Identification of small RNAs in Francisella tularensis.
BACKGROUND
Regulation of bacterial gene expression by small RNAs (sRNAs) have proved to be important for many biological processes. Francisella tularensis is a highly pathogenic Gram-negative bacterium that causes the disease tularaemia in humans and animals. Relatively little is known about the regulatory networks existing in this organism that allows it to survive in a wide array of environments and no sRNA regulators have been identified so far.


RESULTS
We have used a combination of experimental assays and in silico prediction to identify sRNAs in F. tularensis strain LVS. Using a cDNA cloning and sequencing approach we have shown that F. tularensis expresses homologues of several sRNAs that are well-conserved among diverse bacteria. We have also discovered two abundant putative sRNAs that share no sequence similarity or conserved genomic context with any previously annotated regulatory transcripts. Deletion of either of these two loci led to significant changes in the expression of several mRNAs that likely include the cognate target(s) of these sRNAs. Deletion of these sRNAs did not, however, significantly alter F. tularensis growth under various stress conditions in vitro, its replication in murine cells, or its ability to induce disease in a mouse model of F. tularensis infection. We also conducted a genome-wide in silico search for intergenic loci that suggests F. tularensis encodes several other sRNAs in addition to the sRNAs found in our experimental screen.


CONCLUSIONS
Our findings suggest that F. tularensis encodes a significant number of non-coding regulatory RNAs, including members of well conserved families of structural and housekeeping RNAs and other poorly conserved transcripts that may have evolved more recently to help F. tularensis deal with the unique and diverse set of environments with which it must contend.
INSERM U1002, Paris, France.
Guillaume
G
Postic
Eric
E
Frapy
Marion
M
Dupuis
Iharilalao
I
Dubail
Jonathan
J
Livny
Alain
A
Charbit
Karin L
KL
Meibom
D000818:Animals; D001428:Bacterial Vaccines / Q000276:immunology; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D019295:Computational Biology; D018076:DNA, Complementary / Q000235:genetics; D005604:Francisella tularensis / Q000235:genetics* / Q000276:immunology; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D042541:Intracellular Space / Q000382:microbiology; D008264:Macrophages / Q000382:microbiology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D020411:Oligonucleotide Array Sequence Analysis; D034443:RNA Transport / Q000235:genetics; D012329:RNA, Bacterial / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014613:Vaccines, Attenuated / Q000276:immunology
D001428:Bacterial Vaccines; D018076:DNA, Complementary; D012329:RNA, Bacterial; D014613:Vaccines, Attenuated