title: The intracellular sRNA transcriptome of Listeria monocytogenes during growth in macrophages.
abstract: Small non-coding RNAs (sRNAs) are widespread effectors of post-transcriptional gene regulation in bacteria. Currently extensive information exists on the sRNAs of Listeria monocytogenes expressed during growth in extracellular environments. We used deep sequencing of cDNAs obtained from fractioned RNA (<500 nt) isolated from extracellularly growing bacteria and from L. monocytogenes infected macrophages to catalog the sRNA repertoire during intracellular bacterial growth. Here, we report on the discovery of 150 putative regulatory RNAs of which 71 have not been previously described. A total of 29 regulatory RNAs, including small non-coding antisense RNAs, are specifically expressed intracellularly. We validated highly expressed sRNAs by northern blotting and demonstrated by the construction and characterization of isogenic mutants of rli31, rli33-1 and rli50* for intracellular expressed sRNA candidates, that their expression is required for efficient growth of bacteria in macrophages. All three mutants were attenuated when assessed for growth in mouse and insect models of infection. Comparative genomic analysis revealed the presence of lineage specific sRNA candidates and the absence of sRNA loci in genomes of naturally occurring infection-attenuated bacteria, with additional loss in non-pathogenic listerial genomes. Our analyses reveal extensive sRNA expression as an important feature of bacterial regulation during intracellular growth.
authors.affiliation: Institute of Medical Microbiology, Justus-Liebig-University, Frankfurter Strasse 107, 35392 Giessen, Germany.
authors.forename: Mobarak A
authors.initials: MA
authors.lastname: Mraheil
authors.forename: André
authors.initials: A
authors.lastname: Billion
authors.forename: Walid
authors.initials: W
authors.lastname: Mohamed
authors.forename: Krishnendu
authors.initials: K
authors.lastname: Mukherjee
authors.forename: Carsten
authors.initials: C
authors.lastname: Kuenne
authors.forename: Jordan
authors.initials: J
authors.lastname: Pischimarov
authors.forename: Christian
authors.initials: C
authors.lastname: Krawitz
authors.forename: Julia
authors.initials: J
authors.lastname: Retey
authors.forename: Thomas
authors.initials: T
authors.lastname: Hartsch
authors.forename: Trinad
authors.initials: T
authors.lastname: Chakraborty
authors.forename: Torsten
authors.initials: T
authors.lastname: Hain
mesh_terms: D000818:Animals; D002460:Cell Line; D005260:Female; D020869:Gene Expression Profiling; D008087:Listeria / Q000235:genetics; D008089:Listeria monocytogenes / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity; D008088:Listeriosis / Q000382:microbiology; D008264:Macrophages / Q000382:microbiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D016372:RNA, Antisense / Q000378:metabolism; D012329:RNA, Bacterial / Q000378:metabolism*; D058727:RNA, Small Untranslated / Q000378:metabolism*; D038621:Regulatory Sequences, Ribonucleic Acid; D058928:Riboswitch; D017423:Sequence Analysis, RNA
chemical_list: D016372:RNA, Antisense; D012329:RNA, Bacterial; D058727:RNA, Small Untranslated; D038621:Regulatory Sequences, Ribonucleic Acid; D058928:Riboswitch