Differential N-terminal processing of beta and gamma actin

Area of Science:

• Cell biology
• Molecular biology
• Biochemistry

Background:

• Cytoplasmic beta- and gamma-actin are essential cytoskeletal proteins with highly similar N-terminal amino acid sequences.
• Dissecting the individual regulation of these actin isoforms is challenging due to their sequence homology.

Purpose of the Study:

• To investigate potential differences in the post-translational regulation of beta- and gamma-actin.
• To identify the mechanisms and functional consequences of isoform-specific actin processing.

Main Methods:

• Mass spectrometry was employed to analyze actin processing in cultured cells and tissues.
• CRISPR/Cas-9 gene editing was used to delete candidate enzymes involved in N-terminal processing.
• Changes in F-actin levels and cell morphology/migration were assessed.

Main Results:

• Beta-actin, but not gamma-actin, was found to undergo sequential removal of N-terminal aspartic acid residues, generating truncated forms.
• This processing affects up to approximately 3% of beta-actin in various cell types.
• Deletion of candidate processing enzymes significantly reduced beta-actin N-terminal processing and altered F-actin levels, cell spreading, filopodia formation, and migration.

Conclusions:

• A novel mechanism of isoform-specific actin regulation involving N-terminal processing of beta-actin has been identified.
• This processing impacts key cellular functions related to cell shape and motility.
• The findings reveal previously uncharacterized regulatory pathways for actin isoforms.