Missense Mutations in Desmoplakin Plakin Repeat Domains Have Dramatic Effects on Domain Structure and Function.

Area of Science:

• Molecular and Cellular Biology
• Structural Biology
• Genetics and Disease

Background:

• Plakin repeat domains (PRDs) are crucial for plakin protein interaction with intermediate filament (IF) cytoskeletons.
• These interactions are essential for tissue integrity in cardiac and epithelial cells.
• Mutations in PRDs are linked to cardiomyopathies and skin blistering diseases.

Purpose of the Study:

• To investigate the functional and structural impact of 12 disease-associated missense mutations on desmoplakin's PRDs.
• To elucidate the molecular mechanisms by which these mutations cause disease.

Main Methods:

• Expression of mutant PRD proteins in bacterial cells to assess solubility.
• Transfection of HeLa cells to evaluate the effect of mutations on protein localization to vimentin IFs.
• Bioinformatic and structural modeling to predict the impact of mutations on PRD stability and function.

Main Results:

• Five mutations (G2056R, E2193K, G2338R, G2375R, G2647D) resulted in insoluble PRD proteins.
• The G2375R mutation impaired the targeting of desmoplakin to vimentin IFs in HeLa cells.
• Deletion of PRD-B and PRD-C also compromised vimentin targeting, indicating their importance.
• Modeling suggested mechanisms by which mutations destabilize PRDs and disrupt cytoskeletal linkages.

Conclusions:

• Disease-causing mutations in desmoplakin PRDs can lead to protein insolubility and mislocalization.
• These disruptions compromise essential interactions with the intermediate filament cytoskeleton.
• Understanding these molecular mechanisms may inform therapeutic strategies for related cardiomyopathies and skin disorders.