RESUMO
Septins are cytoskeletal proteins that participate in cell adhesion, migration, and polarity establishment. The septin subunit SEPT9 directly interacts with the single LIM domain of epithelial protein lost in neoplasm (EPLIN), an actin-bundling protein. Using a human SEPT9 KO fibroblast cell line, we show that cell adhesion and migration are regulated by the interplay between both proteins. The low motility of SEPT9-depleted cells could be partly rescued by increased levels of EPLIN. The normal organization of actin-related filopodia and stress fibers was directly dependent on the expression level of SEPT9 and EPLIN. Increased levels of SEPT9 and EPLIN enhanced the size of focal adhesions in cell protrusions, correlating with stabilization of actin bundles. Conversely, decreased levels had the opposite effect. Our work thus establishes the interaction between SEPT9 and EPLIN as an important link between the septin and the actin cytoskeleton, influencing cell adhesion, motility, and migration.
Assuntos
Adesão Celular , Movimento Celular , Fibroblastos , Adesões Focais , Proteínas com Domínio LIM , Septinas , Humanos , Septinas/metabolismo , Septinas/genética , Movimento Celular/genética , Fibroblastos/metabolismo , Proteínas com Domínio LIM/metabolismo , Proteínas com Domínio LIM/genética , Adesões Focais/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas do Citoesqueleto/genética , Pseudópodes/metabolismo , Citoesqueleto de Actina/metabolismo , Linhagem Celular , Actinas/metabolismo , Fibras de Estresse/metabolismoRESUMO
The septins of the yeast Saccharomyces cerevisiae assemble into hetero-octameric rods by alternating interactions between neighboring G-domains or N- and C-termini, respectively. These rods polymerize end to end into apolar filaments, forming a ring beneath the prospective new bud that expands during the cell cycle into an hourglass structure. The hourglass finally splits during cytokinesis into a double ring. Understanding these transitions as well as the plasticity of the higher order assemblies requires a detailed knowledge of the underlying structures. Here we present the first X-ray crystal structure of a tetrameric Shs1-Cdc12-Cdc3-Cdc10 complex at a resolution of 3.2 Å. Close inspection of the NC-interfaces of this and other septin structures reveals a conserved contact motif that is essential for NC-interface integrity of yeast and human septins in vivo and in vitro. Using the tetrameric structure in combination with AlphaFold-Multimer allowed us to propose a model of the octameric septin rod.