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Sequence-specific antibodies to connexins block intercellular calcium signaling through gap junctions
S Boitano1, E R Dirksen, W H Evans
1Department of Zoology and Physiology, University of Wyoming, Laramie 82071-3166, USA. sboitano@uwyo.edu
Cell Calcium
|May 7, 1998
Summary
Mechanical stimulation triggers calcium waves in airway cells, likely through connexin 32 gap junctions. Specific antibodies targeting connexin 32 blocked these intercellular calcium waves, confirming its role.
Area of Science:
- Cell Biology
- Molecular Biology
- Physiology
Background:
- Mechanical stimulation of airway epithelial cells causes intercellular calcium (Ca2+) waves.
- Gap junctions are hypothesized to mediate these Ca2+ waves.
- Connexins form gap junctions, facilitating direct cell-to-cell communication.
Purpose of the Study:
- To directly investigate the role of gap junctions in mechanically induced intercellular Ca2+ wave propagation.
- To identify the specific connexin proteins involved in this signaling pathway.
Main Methods:
- Primary airway epithelial cell cultures were mechanically stimulated.
- Sequence-specific antibodies targeting different regions of connexin 32 (Cx32) were introduced via electroporation.
- Antibody effects on intercellular Ca2+ wave propagation were assessed, including competitive reversal experiments with peptides.
Main Results:
- Antibodies targeting the cytosolic loop and carboxyl tail of connexin 32 (Cx32) significantly inhibited Ca2+ wave propagation.
- The inhibitory effect of one antibody (Des 1) was competitively reversed by a specific peptide, supporting target interaction.
- Antibodies against other connexin regions (extracellular loop, amino terminus) and other connexins (Cx43, Cx26) did not affect Ca2+ wave propagation.
Conclusions:
- Mechanically induced Ca2+ waves in airway epithelial cells are propagated via gap junctions.
- Connexin 32 (Cx32) is the primary gap junction protein responsible for mediating this intercellular communication.
- Specific domains of Cx32 are crucial for the propagation of these mechanically stimulated Ca2+ waves.