Macrophage-induced genes of Legionella pneumophila: protection from reactive intermediates and solute imbalance during intracellular growth.
A promoter-probe strategy was devised to identify genes specifically expressed by Legionella pneumophila during growth within the macrophage. Random fragments from the L. pneumophila chromosome were inserted upstream of a promoterless phage T4 td gene, and fragments that led to complementation of thymine auxotrophy during intracellular growth of the bacterium were identified. Two different selection strategies were employed to eliminate promoters that were also active during extracellular growth of the bacterium. Some of these genes were identified independently by using both of the selection strategies. The factors identified include orthologs of efflux-mediated resistance determinants and transporters, a transporter involved in protection from osmotic stress, a stress response GTP-binding protein, a response regulator, a sensor kinase, and two systems that increase the reducing potential of the bacterium, one of which encodes the L. pneumophila ortholog of ahpC. Five of the clones analyzed here were fusions to promoters that were closely linked to genes encoding three-component chemiosmotic efflux pumps that export heavy metals or toxic organic compounds. Analysis of ahpC gene expression indicates that levels increased at least sevenfold during intracellular growth of the bacterium. Inactivation of several of the genes at their chromosomal loci had no effect on the intracellular growth rate of L. pneumophila in cultured macrophages. This suggests that a number of genes with increased expression during intracellular growth may be part of redundant systems that allow survival and growth under the conditions encountered within host cells.
Rankin
Susannah
S
Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
Li
Zhiru
Z
Isberg
Ralph R
RR
D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005798:Genes, Bacterial; D008040:Genetic Linkage; D016680:Genome, Bacterial; D006801:Humans; D007424:Intracellular Fluid / Q000276:immunology / Q000382:microbiology; D016952:Legionella pneumophila / Q000235:genetics* / Q000254:growth & development / Q000276:immunology; D008264:Macrophages / Q000276:immunology / Q000382:microbiology*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010544:Peroxidases / Q000235:genetics; D054464:Peroxiredoxins; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014158:Transcription, Genetic; D020298:U937 Cells
D004269:DNA, Bacterial; D011993:Recombinant Fusion Proteins; D010544:Peroxidases; D054464:Peroxiredoxins