Actin isoforms organize into distinct networks that are essential for the normal function of eukaryotic cells. Despite a high level of sequence and structure conservation, subtle differences in their design principles determine the interaction with myosin motors and actin-binding proteins. Therefore, identifying how the structure of actin isoforms relates to function is important for our understanding of normal cytoskeletal physiology. Here, we report the high-resolution structures of filamentous skeletal muscle α-actin (3.37 Å), cardiac muscle α-actin (3.07 Å), ß-actin (2.99 Å), and γ-actin (3.38 Å) in the Mg2+·ADP state with their native post-translational modifications. The structures revealed isoform-specific conformations of the N-terminus that shift closer to the filament surface upon myosin binding, thereby establishing isoform-specific interfaces. Collectively, the structures of single-isotype, post-translationally modified bare skeletal muscle α-actin, cardiac muscle α-actin, ß-actin, and γ-actin reveal general principles, similarities, and differences between isoforms. They complement the repertoire of known actin structures and allow for a comprehensive understanding of in vitro and in vivo functions of actin isoforms.
Keywords: actin; biochemistry; chemical biology; cytoskeleton; isoforms; macromolecular structure; molecular biophysics; myosin; none; structural biology.
The protein actin is important for many fundamental processes in biology, from contracting muscle to dividing a cell in two. As actin is involved in such a variety of roles, human cells have slightly different versions of the protein, known as isoforms. For example, alpha-actin is vital for contracting muscle, while beta- and gamma-actin drive cellular processes in non-muscle cells. In order to carry out its various functions, actin interacts with many other proteins inside the cell, such as myosin motors which power muscle contraction. These interactions rely on the precise chain of building blocks, known as amino acids, that make up the actin isoforms; even subtle alterations in this sequence can influence the behavior of the protein. However, it is not clear how differences in the amino acid sequence of the actin isoforms impact actin’s interactions with other proteins. Arora et al. addressed this by studying the structure of four human actin isoforms using a technique called cryo-electron microscopy, where the proteins are flash-frozen and bombarded with electrons. These experiments showed where differences between the amino acid chains of each isoform were located in the protein. Arora et al. then compared their structures with previous work showing the structure of actin bound to myosin. This revealed that the tail-end of the protein (known as the N-terminus) differed in shape between the four isoforms, and this variation may influence how actin binds to others proteins in the cell. These results are an important foundation for further work on actin and how it interacts with other proteins. The structures could help researchers design new tools that can be used to target specific isoforms of actin in different types of laboratory experiments.
© 2023, Arora et al.