Effective top-down LC/MS+ method for assessing actin isoforms as a potential cardiac disease marker

Area of Science:

• Biochemistry
• Molecular Biology
• Proteomics

Background:

• Actin is a key cytoskeletal protein essential for cell structure and motility.
• Alpha-cardiac actin (αCAA) and alpha-skeletal actin (αSKA) are major isoforms in striated muscles, with varying expression in disease.
• High sequence similarity (98.9%) between αSKA and αCAA challenges accurate quantification.

Purpose of the Study:

• To develop a robust method for distinguishing and quantifying αSKA and αCAA isoforms.
• To investigate the expression patterns of αSKA and αCAA in healthy and diseased human cardiac and skeletal muscles.
• To assess the potential of αSKA as a biomarker for dilated cardiomyopathy (DCM).

Main Methods:

• Development of a top-down liquid chromatography/mass spectrometry (LC/MS+) based method.
• Purification and comprehensive analysis of α-actin isoforms from muscle tissues.
• Quantification of αSKA and αCAA in healthy and DCM human hearts.

Main Results:

• αSKA is the sole actin isoform in skeletal muscle; αCAA and αSKA are coexpressed in cardiac muscle.
• A novel LC/MS+ method enabled effective purification and analysis of α-actin isoforms.
• αSKA expression was significantly augmented in hearts with DCM (43.1%) compared to healthy controls (23.7%).

Conclusions:

• Top-down LC/MS+ is an effective method for α-actin isoform analysis.
• Quantification of αSKA and αCAA provides insights into muscle physiology and pathology.
• Elevated αSKA in DCM suggests its potential as a clinical marker for cardiac disease.