17327236).
  • 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

Images

  • Anti-Smad3 antibody - ChIP Grade (ab227223) at 1/5000 dilution + Jurkat (human T cell leukemia cell line from peripheral blood) whole cell lysate at 30 µg

    Secondary
    HRP-conjugated anti-rabbit IgG

    Predicted band size: 48 kDa



    7.5% SDS-PAGE gel.

  • Anti-Smad3 antibody - ChIP Grade (ab227223) at 1/1000 dilution + Rat brain tissue extract at 50 µg

    Secondary
    HRP-conjugated anti-rabbit IgG

    Predicted band size: 48 kDa



    10% SDS-PAGE gel.

  • Anti-Smad3 antibody - ChIP Grade (ab227223) at 1/5000 dilution + Mouse brain tissue extract at 50 µg

    Secondary
    HRP-conjugated anti-rabbit IgG

    Predicted band size: 48 kDa



    10% SDS-PAGE gel.

  • Paraffin-embedded human gastric cancer tissue stained for Smad3 using ab227223 at 1/500 dilution in immunohistochemical analysis.

  • Smad3 was immunoprecipitated from HeLa (human epitheilal cell line from cervix adenocarcinoma) whole cell lysate with 2 µg of ab227223. Western blot was performed from the immunoprecipitate using ab227223 at 1/1000 dilution. Anti-rabbit IgG was used as secondary antibody.

    Lane 1: HeLa whole cell lysate 50 µg (Input). 

    Lane 2: Preimmune rabbit IgG instead of ab227223 in HeLa whole cell lysate.

    Lane 3: ab227223 IP in HeLa whole cell lysate. 

     

  • PFA-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained for Smad3 (green) using ab227223 at 1/200 dilution in ICC/IF.

    Nuclear Counterstain: Hoechst 33342.

  • Chromatin was prepared from Hela (human epithelial cell line from cervix adenocarcinoma) whole cell lysate. The ChIP was performed with 5 μg of preimmune rabbit IgG (Lane 1) and 5 µg of ab227223 (Lane 2). The immunoprecipitated DNA was detected by PCR with a primer set targeting to the smad7 promoter.

References

This product has been referenced in:

  • Xu Q  et al. All-trans retinoic acid inhibits lipopolysaccharide-induced inflammatory responses in bovine adipocytes via TGFß1/Smad3 signaling pathway. BMC Vet Res 15:48 (2019). Read more (PubMed: 30709353) »
See 1 Publication for this product

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