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Letter to the EditorInfectious disease Free access | 10.1172/JCI153234

Collateral effects of deletion of nlpD on rpoS and rpoS-dependent genes. Reply.

Inès Ambite,1 Ulrich Dobrindt,2 and Catharina Svanborg1

1Department of Microbiology, Immunology and Glycobiology, Institute of Laboratory Medicine, Lund University, Lund, Sweden.

2Institute of Hygiene, University of Münster, Münster, Germany.

Address correspondence to: Catharina Svanborg, Lund University, Klinikgatan 28, 222 42 Lund, Sweden. Phone: 46.46.222.70.44; Email: catharina.svanborg@med.lu.se.

Find articles by Ambite, I. in: JCI | PubMed | Google Scholar |

1Department of Microbiology, Immunology and Glycobiology, Institute of Laboratory Medicine, Lund University, Lund, Sweden.

2Institute of Hygiene, University of Münster, Münster, Germany.

Address correspondence to: Catharina Svanborg, Lund University, Klinikgatan 28, 222 42 Lund, Sweden. Phone: 46.46.222.70.44; Email: catharina.svanborg@med.lu.se.

Find articles by Dobrindt, U. in: JCI | PubMed | Google Scholar |

1Department of Microbiology, Immunology and Glycobiology, Institute of Laboratory Medicine, Lund University, Lund, Sweden.

2Institute of Hygiene, University of Münster, Münster, Germany.

Address correspondence to: Catharina Svanborg, Lund University, Klinikgatan 28, 222 42 Lund, Sweden. Phone: 46.46.222.70.44; Email: catharina.svanborg@med.lu.se.

Find articles by Svanborg, C. in: JCI | PubMed | Google Scholar

Published September 15, 2021 - More info

Published in Volume 131, Issue 18 on September 15, 2021
J Clin Invest. 2021;131(18):e153234. https://doi.org/10.1172/JCI153234.
© 2021 American Society for Clinical Investigation
Published September 15, 2021 - Version history
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Related articles:

Active bacterial modification of the host environment through RNA polymerase II inhibition
Inès Ambite, … , Ulrich Dobrindt, Catharina Svanborg
Inès Ambite, … , Ulrich Dobrindt, Catharina Svanborg
Research Article Inflammation Microbiology

Active bacterial modification of the host environment through RNA polymerase II inhibition

  • Text
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Abstract

Unlike pathogens, which attack the host, commensal bacteria create a state of friendly coexistence. Here, we identified a mechanism of bacterial adaptation to the host niche, where they reside. Asymptomatic carrier strains were shown to inhibit RNA polymerase II (Pol II) in host cells by targeting Ser2 phosphorylation, a step required for productive mRNA elongation. Assisted by a rare, spontaneous loss-of-function mutant from a human carrier, the bacterial NlpD protein was identified as a Pol II inhibitor. After internalization by host cells, NlpD was shown to target constituents of the Pol II phosphorylation complex (RPB1 and PAF1C), attenuating host gene expression. Therapeutic efficacy of a recombinant NlpD protein was demonstrated in a urinary tract infection model, by reduced tissue pathology, accelerated bacterial clearance, and attenuated Pol II–dependent gene expression. The findings suggest an intriguing, evolutionarily conserved mechanism for bacterial modulation of host gene expression, with a remarkable therapeutic potential.

Authors

Inès Ambite, Nina A. Filenko, Elisabed Zaldastanishvili, Daniel S.C. Butler, Thi Hien Tran, Arunima Chaudhuri, Parisa Esmaeili, Shahram Ahmadi, Sanchari Paul, Björn Wullt, Johannes Putze, Swaine L. Chen, Ulrich Dobrindt, Catharina Svanborg

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Collateral effects of deletion of nlpD on rpoS and rpoS-dependent genes
Manami Tsunoi, … , Sunao Iyoda, Tadayuki Iwase
Manami Tsunoi, … , Sunao Iyoda, Tadayuki Iwase
Letter to the Editor Infectious disease

Collateral effects of deletion of nlpD on rpoS and rpoS-dependent genes

  • Text
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Abstract

Authors

Manami Tsunoi, Sunao Iyoda, Tadayuki Iwase

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The authors reply:

