Phospho-Smad2 (Ser465/467) Ant

3101:

Western Blotting

Phospho-Smad2 (Ser465/467) Antibody detects endogenous levels of Smad2 only when dually phosphorylated at Ser465 and Ser467, and may detect phosphorylated Smad3 at its equivalent site. This antibody does not cross-react with other Smad-related proteins.

Polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser465/467 of human Smad2. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western blot analysis of extracts from HepG2 cells treated with TGFbeta for the indicated times, using Phospho-Smad2 (Ser465/467) Antibody (upper) or Smad2 antibody (lower).

Members of the Smad family of signal transduction molecules are components of a critical intracellular pathway that transmit TGF-β signals from the cell surface into the nucleus. Three distinct classes of Smads have been defined: the receptor-regulated Smads (R-Smads), which include Smad1, 2, 3, 5, and 8; the common-mediator Smad (co-Smad), Smad4; and the antagonistic or inhibitory Smads (I-Smads), Smad6 and 7 (1-5). Activated type I receptors associate with specific R-Smads and phosphorylate them on a conserved carboxy-terminal SSXS motif. The phosphorylated R-Smad dissociates from the receptor and forms a heteromeric complex with the co-Smad (Smad4), allowing translocation of the complex to the nucleus. Once in the nucleus, Smads can target a variety of DNA binding proteins to regulate transcriptional responses (6-8).

Following stimulation by TGF-beta, Smad2 and Smad3 become phosphorylated at their carboxyl termini by the receptor kinase (serines 465 and 467 on Smad2; serines 423 and 425 on Smad3) by TbetaR-I (9-11). Following phosphorylation, Smad2 and Smad3 form a heteromeric complex with the co-smad family member Smad4. These complexes are translocated to the nucleus where they bind DNA and regulate gene transcription.

1. Heldin, C.H. et al. (1997) Nature 390, 465-471.
2. Attisano, L. and Wrana, J.L. (1998) Curr. Opin. Cell Biol. 10, 188-194.
3. Derynck, R. et al. (1998) Cell 95, 737-740.
4. Massague, J. (1998) Annu. Rev. Biochem. 67, 753-791.
5. Whitman, M. (1998) Genes Dev. 12, 2445-2462.
6. Wu, G. et al. (2000) Science 287, 92-97.
7. Attisano, L. and Wrana, J.L. (2002) Science 296, 1646-1647.
8. Moustakas, A. et al. (2001) J. Cell Sci. 114, 4359-4369.
9. Abdollah, S. et al. (1997) J. Biol. Chem. 272, 27678-27685.
10. Souchelnytskyi, S. et al. (1997) J. Biol. Chem. 272, 28107-28115.
11. Liu, X. et al. (1997) Proc. Natl. Acad. Sci. 94, 10669-10674.

• Watabe, T. et al. (2003) TGF-beta receptor kinase inhibitor enhances growth and integrity of embryonic stem cell-derived endothelial cells. J. Cell Biol. 163, 1303-1311. Applications: Western Blotting
• Stove, C. et al. (2004) Melanoma cells secrete follistatin, an antagonist of activin-mediated growth inhibition. Oncogene 23, 5330-5339. Applications: Western Blotting
• Tomita, M. et al. (2004) The Kaposi's sarcoma-associated herpesvirus K-bZIP protein represses transforming growth factor beta signaling through interaction with CREB-binding protein. Oncogene 23, 8272-8281. Applications: Western Blotting
• Kondo, M. et al. (2004) Cell Death Differ 11, 1092-101. Applications: Western Blotting
• Nagata, M. et al. (2006) Genes Cells 11, 1267-80. Applications: Western Blotting
• Ehata, S. et al. (2007) Cancer Res 67, 9694-703. Applications: Western Blotting
• Louafi, F. et al. (2010) J Biol Chem 285, 41328-36. Applications: Western Blotting
• Kroening, S. et al. (2010) Am J Physiol Renal Physiol 298, F796-806. Applications: Western Blotting

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For Research Use Only. Not For Use In Diagnostic Procedures.