Phospho-smad1/5/8
Phospho-smad1/5/8 - Related Videos
10:52
Radiolabeling and Quantification of Cellular Levels of Phosphoinositides by High Performance Liquid Chromatography-coupled Flow Scintillation
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Cited by 21 •
2016
Phosphoinositides are signaling lipids whose relative abundance rapidly changes in response to various stimuli. This article describes a method to measure the abundance of phosphoinositides by metabolically labeling cells with 3H-myo-inositol, followed by extraction and deacylation. Extracted glycero-inositides are then separated by high-performance liquid chromatography and quantified by flow scintillation.
01:42
Phosphoinositides and PIPs
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2023
Phosphoinositides are a group of phospholipids containing a glycerol backbone with two fatty acid chains and a phosphate attached to a myoinositol sugar ring. The inositol head group extends into the cytoplasm, where it is modified by adding phosphate groups to form phosphatidylinositol phosphates or PIPs. Different phosphoinositides are synthesized and recruited on the cytosolic face of the plasma membrane. The localization of specific phosphoinositides concentrated in separate membrane...
10:58
PIP-on-a-chip: A Label-free Study of Protein-phosphoinositide Interactions
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Cited by 7 •
2017
Here we present a supported lipid bilayer in the context of a microfluidic platform to study protein-phosphoinositide interactions using a label-free method based on pH modulation.
06:23
pH Modulation Assay on Supported Lipid Bilayers to Detect Protein-Phosphoinositide Interactions
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2025
In this video, we demonstrate protein-phosphoinositide interactions in a microfluidic pattern using the pH modulation assay, using pH-sensitive fluorescent dye tagged to phosphatidylethanolamine lipid of a supported lipid bilayer, called a microfluidic platform. The modulations in pH after protein binding to phospholipid cause fluorescence quenching.
08:07
Identification of Inositol Phosphate or Phosphoinositide Interacting Proteins by Affinity Chromatography Coupled to Western Blot or Mass Spectrometry
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Cited by 13 •
2019
This protocol focuses on the identification of proteins that bind to inositol phosphates or phosphoinositides. It uses affinity chromatography with biotinylated inositol phosphates or phosphoinositides that are immobilized via streptavidin to agarose or magnetic beads. Inositol phosphate or phosphoinositide binding proteins are identified by Western blotting or mass spectrometry.
FAQs
Phosphorylation acts as the key intracellular switch that converts receptor stimulation into a transcriptional response. Once modified by BMP receptor serine/threonine kinases, SMAD1, SMAD5, or SMAD8 can associate with SMAD4 and move into the nucleus. This sequence links an extracellular BMP signal to changes in gene expression that may affect cell fate and tissue organization.
Phosphorylated SMAD1/5/8 do not function only as isolated signaling components. Their association with SMAD4 supports nuclear entry, where the signaling complex can influence gene expression. Consequently, detecting phosphorylation identifies activation of an important pathway step, while the biological response depends on subsequent complex formation and nuclear activity.
These cell-surface receptors provide the direct enzymatic step that transfers BMP pathway activation to intracellular SMAD proteins. After BMP ligands bind the receptors, receptor kinase activity phosphorylates SMAD1, SMAD5, and SMAD8. The resulting signal can then proceed through SMAD4 association and nuclear entry, making receptor-mediated phosphorylation a useful indicator of pathway engagement.
Western blotting, immunofluorescence, and immunohistochemistry provide complementary ways to assess pathway activity. Western blotting supports measurement of the detected phospho-SMAD1/5/8 signal in a sample, whereas immunofluorescence and immunohistochemistry can show where the signal occurs in cells or tissue. Method selection therefore depends on whether signal assessment or spatial localization is most relevant.
The measurement helps researchers evaluate whether BMP pathway signaling is active in the examined biological context. Interpreting the signal alongside the sample type can connect extracellular BMP stimulation with intracellular pathway engagement, rather than treating phosphorylation as an isolated molecular event. This makes the readout useful for studying changes in cell behavior, differentiation, or tissue organization.
This analysis is relevant when investigators need to examine BMP signaling during development, stem cell differentiation, tissue formation, or disease modeling. In these settings, the readout can help relate pathway activity to changes in cell fate and tissue organization. It is especially informative when comparing how different biological conditions influence BMP-dependent signaling.
Phospho-SMAD1/5/8 provides a molecular measurement that can be related to larger biological processes governed by BMP pathways. Because activated SMAD complexes influence gene expression, their detection can help connect extracellular signaling with developmental events, differentiation programs, and tissue formation. In disease models, the same approach can clarify how altered pathway activity relates to disrupted tissue organization.