Abrams, M. E. | bioRxiv Science

bioRxiv2019

The mechanisms of dynamin-actin interaction

Cell-cell fusion is an indispensable process in the conception, development and physiology of multicellular organisms. Here we demonstrate a direct and noncanonical role for dynamin, best known as a fission GTPase in endocytosis, in cell-cell fusion. Our genetic and cell biological analyses show that dynamin colocalizes within the F-actin-enriched podosome-like structures at the fusogenic synapse, which is required for generating invasive membrane protrusions and myoblast fusion in vivo, in an endocytosis-independent manner. Biochemical, negative stain EM and cryo-electron tomography (cryo-ET) analyses revealed that dynamin forms helices that directly bundles actin filaments by capturing multiple actin filaments at their outer rim via interactions with dynamins proline-rich domain. GTP hydrolysis by dynamin triggers disassembly of the dynamin helix, exposes the sides of the actin filaments, promotes dynamic Arp2/3-mediated branched actin polymerization, and generates a mechanically stiff actin network. Thus, dynamin functions as a unique actin-bundling protein that enhances mechanical force generation by the F-actin network in a GTPase-dependent manner. Our findings have universal implications for understanding dynamin-actin interactions in various cellular processes beyond cell-cell fusion.

cell biology↗

bioRxiv2017

Actin assembly-inducing protein ActA promotes FcγRIa-mediated Listeria internalization

Listeria monocytogenes is a Gram-positive intracellular pathogen and the causative agent of human listeriosis. While the ability of L. monocytogenes to enter and survive in professional phagocytes is critical to establish a successful infection, mechanisms of invasion are poorly understood. Our previous investigation into the role of type I interferon-stimulated genes in bacterial infection revealed that the human immunoglobulin receptor Fc{gamma}RIa served as a L. monocytogenes invasion factor. Fc{gamma}RIa-mediated L. monocytogenes entry occurred independently of immunoglobulin interaction or bacterial internalins. However, the bacterial determinants that mediate Fc{gamma}RIa interaction remain unclear. Using a comparative genomics approach, we identify actin assembly-inducing protein ActA as a pathogen specific ligand of Fc{gamma}RIa. Fc{gamma}RIa enhanced entry of pathogenic L. monocytogenes and L. ivanovii strain but not non-pathogenic L. innocua. We found that the major virulence regulator PrfA is required for pathogen entry into Fc{gamma}RIa-expressing cells and identify its gene target actA as the critical Listeria ligand. ActA alone was sufficient to promote entry into Fc{gamma}RIa-expressing cells, and this function is independent of its actin nucleating activity. Together, these studies present an unexpected role of ActA beyond its canonical function in actin-based motility and expand our understanding of Listeria strategies for host cell invasion.\n\nImportanceListeria monocytogenes is a food-borne bacterial pathogen and a causative agent of listeriosis with up to 50% mortality rate in immunocompromised individuals. While the mechanisms of Listeria entry into non-phagocytic cells have been extensively characterized, the details of phagocytic cell invasion are still poorly understood. We have recently discovered that human immunoglobulin receptor Fc{gamma}RIa mediates Listeria uptake by monocytic cells. This process occurred independently of canonical immunoglobulin interactions as well as classic Listeria internalization factors. Importantly, molecular determinants of Listeria-Fc{gamma}RIa interaction leading to bacterial entry, remained unknown. In this study, we demonstrate that Listeria virulence factor actin-assembly inducing protein ActA is required for Fc{gamma}RIa-mediated entry. Further, ActA was found to be sufficient for the internalization, suggesting its role as a bacterial ligand of Fc{gamma}RIa. Together, these findings expand our knowledge of mechanisms that Listeria has evolved to exploit cellular signaling pathways and immune defense of the host.

microbiology↗