In their Letter to the Editor, Tsunoi et al. (1) discuss a very interesting and relevant aspect of our published work (2) and suggest topics for further investigation. This published study identifies the E. coli NlpD protein as a potent regulator of gene expression in human cells. This effect is attributed to the inhibition of RNA polymerase II phosphorylation (Pol II), mediated, in part, by effects of NlpD on the Pol II RPB1 subunit and PAF1C (2, 3). The discovery of the NlpD effect was made possible by the isolation of a loss-of-function mutant (SN25) from a human carrier of the parent strain E. coli 83972, and a mutation strategy was devised to introduce the nlpD point mutation from SN25 into the parent strain and to complement SN25 with a fully functional nlpD gene cluster. To address whether NlpD exerts its effects on host cells independently or in synergy with sigma38 (σ38), the 5′nlpD coding sequence was deleted in E. coli 83972, while leaving the rpoS promoter intact. The partial nlpD deletion mutant lost the ability to inhibit PAF1C and Pol II phosphorylation.

In the E. coli genome, nlpD and rpoS form an operon. σ38 expression is regulated, in part, from a promoter located within nlpD. Elegant studies addressing the role of σ38 as a global regulator of bacterial gene expression are referenced in the letter by Tsunoi et al. (1). Global analyses of the rpoS regulon in E. coli K-12 have identified more than 1000 genes (4, 5). As pointed out by Dr. Iwase and colleagues, effects on σ38 are therefore likely to affect bacterial phenotypes, under a variety of conditions. A loss of σ38 expression changes metabolic activity or virulence in uropathogenic E. coli (6). Our preliminary comparison of E. coli 83972 and SN25 gene expression profiles indicates that typical σ38-dependent stress response genes are downregulated in SN25 relative to the 83972 wild-type strain.

Importantly, the effects of NlpD on human cells were characterized using recombinant NlpD protein rather than whole bacteria. Therapeutic efficacy was demonstrated as inhibition of inflammation and accelerated bacterial clearance from infected tissues in a murine urinary tract infection model (Figure 1). In contrast to NlpD, recombinant σ38 displayed no affinity for RPB1 or PAF1C in pull-down experiments. Instead, the data suggest that, by inhibiting TBP binding, σ38 itself may act as an additional regulator of gene expression in infected hosts (2).

Recombinant NlpD treatment accelerated bacterial clearance in CY17-infectedFigure 1

Recombinant NlpD treatment accelerated bacterial clearance in CY17-infected C57BL/6 mice (A) and attenuated inflammation (B). Data are presented as mean ± SEM (n = 5 mice). Two-way ANOVA with Sidak’s multiple comparison tests, *P < 0.05, **P < 0.01, ***P < 0.001.

Thanks to Drs. Tsunoi, Iyoda, and Iwase for initiating this interesting discussion.

Footnotes

Conflict of interest: IA and CS are inventors on patents related to the therapeutic use of NlpD (US 16/341,962, Europe 17797991.1, Australia 2017344453, India 20194718316, Singapore 11201903336U). IA and CS are shareholders of SelectImmune Pharma, a biotech startup company developing alternatives to antibiotics in UTI treatment, and CS is chairman of the board.

Reference information: J Clin Invest. 2021;131(18):e153234. https://doi.org/10.1172/JCI153234.

See the related article at Active bacterial modification of the host environment through RNA polymerase II inhibition.

See the related article at Collateral effects of deletion of nlpD on rpoS and rpoS-dependent genes.

References
  1. Tsunoi M, Iyoda S, Iwase T. Collateral effects of deletion of nlpD on rpoS and rpoS-dependent genes. J Clin Invest. 2021;131(18):e152693.
  2. Ambite I, et al. Active bacterial modification of the host environment through RNA polymerase II inhibition. J Clin Invest. 2021;131(4):e140333.
    View this article via: JCI CrossRef PubMed Google Scholar
  3. Lutay N, et al. Bacterial control of host gene expression through RNA polymerase II. J Clin Invest. 2013;123(6):2366–2379.
    View this article via: JCI CrossRef PubMed Google Scholar
  4. Cho BK, et al. Genome-scale reconstruction of the sigma factor network in Escherichia coli: topology and functional states. BMC Biol. 2014;12(4):4.
    View this article via: PubMed Google Scholar
  5. Wong GT, et al. Genome-wide transcriptional response to varying RpoS levels in Escherichia coli K-12. J Bacteriol. 2017;199(7):755–771.
    View this article via: PubMed Google Scholar
  6. Zlatkov N, Uhlin BE. Absence of global stress regulation in Escherichia coli promotes pathoadaptation and novel c-di-GMP-dependent metabolic capability. Sci Rep. 2019;9(1):2600.
    View this article via: CrossRef PubMed Google Scholar
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