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Isolation of neonatal cardiomyocytes reduces the expression of the GTP-binding protein, Gh
E Vincan1, C B Neylon, R M Graham
1Cellular Biochemistry Laboratory, Baker Medical Research Institute, Melbourne, Victoria, Australia.
Insights
Alpha 1-adrenoceptors in the heart use a unique GTP-binding protein, Gh, not found in cultured cells. Gh expression is specific to intact cardiac tissue, distinguishing it from Gq and Gi proteins.
Area of Science:
- Cardiovascular Physiology
- Molecular Pharmacology
- G Protein-Coupled Receptors
Background:
- Alpha 1-adrenoceptors typically signal via Gq proteins, activating phospholipase C-beta.
- However, cardiac alpha 1-adrenoceptors have been an exception, coupling to a distinct GTP-binding protein, Gh.
- Gh possesses dual functionality as a transglutaminase and receptor-coupling protein, activating a different phospholipase isoform.
Purpose of the Study:
- To investigate the expression and regulation of the GTP-binding protein Gh in cardiac tissue.
- To determine if Gh expression is maintained in isolated cardiomyocytes or specific to intact heart tissue.
- To compare Gh expression with other G proteins like Gq and Gi in cardiac contexts.
Main Methods:
- Isolation and culture of neonatal cardiomyocytes.
- Western blot analysis to detect and quantify Gh, Gq, and Gi protein levels.
- Analysis of protein expression in both intact neonatal/adult heart tissues and cultured cells.
Main Results:
- Isolation and culture of neonatal cardiomyocytes led to a significant decrease in Gh expression.
- Gh expression levels remained unaffected in cultured cells for Gq and Gi proteins.
- Gh was readily detected in protein extracts from intact neonatal and adult heart tissues.
Conclusions:
- Gh expression is a characteristic feature of intact cardiac tissue, not isolated cardiomyocytes.
- The unique signaling pathway involving Gh in the heart is dependent on the tissue's structural integrity.
- These findings highlight tissue-specific G protein coupling mechanisms for alpha 1-adrenoceptors.
Abstract:
alpha 1-Adrenoceptors in most tissues couple with the heterotrimeric GTP-binding protein Gq, the alpha subunit of which activates the beta-isoforms of phospholipase C. However, in heart (and in liver) alpha 1-adrenoceptors have been reported to couple to a high molecular weight GTP-binding protein. Gh, which functions both as a type II transglutaminase and as a receptor coupling protein. Gh activates a phospholipase isoform distinct from phospholipase C-beta. Here we report that isolation and culture of neonatal cardiomyocytes decreased the expression of Gh without reducing the content of Gq or Gi. Gh was readily detected in extracts from intact neonatal and adult heart tissues. The expression of Gh thus appears to be a feature of intact cardiac tissue.