Visualization of Compartmentalized Kinase Activity Dynamics Using Adaptable BimKARs

Area of Science:

• Cellular signaling and kinase activity monitoring
• Molecular imaging and biosensor development

Background:

• Real-time monitoring of kinase activity is crucial for understanding spatiotemporal regulation of cellular signaling.
• Existing methods may lack the dynamic range to capture subtle changes in kinase activity.

Purpose of the Study:

• To report on the adaptability and utility of bimolecular kinase activity reporters (bimKARs) for live-cell imaging.
• To demonstrate the enhanced dynamic range of bimKARs compared to unimolecular reporters.
• To investigate the regulation of AMP-activated protein kinase (AMPK) by protein kinase A (PKA) at the plasma membrane.

Main Methods:

• Development and application of bimolecular kinase activity reporters (bimKARs).
• Monitoring of kinase activity for JNK, ERK, and AMPK in live cells.
• Investigation of AMPK regulation by PKA using bimKARs.

Main Results:

• bimKARs demonstrated versatility in monitoring various kinases, including JNK, ERK, and AMPK.
• bimKARs exhibited significantly enhanced dynamic ranges compared to unimolecular reporters.
• The study revealed dual regulatory roles (negative and positive) of PKA on AMPK activity within the same cell at the plasma membrane.

Conclusions:

• bimKARs are adaptable and powerful molecular tools for real-time visualization of kinase activity dynamics.
• The enhanced dynamic range of bimKARs allows for the detection of previously unresolvable signaling events.
• PKA exhibits complex, context-dependent regulation of AMPK activity at the plasma membrane.