Related Experiment Video
Updated: Aug 1, 2026

Detection of Signaling Effector-Complexes Downstream of BMP4 Using in situ PLA, a Proximity Ligation Assay
Published on: March 3, 2011
Characterization of functional domains within Smad4/DPC4
M P de Caestecker1, P Hemmati, S Larisch-Bloch
1Laboratory of Chemoprevention, NCI, National Institutes of Health, Bethesda, Maryland 20892-5055, USA. decaestm@dce41.nci.nih.gov
Abstract:
Smad proteins are a family of highly conserved, intracellular proteins that signal cellular responses downstream of transforming growth factor-beta (TGF-beta) family serine/threonine kinase receptors. One of these molecules, Smad4, originally identified as the candidate tumor suppressor gene dpc-4, reconstitutes TGF-beta- and activin-dependent transcriptional responses in Smad4 null cell lines and interacts in a ligand-dependent manner with other Smad family members in both TGF-beta, activin, and bone morphogenetic protein-2/-4 pathways. Here, we used an assay based on the restoration of ligand-dependent transcriptional responses in a Smad4 null cell line to characterize functional domain structures within Smad4. We showed that restoration of TGF-beta-induced transcriptional responses by Smad4 was inhibited by co-transfection with a kinase dead TGF-beta type II receptor and that constitutive activation was blocked with TGF-beta neutralizing antibodies, confirming the essential role of Smad4 in TGF-beta signaling. Using a series of Smad4 mutation, deletion, and Smad1/Smad4 chimera constructs we identified a 47-amino acid deletion within the middle-linker region of Smad4 that is essential for the mediation of signaling responses. In addition, we showed that the NH2-terminal domain of Smad4 augments ligand-dependent activation associated with the middle-linker region, indicating that there is a distinct ligand-response domain within the N terminus of this molecule.
More Related Videos
14:47Identification of MyoD Interactome Using Tandem Affinity Purification Coupled to Mass Spectrometry
Published on: May 17, 2016
09:34Identification of Modified Histones as Binding Substrates of Human Spindlin Family Member 4 (SPIN4) by Peptide Arrays and Native Nucleosome Pulldown
Published on: March 27, 2026
Related Concept Videos
Conservation of Protein Domains Over Different Proteins
A limited set of protein domains often duplicate and recombine during evolution. These domains can be organized in different combinations to form...
Master Transcription Regulators
Membrane Domains
Protein Domains
The membrane comprises a group of distinct proteins responsible for carrying out a cell's specific function. For example, the plasma membrane of the human sperm, or a single germ cell, contains a unique set of proteins in the anterior...
Assembly of Signaling Complexes
Interaction domains in cell signaling
Interaction domains recognize exposed features of their binding partners containing post-translationally modified sequences,...
The JAK-STAT Signaling Pathway
TGF - β Signaling Pathway