Targeted proteolysis sustains calcineurin activation.

Area of Science:

• Cardiovascular Biology
• Molecular Mechanisms of Disease
• Enzymology

Background:

• Calcineurin (CnA) plays a crucial role in regulating myocardial hypertrophy.
• Pathological workload induces targeted proteolysis of the CnA autoinhibitory domain, enhancing its activity in human myocardium.
• The precise proteolytic mechanism underlying CnA activation requires investigation.

Purpose of the Study:

• To elucidate the proteolytic mechanism responsible for calcineurin (CnA) activation in cardiomyocytes.
• To investigate the role of calpain in the proteolysis and activation of CnA under hypertrophic stimuli.
• To determine the consequences of CnA proteolysis on its activity and subcellular localization.

Main Methods:

• Stimulation of rat cardiomyocytes with Angiotensin II (Ang II).
• Assay of calpain activity and measurement of CnA proteolysis.
• Inhibition of calpain using a membrane-permeable inhibitor.
• Quantification of CnA activity and assessment of nuclear translocation via immunohistochemistry and GFP-tagging.

Main Results:

• Ang II significantly increased calpain activity and induced proteolysis of CnA's autoinhibitory domain.
• Calpain inhibition prevented Ang II-induced proteolysis and subsequent CnA activation.
• Ang II stimulation led to CnA nuclear translocation, which was reversed by calpain inhibition.
• Proteolysis of the autoinhibitory domain resulted in constitutively active and nuclear CnA.

Conclusions:

• Ang II-induced calpain activation mediates the proteolysis of CnA's autoinhibitory domain.
• This proteolysis increases CnA activity and promotes its nuclear translocation, contributing to myocardial hypertrophy.
• The loss of the autoinhibitory domain leads to persistent nuclear localization and activity of CnA, even after stimulus removal.