Photorhabdus and Xenorhabdus species have mutualistic associations with nematodes and an entomopathogenic stage1,2 in their life cycles. In both stages, numerous specialized metabolites are produced that have roles in symbiosis and virulence3,4. Although regulators have been implicated in the regulation of these specialized metabolites3,4, how small regulatory RNAs (sRNAs) are involved in this process is not clear. Here, we show that the Hfq-dependent sRNA, ArcZ, is required for specialized metabolite production in Photorhabdus and Xenorhabdus. We discovered that ArcZ directly base-pairs with the mRNA encoding HexA, which represses the expression of specialized metabolite gene clusters. In addition to specialized metabolite genes, we show that the ArcZ regulon affects approximately 15% of all transcripts in Photorhabdus and Xenorhabdus. Thus, the ArcZ sRNA is crucial for specialized metabolite production in Photorhabdus and Xenorhabdus species and could become a useful tool for metabolic engineering and identification of commercially relevant natural products.
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All .mzXML files from the HPLC–MS/MS runs are available at MassIVE (https://massive.ucsd.edu) under the ID MSV000084163. Raw sequence data are available at the European nucleotide archive (https://www.ebi.ac.uk/ena/) under project accession numbers PRJEB33827 and PRJEB24159. The proteomic data can be accessed at PRIDE (https://www.ebi.ac.uk/pride/) with the project accession number PXD019095. Source data are provided with this paper.
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This work was funded in part by the DFG (SFB 902, project no. B17) and the LOEWE Centre for Translational Biodiversity Genomics (LOEWE TBG), supported by the State of Hesse. K.P. acknowledges funding by the DFG (EXC 2051, grant no. 390713860), Vallee Foundation and European Research Council (grant no. StG-758212). We thank A. Goodman for providing pSAM-BT and for helpful discussions. We thank L. Pöschel and A. K. Heinrich for the plasmid construction.
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Expression of various sRNAs in P. laumondii at different time points. RNA samples of P. laumondii WT and ∆hfq strains were taken at three different OD600 values (0.5, 2 and 4) and after 24 h of growth. The RNA was loaded on northern blots and probed for the indicated sRNAs. Probing for 5 S rRNA served as loading control. Source data
Expression of various sRNAs in X. szentirmaii at different time points. RNA samples of X. szentirmaii WT and ∆hfq strains were taken at three different OD600 values (0.5, 2 and 4) and after 24 h of growth. The RNA was loaded on northern blots and probed for the indicated sRNAs. Probing for 5 S rRNA served as loading control. Source data
Extended Data Fig. 3 Phenotype and specialized metabolite profile of transposon-insertion mutants of P. laumondii.
Phenotype of transposon-insertion mutants of P. laumondii. a. Differences in pigmentation of transposon-insertion mutant liquid cultures compared to WT. Depicted are eleven transposon-insertion mutants and a WT culture after 3 d of cultivation at 30 °C with shaking. b. SM profiles of the transposon-insertion mutants. Relative SM production was quantified from duplicates using TargetAnalysis (Bruker) and compared to the WT of P. laumondii after 72 h cultivation at 30 °C with shaking. Mutant 3 was analysed further and the transposon insertion was identified in the arcZ gene.
Infective juvenile development to hermaphrodites with strains of P. laumondii and X. szentirmaii. Data are presented as the mean ± s.e.m. Dots represent biologically independent replicates (n = 10). Asterisks indicate statistical significance (*P < 0.05, **P < 0.005, ***P < 0.0005, ****P < 0.00005) of relative recovery compared to WT recovery levels. Statistical significances were calculated using a two-sided unpaired t-test. Exact p values (left to right, respectively) for P. laumondii TTO1 correspond to p = <0.0001, 0.0006 and for X. szentirmaii to P = 0.56, 0.0094. Source data
Unprocessed northern blots.
Raw data of specialized metabolite quantification using TargetAnalysis.
Raw data of the GFP measurements (b) and specialized metabolite quantification with TargetAnalysis (e).
Unprocessed northern blots.
Unprocessed northern blots.
Unprocessed northern blots.
Raw data of nematode bioassays.
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Neubacher, N., Tobias, N.J., Huber, M. et al. Symbiosis, virulence and natural-product biosynthesis in entomopathogenic bacteria are regulated by a small RNA. Nat Microbiol 5, 1481–1489 (2020). https://doi.org/10.1038/s41564-020-00797-5