L. monocytogenes-induced actin assembly requires the actA gene product, a surface protein

The intracellular pathogenic bacterium L. monocytogenes can spread directly from cell to cell without leaving the cytoplasm. The mechanism of this movement, generated through bacterially induced actin polymerization, is not understood. By analyzing an avirulent Tn917-lac mutant defective for actin polymerization, we have identified a bacterial component involved in this process. The transposon had inserted in actA, the second gene of an operon. Gene disruption of downstream genes and transformation of the mutant strain with actA showed that the actA gene encodes a surface protein necessary for bacterially induced actin assembly. Our results indicate that it is a 610 amino acid protein with an apparent molecular weight of 90 kd.

[1]  P. Sansonetti,et al.  Identification of icsA, a plasmid locus of Shigella flexneri that governs bacterial intra- and intercellular spread through interaction with F-actin. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[2]  D. Portnoy,et al.  Actin filament nucleation by the bacterial pathogen, Listeria monocytogenes , 1990, The Journal of cell biology.

[3]  F. Young,et al.  New shuttle vectors for Bacillus subtilis and Escherichia coli which allow rapid detection of inserted fragments. , 1984, Gene.

[4]  D. Portnoy,et al.  Listeria monocytogenes moves rapidly through the host-cell cytoplasm by inducing directional actin assembly. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[5]  M. Lisanti,et al.  Polarized apical distribution of glycosyl-phosphatidylinositol-anchored proteins in a renal epithelial cell line. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[6]  E. A. Havell Synthesis and secretion of interferon by murine fibroblasts in response to intracellular Listeria monocytogenes , 1986, Infection and immunity.

[7]  E. Wulf,et al.  Fluorescent phallotoxin, a tool for the visualization of cellular actin. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[8]  B. Tall,et al.  Intracellular spread of Shigella flexneri associated with the kcpA locus and a 140-kilodalton protein , 1989, Infection and immunity.

[9]  M. Füzi,et al.  Production of Opacity in Egg-Yolk Medium by Listeria monocytogenes , 1962, Nature.

[10]  D. Werner,et al.  The complete cDNA sequence of mouse elongation factor 1 alpha (EF 1 alpha) mRNA. , 1989, Nucleic acids research.

[11]  S. Makino,et al.  A genetic determinant required for continuous reinfection of adjacent cells on large plasmid in S. flexneri 2a , 1986, Cell.

[12]  P. Sansonetti,et al.  In vitro model of penetration and intracellular growth of Listeria monocytogenes in the human enterocyte-like cell line Caco-2 , 1987, Infection and immunity.

[13]  D. Portnoy,et al.  Role of hemolysin for the intracellular growth of Listeria monocytogenes , 1988, The Journal of experimental medicine.

[14]  D. Lipman,et al.  Rapid and sensitive protein similarity searches. , 1985, Science.

[15]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[16]  S. Ehrlich,et al.  Tn10-derived transposons active in Bacillus subtilis , 1990, Journal of bacteriology.

[17]  S. Bernard,et al.  Immunochemical identification and biological characterization of cytotoxic necrotizing factor from Escherichia coli , 1989, Journal of clinical microbiology.

[18]  G. Blobel,et al.  Intracellular protein topogenesis. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[19]  P. Sansonetti,et al.  Intracellular and cell-to-cell spread of Listeria monocytogenes involves interaction with F-actin in the enterocytelike cell line Caco-2 , 1990, Infection and immunity.

[20]  W. Goebel,et al.  Molecular cloning, sequencing, and identification of a metalloprotease gene from Listeria monocytogenes that is species specific and physically linked to the listeriolysin gene , 1991, Infection and immunity.

[21]  J. Mounier,et al.  A nonvirulent mutant of Listeria monocytogenes does not move intracellularly but still induces polymerization of actin , 1990, Infection and immunity.

[22]  D. Portnoy,et al.  Isolation of Listeria monocytogenes small-plaque mutants defective for intracellular growth and cell-to-cell spread , 1990, Infection and immunity.

