Different translation dynamics of β- and γ-actin regulates cell migration

Abstract

β- and γ-cytoplasmic actins are ubiquitously expressed in every cell type and are nearly identical at the amino acid level but play vastly different roles in vivo. Their essential roles in embryogenesis and mesenchymal cell migration critically depend on the nucleotide sequences of their genes, rather than their amino acid sequence, however it is unclear which gene elements underlie this effect. Here we address the specific role of the coding sequence in β- and γ-cytoplasmic actins' intracellular functions, using stable polyclonal populations of immortalized mouse embryonic fibroblasts with exogenously expressed actin isoforms and their 'codon-switched' variants. When targeted to the cell periphery using the β-actin 3′UTR, β-actin and γ-actin have differential effects on cell migration. These effects directly depend on the coding sequence. Single molecule measurements of actin isoform translation, combined with fluorescence recovery after photobleaching, demonstrate a pronounced difference in β- and γ-actins' translation elongation rates in cells, leading to changes in their dynamics at the focal adhesions, impairments in actin bundle formation, and reduced cell anchoring to the substrate during migration. Our results demonstrate that coding sequence-mediated differences in actin translation play a key role in cell migration.

Cofilin and Actin Dynamics: Multiple Modes of Regulation and Their Impacts in Neuronal Development and Degeneration

“…The β-actin mRNA contains a 3 UTR zip code which helps direct its localization to the cell periphery for translation, unlike the γ-actin which is translated nearby the cell nucleus [180]. Other modifications are associated with β-actin translation, such as arginylation [181], suggesting that the location where a protein is made is as important to its function as when it is being made. Coordinated translation of families of proteins might explain their mechanisms of sorting easier than a post-translational code, although one does not exclude the other.…”

“…Thus, it takes a village of actin regulatory proteins working in a cooperative manner to coordinate the diverse and spatially specific functions required for the vast array of actin-dependent cellular processes, especially in metazoans where tissue specific functions are required. These processes are spatially regulated by the localized components of upstream regulatory pathways that can modulate cofilin directly or modulate one or more of the many other proteins that impact cofilin localization, activation, filament severing, monomer binding, etc., resulting in an exquisite and coordinated control of actin-mediated cellular behavior [181]. Understanding the coordinated regulation of this multitude of proteins has become more complex due to the discovery of additional posttranslational modifications of cytoplasmic actin [182], including oxidation/reduction (redox regulation and S-glutathionylation) [121,183], N-terminal arginylation [184], and lysine acetylation [185].…”

Structural Mechanisms of Actin Isoforms

“…Actin isoforms are subject to extensive PTM mechanisms that may drive structural changes, distinct biochemistries, and cellular activities (3,12,41). Prevalent PTMs of actin isoforms include acetylation, phosphorylation, and methylation (Table 2).…”

“…Actin isoforms differ by conservative and nonconservative substitutions (Figure 1-figure supplement 1C) that contribute to their distinct biochemical and in vivo function (1,11,12). Overall, the amino acid sequence divergence is higher between muscle and nonmuscle actins than within muscle or nonmuscle actin sequences (Figure 1 There are no substitutions in SD2, the smallest and most flexible subdomain (34).…”

“…Actin isoforms display differential biochemistries, cellular localization, and interactions with myosin motors and actin-binding proteins (ABPs) (2,3,(6)(7)(8)(9)(10). These characteristics result in the formation of diverse cellular actin networks with distinct compositions, architectures, dynamics, and mechanics that enable fundamental processes such as cell adhesion, migration, and contractility (1,11,12). Actin isoforms share a high sequence identity at the protein level (~93-99%) and the ability to self-assemble into helical, polarized filaments (F-actin) from monomers (G-actin) (2,(13)(14)(15).…”