Genome-wide profiling of Hfq-binding RNAs uncovers extensive post-transcriptional rewiring of major stress response and symbiotic regulons in Sinorhizobium meliloti.
The RNA chaperone Hfq is a global post-transcriptional regulator in bacteria. Here, we used RNAseq to analyze RNA populations from the legume symbiont Sinorhizobium meliloti that were co-immunoprecipitated (CoIP-RNA) with a FLAG-tagged Hfq in five growth/stress conditions. Hfq-bound transcripts (1315) were largely identified in stressed bacteria and derived from small RNAs (sRNAs), both trans-encoded (6.4%) and antisense (asRNAs; 6.3%), and mRNAs (86%). Pull-down with Hfq recovered a small proportion of annotated S. meliloti sRNAs (14% of trans-sRNAs and 2% of asRNAs) suggesting a discrete impact of this protein in sRNA pathways. Nonetheless, Hfq selectively stabilized CoIP-enriched sRNAs, anticipating that these interactions are functionally significant. Transcription of 26 Hfq-bound sRNAs was predicted to occur from promoters recognized by the major stress σ factors σ(E2) or σ(H1/2). Recovery rates of sRNAs in each of the CoIP-RNA libraries suggest a large impact of Hfq-assisted riboregulation in S. meliloti osmoadaptation. Hfq directly targeted 18% of the predicted S. meliloti mRNAs, which encode functionally diverse proteins involved in transport and metabolism, σ(E2)-dependent stress responses, quorum sensing, flagella biosynthesis, ribosome, and membrane assembly or symbiotic nitrogen fixation. Canonical targeting of the 5' regions of two of the ABC transporter mRNAs by the homologous Hfq-binding AbcR1 and AbcR2 sRNAs leading to inhibition of protein synthesis was confirmed in vivo. We therefore provide a comprehensive resource for the systems-level deciphering of hitherto unexplored S. meliloti stress and symbiotic post-transcriptional regulons and the identification of Hfq-dependent sRNA-mRNA regulatory pairs.
Grupo de Ecología Genética de la Rizosfera; Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; CSIC, Granada, Spain.
Omar
O
Torres-Quesada
Center for Biotechnology (CeBiTec); Bielefeld University; Bielefeld, Germany.
Jan
J
Reinkensmeier
LOEWE Center for Synthetic Microbiology (SYNMIKRO) and Department of Biology; Philipps-Universität Marburg; Marburg, Germany.
Jan-Philip
JP
Schlüter
LOEWE Center for Synthetic Microbiology (SYNMIKRO) and Department of Biology; Philipps-Universität Marburg; Marburg, Germany.
Marta
M
Robledo
Grupo de Ecología Genética de la Rizosfera; Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; CSIC, Granada, Spain.
Alexandra
A
Peregrina
Center for Biotechnology (CeBiTec); Bielefeld University; Bielefeld, Germany.
Robert
R
Giegerich
Grupo de Ecología Genética de la Rizosfera; Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; CSIC, Granada, Spain.
Nicolás
N
Toro
LOEWE Center for Synthetic Microbiology (SYNMIKRO) and Department of Biology; Philipps-Universität Marburg; Marburg, Germany.
Anke
A
Becker
Grupo de Ecología Genética de la Rizosfera; Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; CSIC, Granada, Spain.
Jose I
JI
Jiménez-Zurdo
D020029:Base Pairing; D001665:Binding Sites; D015964:Gene Expression Regulation, Bacterial; D035001:Host Factor 1 Protein / Q000378:metabolism*; D011485:Protein Binding; D012323:RNA Processing, Post-Transcriptional; D012329:RNA, Bacterial / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D058727:RNA, Small Untranslated / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D016962:Sinorhizobium meliloti / Q000235:genetics* / Q000378:metabolism*; D013312:Stress, Physiological
D035001:Host Factor 1 Protein; D012329:RNA, Bacterial; D012333:RNA, Messenger; D058727:RNA, Small Untranslated
RNA-binding protein; nitrogen-fixation; riboregulation; sRNA; α-proteobacteria