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Characterization of the gap junction protein, connexin45
J G Laing1, E M Westphale, G L Engelmann
1Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri 63110.
The Journal of Membrane Biology
|April 1, 1994
Summary
Connexin45, a gap junction protein, is expressed in BWEM cells and colocalizes with connexin43. Tumor promoter TPA affects connexin45 and connexin43 expression and phosphorylation differently.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Connexin45 is a gap junction protein forming channels with unique characteristics.
- Connexin45 expression was detected in multiple cell lines, including BWEM cells.
Purpose of the Study:
- To investigate the expression, characteristics, and regulation of connexin45 in BWEM cells.
- To compare the behavior of connexin45 with connexin43 under specific treatments.
Main Methods:
- RNA blot analysis for connexin45 expression.
- Immunofluorescence microscopy to study connexin localization.
- Metabolic labeling ([35S]-methionine, [32P]-orthophosphoric acid) and immunoprecipitation to characterize connexin45 polypeptide.
- Treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) and assessment of intercellular communication and connexin phosphorylation.
Main Results:
- Connexin45 and connexin43 were co-expressed and colocalized in BWEM cells.
- Connexin45 is a 48 kD polypeptide and is phosphorylated.
- TPA treatment inhibited intercellular passage, reduced connexin45 expression, and prevented its phosphorylation, while inducing connexin43 phosphorylation.
Conclusions:
- Connexin43 and connexin45 are differentially regulated by TPA treatment in BWEM cells, involving distinct mechanisms of phosphorylation and expression.
- Protein phosphorylation plays a role in regulating connexin43 and connexin45 function.