Host succinate is an activation signal for
            <i>Salmonella</i>
            virulence during intracellular infection

Rosenberg, Gili; Yehezkel, Dror; Hoffman, Dotan; Mattioli, Camilla Ciolli; Fremder, Moran; Ben-Arosh, Hadar; Vainman, Leia; Nissani, Noa; Hen-Avivi, Shelly; Brenner, Shirley; Itkin, Maxim; Malitsky, Sergey; Ohana, Ehud; Ben-Moshe, Noa Bossel; Avraham, Roi

doi:10.1126/science.aba8026

Cited by 93 publications

(65 citation statements)

References 40 publications

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“…ProQ suppresses growth in succinate-containing media Succinate has been identified as an important carbon source for Salmonella during colonization of the murine gastrointestinal tract (Spiga et al, 2017), and plays an important role during macrophage infection (Jiang et al, 2021;Rosenberg et al, 2021).…”

Section: Resultsmentioning

confidence: 99%

“…Mineral M9 contained: i) 12.8 g/l Na2HPO4×7H2O, 3 g/l KH2PO4, 0.5 g/l NaCL, 1 g/l NH4Cl, 2mM MgSO4, 0.1 mM CaCl2, 0.0004% Histidine, 0.5 g/ml Thiamine; ii) trace elements: 134 M EDTA, 31 M FeCl3×6H2O, 6.2 M ZnCl2, 0.76 M CuCl2×2H2O, 0.42 M CoCl2×2H2O, 1.62 M H3BO3, 81 nM MnCl2×4H2O; and iii) supplemented with 40 mM Na-succinate (Sigma-Aldrich) as carbon source. Bacterial cultures were inoculated to an OD600 nm 0.01 and incubated at 37°C without aeration for microaerobic conditions as previously described (Rosenberg et al, 2021;Serna et al, 2010) or with 200 rpm shaking for aerobic conditions. For solid growth, Salmonella strains were streaked on LB or M9 Agar supplemented with 40mM Na-succinate as sole carbon source.…”

Section: Bacterial Strains and Growth Conditionsmentioning

confidence: 99%

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Scanning mutagenesis of RNA-binding protein ProQ reveals a quality control role for the Lon protease

Mouali

Ponath

Scharrer

et al. 2021

RNA

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The FinO-domain protein ProQ belongs to a widespread family of RNA-binding proteins (RBPs) involved in gene regulation in bacterial chromosomes and mobile elements. Whilst the cellular RNA targets of ProQ have been established in diverse bacteria, the functionally crucial ProQ residues remain to be identified under physiological conditions. Following our discovery that ProQ deficiency alleviates growth suppression of Salmonella with succinate as the sole carbon source, an experimental evolution approach was devised to exploit this phenotype. By coupling mutational scanning with loss-of-function selection, we identified multiple ProQ residues in both the N-terminal FinO domain and the variable C-terminal region that are required for ProQ activity. Two C-terminal mutations abrogated ProQ function and mildly impaired binding of a model RNA target. By contrast, several mutations in the FinO domain rendered ProQ both functionally inactive and unable to interact with target RNA in vivo. Alteration of the FinO domain stimulated the rapid turnover of ProQ by Lon-mediated proteolysis, suggesting a quality control mechanism that prevents the accumulation of non-functional ProQ molecules. We extend this observation to Hfq, the other major sRNA chaperone of enteric bacteria. The Hfq Y55A mutant protein, defective in RNA-binding and oligomerization, proved to be labile and susceptible to degradation by Lon. Taken together, our findings connect the major AAA+ family protease Lon with RNA-dependent quality control of Hfq and ProQ, the two major sRNA chaperones of Gram-negative bacteria.

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Section: Resultsmentioning

confidence: 99%

Section: Bacterial Strains and Growth Conditionsmentioning

confidence: 99%

Scanning mutagenesis of RNA-binding protein ProQ reveals a quality control role for the Lon protease

Mouali

Ponath

Scharrer

et al. 2021

RNA

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show abstract

“…S12). Itaconate may also be transported into S. Typhimurium by the dicarboxylate transporter DcuB, which is know to transport succinate [45].…”

Section: Resultsmentioning

confidence: 99%

Effect of pH on the antimicrobial activity of the macrophage metabolite itaconate

et al. 2021

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The production of itaconate by macrophages was only discovered in 2011. An increasing number of studies have since revealed essential biological functions for this small molecule, ranging from antimicrobial to immunomodulator. The antibacterial role of itaconate has however been questioned because the estimated concentration of itaconate in macrophages (low-millimolar) is lower than the minimum inhibitory concentration (MIC) of itaconate reported for several bacterial strains (low-to-mid-millimolar). We note that some of these investigations have tended to ignore the high acidity of this small diacid (pKas 3.85 and 5.45), thereby potentially biassing activity measurements. We measured the MIC of itaconate in Escherichia coli (not known to metabolize itaconate) and in Salmonella enterica serovar Typhimurium (known to metabolize itaconate) at varying pH values to probe the effect that pH has on itaconate toxicity. Herein, we demonstrate that the antimicrobial effect of itaconate is dependent upon the pH of the media and that itaconate does have antimicrobial activity at biologically relevant pH and concentrations. Under nutrient-poor conditions, the antimicrobial activity of itaconate in both E. coli and S. Typhimurium increased approximately 200-fold when the pH was dropped by one unit, whereas itaconate was not found to be toxic under nutrient rich conditions. Our results also reveal that the activity of itaconate is synergistic with acidity, yet is not a function of increased permeability with protonation. Similar experiments performed with succinate (a pKa-matched diacid) yielded drastically different results, consistent with a target-based mechanism of action for itaconate. Overall, our work shows the importance of controlling the pH when performing experiments with itaconic acid.

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“…Overall, METLIN-NL empirically derived data will enable new types of analyses facilitating more rapid identification of unknown compounds via both fragment ion and neutral loss similarity searching. 2 Both biologists and chemists will be able applying METLIN-NL to the structure elucidation of unknowns derived from animals, 19 plants, 14,20 or microbiota 21 ; and METLIN-NL can also be used as a resource for identifying unexpected synthetic chemical or enzymatically modified drug products (e.g. pharmaceuticals 22 ) as it is populated with both biological and chemical entities.…”

Section: Figure 2 Ms 2 and Neutral Loss Data On Two Related Oxylipins (16 Keto 16-b1-phytop And 16-b1-phytop) And The Statin Drugs Rosuvamentioning

confidence: 99%

METLIN Neutral Loss Database Enhances Similarity Analysis

Siuzdak

Aisporna

et al. 2021

Preprint

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Tandem mass spectrometry (MS2) data is an effective resource for the identification of known molecules and the putative identification of novel, previously uncharacterized molecules (unknowns). Yet, MS2 data alone is limited in characterizing structurally closely related molecules with different masses. Neutral loss data is key in retrieving this structural similarity. To facilitate unknown identification and complement METLIN fragment ion data for characterizing structurally related molecules, we have created the METLIN neutral loss database (https://metlin-nl.scripps.edu).

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Host succinate is an activation signal for Salmonella virulence during intracellular infection

Cited by 93 publications

References 40 publications

Scanning mutagenesis of RNA-binding protein ProQ reveals a quality control role for the Lon protease

Scanning mutagenesis of RNA-binding protein ProQ reveals a quality control role for the Lon protease

Effect of pH on the antimicrobial activity of the macrophage metabolite itaconate

METLIN Neutral Loss Database Enhances Similarity Analysis

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