Salmonella enterica Serovar Typhi Uses Type IVB Pili To Enter Human Intestinal Epithelial Cells
| Autor: | Xiao-Lian Zhang, Christina Morris, Jim Hackett, Xiaoyun Dai, Ada W. Y. Fung, Yanhua Yang, Cecilia M. C. Yip, Danny Ka-Ho Wong, Inez S. M. Tsui |
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| Rok vydání: | 2000 |
| Předmět: |
Operon
Molecular Sequence Data Immunology Fimbria Biology Salmonella typhi medicine.disease_cause Microbiology Bacterial Adhesion Pilus Plasmid Bacterial Proteins medicine Humans Amino Acid Sequence Intestinal Mucosa Protein Precursors Serotyping Cells Cultured Sequence Homology Amino Acid Virulence Membrane Proteins Epithelial Cells Molecular biology Pathogenicity island Infectious Diseases Vibrio cholerae Fimbriae Bacterial Pilin Molecular and Cellular Pathogenesis biology.protein Parasitology Fimbriae Proteins Transcription Factors |
| Zdroj: | Infection and Immunity. 68:3067-3073 |
| ISSN: | 1098-5522 0019-9567 |
| DOI: | 10.1128/iai.68.6.3067-3073.2000 |
| Popis: | DNA sequencing upstream of the Salmonella enterica serovar Typhi pilV and rci genes previously identified in the ca. 118-kb major pathogenicity island (X.-L. Zhang, C. Morris, and J. Hackett, Gene 202:139‐146, 1997) identified a further 10 pil genes apparently forming a pil operon. The product of the pilS gene, prePilS protein (a putative type IVB structural prepilin) was purified, and an anti-prePilS antiserum was raised in mice. Mutants of serovar Typhi either lacking the whole pil operon or with an insertion mutation in the pilS gene were constructed, as was a strain in which the pilN to pilV genes were driven by the tac promoter. The pil 1 strains synthesized type IVB pili, as judged by (i) visualization in the electron microscope of thin pili in culture supernatants of one such strain and (ii) the presence of PilS protein (smaller than the prePilS protein by removal of the leader peptide) on immunoblotting of material pelleted by high-speed centrifugation of either the culture supernatant or sonicates of pil 1 strains. Control pil mutants did not express the PilS protein. A pilS mutant of serovar Typhi entered human intestinal INT407 cells in culture to levels only 5 to 25% of those of the wild-type strain, and serovar Typhi entry was strongly inhibited by soluble prePilS protein (50% inhibition of entry at 1.4 mM prePilS). Earlier, it was reported that the major pathogenicity island of Salmonella enterica serovar Typhi, which is ca. 118 kb in size (11), contained pilV and rci genes, which were cloned and sequenced (22). The Rci gene product was shown to be a site-specific recombinase, active to invert DNA in the C-terminal region of the pilV gene, so that two PilV proteins could be synthesized. Comparisons with database sequences indicated that the two possible pilV genes might code for pilus-tip adhesins, as the serovar Typhi PilV sequence was similar to that of PilV proteins encoded by the Escherichia coli R64 plasmid. In R64-bearing strains, different PilV proteins, borne on type IV pili, select various recipients in liquid mating (the R64-bearing cell is the donor) (10). Both serovar Typhi PilV proteins were seen when the two pilV genes were transcribed from the T7 promoter. The discovery of the serovar Typhi pilV and rci genes in the ca. 118-kb pathogenicity island (henceforth in this work termed the large pathogenicity island) suggested that serovar Typhi might synthesize thin pili belonging to the type IV pilin family (9). As type IV pili, encoded in a Vibrio cholerae pathogenicity island (7, 8) are used by V. cholerae as mediators of adhesion to human cells (13, 18), it was of interest to ask (i) if serovar Typhi also synthesizes type IV pili and (ii) if such pili are important in adherence to or invasion of human intestinal cells. These topics are the subject of this paper. MATERIALS AND METHODS Materials. All reagents were of molecular biology grade. Enzymes active on DNA were obtained from either GibcoBRL or Boehringer Mannheim and were |
| Databáze: | OpenAIRE |
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