title: Actin restructuring during Salmonella typhimurium infection investigated by confocal and super-resolution microscopy.
abstract: We have used super-resolution optical microscopy and confocal microscopy to visualize the cytoskeletal restructuring of HeLa cells that accompanies and enables Salmonella typhimurium internalization. Herein, we report the use of confocal microscopy to verify and explore infection conditions that would be compatible with super-resolution optical microscopy, using Alexa-488 labeled phalloidin to stain the actin cytoskeletal network. While it is well known that actin restructuring and cytoskeletal rearrangements often accompany and assist in bacterial infection, most studies have employed conventional diffraction-limited fluorescence microscopy to explore these changes. Here we show that the superior spatial resolution provided by single-molecule localization methods (such as direct stochastic optical reconstruction microscopy) enables more precise visualization of the nanoscale changes in the actin cytoskeleton that accompany bacterial infection. In particular, we found that a thin (100-nm) ring of actin often surrounds an invading bacteria 10 to 20 min postinfection, with this ring being transitory in nature. We estimate that a few hundred monofilaments of actin surround the S. typhimurium in this heretofore unreported bacterial internalization intermediate.
authors.affiliation: Los Alamos National Laboratory, Center for Integrated Nanotechnologies, Los Alamos, New Mexico 87545.
authors.forename: Jason J
authors.initials: JJ
authors.lastname: Han
authors.affiliation: Los Alamos National Laboratory, Biosciences Division, Los Alamos, New Mexico 87545.
authors.forename: Yuliya A
authors.initials: YA
authors.lastname: Kunde
authors.affiliation: Los Alamos National Laboratory, Biosciences Division, Los Alamos, New Mexico 87545.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Hong-Geller
authors.affiliation: Los Alamos National Laboratory, Center for Integrated Nanotechnologies, Los Alamos, New Mexico 87545.
authors.forename: James H
authors.initials: JH
authors.lastname: Werner
mesh_terms: D000199:Actins / Q000378:metabolism*; D004396:Coloring Agents / Q000737:chemistry; D003599:Cytoskeleton / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006834:Hydrazines / Q000737:chemistry; D008853:Microscopy / Q000379:methods*; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence; D010590:Phalloidine / Q000737:chemistry; D012480:Salmonella Infections / Q000378:metabolism*; D012486:Salmonella typhimurium; D013269:Stochastic Processes
chemical_list: D000199:Actins; C417561:Alexa 488 hydrazide; D004396:Coloring Agents; D006834:Hydrazines; D010590:Phalloidine