[23]  P. Rácz,et al.  Electron microscopic studies in experimental keratoconjunctivitis listeriosa. I. Penetration of Listeria monocytogenes into corneal epithelial cells. , 1970, Acta microbiologica Academiae Scientiarum Hungaricae.

[24]  D. Lipman,et al.  Improved tools for biological sequence comparison. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[25]  S. Craig,et al.  cDNA-derived sequence of chicken embryo vinculin. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[26]  J. Moulder Comparative biology of intracellular parasitism. , 1985, Microbiological reviews.

[27]  R. Levin,et al.  Genetic transformation between strains of Listeria monocytogenes , 1989 .

[28]  P. Cossart,et al.  Transcriptional mapping and nucleotide sequence of the Listeria monocytogenes hlyA region reveal structural features that may be involved in regulation , 1989, Infection and immunity.

[29]  J. Olmsted,et al.  A model for microtubule-associated protein 4 structure. Domains defined by comparisons of human, mouse, and bovine sequences. , 1991, The Journal of biological chemistry.

[30]  J. Vandekerckhove Actin-binding proteins. , 1990, Current opinion in cell biology.

[31]  P. Cossart,et al.  Attenuated mutants of the intracellular bacterium Listeria monocytogenes obtained by single amino acid substitutions in listeriolysin O , 1990, Molecular microbiology.

[32]  J. Condeelis,et al.  Identification of an actin-binding protein from Dictyostelium as elongation factor 1a , 1990, Nature.

[33]  R. Dernick,et al.  Simplified method for silver staining of proteins in polyacrylamide gels and the mechanism of silver staining , 1985 .

[34]  D. Portnoy,et al.  Actin filaments and the growth, movement, and spread of the intracellular bacterial parasite, Listeria monocytogenes , 1989, The Journal of cell biology.

[35]  A. Killinger,et al.  Listeria monocytogenes and listeric infections. , 1966, Bacteriological reviews.

[36]  J. Bryan,et al.  Cloning and expression of a smooth muscle caldesmon. , 1989, The Journal of biological chemistry.

[37]  P. Cossart,et al.  Listeria monocytogenes. A model system for the molecular study of intracellular parasitism. , 1989, Molecular biology & medicine.

[38]  P. Cossart,et al.  Identification of a new operon involved in Listeria monocytogenes virulence: its first gene encodes a protein homologous to bacterial metalloproteases , 1991, Infection and immunity.

[39]  C. M. Goodloe-Holland,et al.  Junctional plasma membrane domains isolated from aggregating Dictyostelium discoideum amebae. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[40]  C. Kocks,et al.  Nucleotide sequence of the lecithinase operon of Listeria monocytogenes and possible role of lecithinase in cell-to-cell spread , 1992, Infection and immunity.

[41]  I. Eperon,et al.  Isolation and characterization of a vinculin cDNA from chick-embryo fibroblasts. , 1987, Biochemical Journal.

[42]  J. Hartwig,et al.  Actin-binding proteins. , 1991, Current opinion in cell biology.

[43]  K. Timmis,et al.  Intercellular spread of Shigella flexneri through a monolayer mediated by membranous protrusions and associated with reorganization of the cytoskeletal protein vinculin , 1991, Infection and immunity.

[44]  D. Clewell,et al.  Complete nucleotide sequence of macrolide-lincosamide-streptogramin B-resistance transposon Tn917 in Streptococcus faecalis , 1985, Journal of bacteriology.

[45]  P. Cossart,et al.  Pleiotropic control of Listeria monocytogenes virulence factors by a gene that is autoregulated , 1991, Molecular microbiology.

[46]  E. Oaks,et al.  Plaque formation by virulent Shigella flexneri , 1985, Infection and immunity.

[47]  S. Makino,et al.  virG, a plasmid-coded virulence gene of Shigella flexneri: identification of the virG protein and determination of the complete coding sequence , 1989, Journal of bacteriology.