Related Experiment Video

Updated: Jul 30, 2026

Real-time Live Imaging of T-cell Signaling Complex Formation
10:31

Real-time Live Imaging of T-cell Signaling Complex Formation

Published on: June 23, 2013

Overcoming the signaling defect of Lyn-sequestering, signal-curtailing FcepsilonRI dimers: aggregated dimers can

M Lara1, E Ortega, I Pecht

  • 1Departamento de Inmunologia, Instituto de Investigaciones Biomedicas, Universidad Nacional Autónoma de Mexico, Mexico DF, Mexico.

Journal of Immunology (Baltimore, Md. : 1950)
|October 10, 2001
PubMed

Clustering the tetrameric (alphabetagamma(2)) IgE receptor, FcepsilonRI, on basophils and mast cells activates the Src-family tyrosine kinase, Lyn, which phosphorylates FcepsilonRI beta and gamma subunit tyrosines, creating binding sites for the recruitment and activation of Syk. We reported previously that FcepsilonRI dimers formed by a particular anti-FcepsilonRI alpha mAb (H10) initiate signaling through Lyn activation and FcepsilonRI subunit phosphorylation, but cause only modest activation of Syk and little Ca(2+) mobilization and secretion. Curtailed signaling was linked to the formation of unusual, detergent-resistant complexes between Lyn and phosphorylated receptor subunits. Here, we show that H10-FcepsilonRI multimers, induced by adding F(ab')(2) of goat anti-mouse IgG to H10-treated cells, support strong Ca(2+) mobilization and secretion. Accompanying the recovery of signaling, H10-FcepsilonRI multimers do not form stable complexes with Lyn and do support the phosphorylation of Syk and phospholipase Cgamma2. Immunogold electron microscopy showed that H10-FcepsilonRI dimers colocalize preferentially with Lyn and are rarely within the osmiophilic "signaling domains" that accumulate FcepsilonRI and Syk in Ag-treated cells. In contrast, H10-FcepsilonRI multimers frequently colocalize with Syk within osmiophilic patches. In sucrose gradient centrifugation analyses of detergent-extracted cells, H10-treated cells show a more complete redistribution of FcepsilonRI beta from heavy (detergent-soluble) to light (Lyn-enriched, detergent-resistant) fractions than cells activated with FcepsilonRI multimers. We hypothesize that restraints imposed by the particular orientation of H10-FcepsilonRI dimers traps them in signal-initiating Lyn microdomains, and that converting the dimers to multimers permits receptors to dissociate from Lyn and redistribute to separate membrane domains that support Syk-dependent signal propagation.

More Related Videos

SorLA and CLC:CLF-1-dependent Downregulation of CNTFRα as Demonstrated by Western Blotting, Inhibition of Lysosomal Enzymes, and Immunocytochemistry10:16

SorLA and CLC:CLF-1-dependent Downregulation of CNTFRα as Demonstrated by Western Blotting, Inhibition of Lysosomal Enzymes, and Immunocytochemistry

Published on: January 6, 2017

Using In Vitro Fluorescence Resonance Energy Transfer to Study the Dynamics Of Protein Complexes at a Millisecond Time Scale10:50

Using In Vitro Fluorescence Resonance Energy Transfer to Study the Dynamics Of Protein Complexes at a Millisecond Time Scale

Published on: March 14, 2019

Related Concept Videos

Assembly of Signaling Complexes01:30

Assembly of Signaling Complexes

Multiprotein signaling complexes are formed in a dynamic process involving protein-protein interactions at the cytoplasmic domain of transmembrane receptors or enzymatic and non-enzymatic proteins associated with the receptor. These complexes ensure the activation and propagation of intracellular signals that regulate cell functions.
Interaction domains in cell signaling
Interaction domains recognize exposed features of their binding partners containing post-translationally modified sequences,...
Interactions Between Signaling Pathways01:19

Interactions Between Signaling Pathways

Signaling cascades usually lack linearity. Multiple pathways interact and regulate one another, allowing cells to integrate and respond to diverse environmental stimuli.
Convergence and divergence, and cross-talk between signaling pathways
Two distinct signaling pathways can converge on a single functional unit, which may either be a single protein or a complex of proteins. The response is either functionally distinct or synergistic between the two pathways but different from the response...
Amplifying Signals via Enzymatic Cascade01:22

Amplifying Signals via Enzymatic Cascade

When a ligand binds to a cell-surface receptor, the receptor's intracellular domain changes shape, which may either activate its enzyme function or allow its binding to other molecules. The initial signal is amplified by most signal transduction pathways. This means that a single ligand molecule can activate multiple molecules of a downstream target. Proteins that relay a signal are most commonly phosphorylated at one or more sites, activating or inactivating the protein. Kinases catalyze the...
Receptor Tyrosine Kinases01:26

Receptor Tyrosine Kinases

Receptor tyrosine kinases or RTKs are membrane-bound receptors that phosphorylate specific tyrosine on protein substrates. RTKs regulate cellular growth, differentiation, survival, and migration. They contain an extracellular ligand binding domain, a transmembrane domain, and a cytosolic tail with intrinsic kinase activity. Several extracellular signaling molecules activate RTKs in one or more ways and relay the signal downstream. Ligands such as platelet-derived growth factor (PDGF) or...
The JAK-STAT Signaling Pathway01:20

The JAK-STAT Signaling Pathway

Several cytokine receptors have tightly bound Janus kinase or JAK proteins attached at their cytosolic tail. Small signaling molecules such as cytokines, growth hormones, or prolactins bind to the cytokine receptors and initiate their dimerization. The dimerization brings the cytosolic JAKs together that trans-phosphorylate and activates each other. The activated JAKs now phosphorylate cytosolic tails of the cytokine receptors, which serve as binding sites for adaptor proteins such as  SH2...
Intracellular Signaling Affects Focal Adhesions01:17

Intracellular Signaling Affects Focal Adhesions

Integrins act both as extracellular input receivers and as intracellular processing activators. As their name suggests, integrins are entirely integrated into the membrane structure. Their hydrophobic membrane-spanning regions interact with the phospholipid bilayer's hydrophobic region. These membrane receptors provide extracellular attachment sites for effectors like hormones and growth factors. They activate intracellular response cascades when their effectors are bound and active.
Some...