Asymmetric distribution of the Listeria monocytogenes ActA protein is required and sufficient to direct actin-based motility.
Listeria monocytogenes is a Gram-positive facultative intracytoplasmic bacterial pathogen that exhibits rapid actin-based motility in eukaryotic cells and in cell-free cytoplasmic extracts. The protein product of the actA gene is required for bacterial movement and is normally expressed in a polarized fashion on the bacterial surface. Here we demonstrate that the ActA protein is sufficient to direct motility in the absence of other L. monocytogenes gene products, and that polarized localization of the protein is required for efficient unidirectional movement. We have engineered a fusion protein combining ActA with the C-terminal domain of the LytA protein of Streptococcus pneumoniae, which mediates high-affinity binding to DEAE-cellulose and to choline moieties present in the S. pneumoniae cell wall. DEAE-cellulose fragments or S. pneumoniae coated uniformly with the ActA/LytA fusion protein nucleate actin filament growth in cytoplasmic extracts, but do not move efficiently. However, when ActA/LytA-coated S. pneumoniae is grown to polarize the distribution of the fusion protein, the bacteria exhibit unidirectional actin-based movement similar to the normal movement of L. monocytogenes.
Smith
G A
GA
Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Portnoy
D A
DA
Theriot
J A
JA
D000199:Actins / Q000502:physiology*; D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002465:Cell Movement; D003001:Cloning, Molecular; D017931:DNA Primers; D005260:Female; D005798:Genes, Bacterial; D008089:Listeria monocytogenes / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000502:physiology; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D014020:Tissue Extracts; D014982:Xenopus laevis
D000199:Actins; D001426:Bacterial Proteins; D017931:DNA Primers; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D014020:Tissue Extracts; C072980:actA protein, Listeria monocytogenes