Salmonella- induced SIRT1 and SIRT3 are crucial for maintaining the metabolic switch in bacteria and host for successful pathogenesis

  1. Dipasree Hajra
  2. Raju S. Rajmani
  3. Ayushi Devendrasingh Chaudhary
  4. Shashi Kumar Gupta
  5. Dipshikha Chakravortty

  • Curated by eLife

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    eLife assessment

    This study presents valuable findings on the role of the sirtuins SIRT1 and SIRT3 during Salmonella Typhimurium infection. Although the work increases our understanding of the mechanisms used by this pathogen to interact with its host and may have implications for other intracellular pathogens, the reviewers found that the evidence to support the claims is incomplete. In particular, the discrepancy between results obtained using cultured cell lines and the animal model of infection stands out.

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Abstract

Sirtuins are the major players in host immuno-metabolic regulation. However, the role of sirtuins in the modulation of the immune metabolism pertaining to Salmonellosis is largely unknown. Here, our investigation focussed on the role of two important sirtuins, SIRT1 and SIRT3, shedding light on their impact on intracellular Salmonella ’s metabolic switch and pathogenesis establishment. Our study indicated the ability of the live Salmonella Typhimurium to differentially regulate the levels of SIRT1 and SIRT3 for maintaining the high glycolytic metabolism and low fatty acid metabolism in Salmonella . Perturbing SIRT1 or SIRT3 through knockdown or inhibition, resulted in a remarkable shift in the host metabolism to low fatty acid oxidation and high glycolysis. This switch led to decreased proliferation of Salmonella in the macrophages. Further, Salmonella -induced higher levels of SIRT1 and SIRT3 led to a skewed polarization state of the macrophages from a pro-inflammatory M1 state toward an immunosuppressive M2 making it more conducive for the intracellular life of Salmonella . Alongside, governing immunological functions by modulating p65 NF-κB acetylation, SIRT1, and SIRT3 also skew Salmonella- induced host metabolic switch by regulating the acetylation status of HIF-1α and PDHA1. Interestingly, though knock-down of SIRT1/3 attenuated Salmonella proliferation in macrophages, in in vivo mice-model of infection, inhibition or knockdown of SIRT1/3 led to more dissemination and higher organ burden which can be attributed to enhanced ROS and IL-6 production. Our study hence reports for the first time that Salmonella modulates SIRT1/3 levels to maintain its own metabolism for successful pathogenesis.

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  1. Version published to 10.1101/2022.11.21.517246v4 on bioRxiv
  2. Version published to 10.7554/elife.93125 on eLife
  3. Version published to 10.7554/elife.93125.1 on eLife
  4. eLife

    eLife assessment

    This study presents valuable findings on the role of the sirtuins SIRT1 and SIRT3 during Salmonella Typhimurium infection. Although the work increases our understanding of the mechanisms used by this pathogen to interact with its host and may have implications for other intracellular pathogens, the reviewers found that the evidence to support the claims is incomplete. In particular, the discrepancy between results obtained using cultured cell lines and the animal model of infection stands out.

  5. eLife

    Reviewer #1 (Public Review):

    Summary:
    The current manuscript by Hajra et al deals with the role of the prominent Sirtuins SIRT1 and -3 during infection of macrophages with Salmonella Typhimurium (ST). Apparently, ST infection induces upregulation of host cell SRTs to aid its own metabolism during the intracellular lifestyle and to help reprogramming macrophage polarization. The manuscript has two parts, namely one part that deals with Salmonella infection in cells, where RAW 264.7 murine macrophage-like cells, sharing some features with primary macrophages, were employed. Infected RAW cells displayed a tendency to polarize towards wound-healing M2 and not inflammatory M1 macrophages, which was dependent on SRT. Consequently, the inflammatory response in RAW was more robust in the absence of SRT. Moreover, loss of SRTs leads to impaired bacterial proliferation in these cells, which was attributed to defects in metabolic adaption of the bacteria in the absence of SRT-activity and to the increased M1 inflammatory response.

    Unfortunately, the line of argumentation remains incomplete because corresponding assays in mice showed the opposite result as compared to the experiments using RAW 264.7 cells. i.e. loss of SRTs leads to increased bacterial load in animals (versus impaired proliferation in RAW 264.7 cells). The authors cannot explain this discrepancy.

    Strengths:
    Extensive analysis of Salmonella infection in RAW macrophage-like cells and mice in the context of SRT1/3 function.

    Weaknesses:
    Lack of connection between the cell-based and organismic data, which are not supportive of each other.

  6. eLife

    Reviewer #2 (Public Review):

    Dipasree Hajra et al demonstrated that Salmonella was able to modulate the expression of Sirtuins (Sirt1 and Sirt3) and regulate the metabolic switch in both host and Salmonella, promoting its pathogenesis. The authors found Salmonella infection induced high levels of Sirt1 and Sirt3 in macrophages, which were skewed toward the M2 phenotype allowing Salmonella to hyper-proliferate. Mechanistically, Sirt1 and Sirt3 regulated the acetylation of HIF-1alpha and PDHA1, therefore mediating Salmonella-induced host metabolic shift in the infected macrophages. Interestingly, Sirt1 and Sirt3-driven host metabolic switch also had an effect on the metabolic profile of Salmonella. Counterintuitively, inhibition of Sirt1/3 led to increased pathogen burdens in an in vivo mouse model. Overall, this is a well-designed study. There are a few comments below that would further strengthen the current study.

    Major comments:
    In the in vivo study (lines 436-446) - the authors noticed increased pathogen burden in the EX-527 or the 3TYP-treated mice cohorts but decreased pathogen burden within the F4/80+ macrophage population. What are the other cell types that have increased pathogen burden in splenocytes from EX-527 or the 3TYP treated? Can this be further explored and explained?

    While the authors indicated that IL-6 cytokine storm and elevated ROS production could result in bacterial dissemination in vivo, one could also argue that Sirt1/3 inhibitors might have an impact on gut function and/or gut microbiota (PMID: 22115311). Did Sirt1/3 inhibitors also lead to increased pathogen burdens in the gut? If so, the potential effect of these in vivo treatments on gut microbiota/colonization resistance should be discussed.

    Minor comment:
    Sirt1 has been shown to be degraded during Salmonella infection (PMID: 28192515), which is different from the current study. An explanation should be provided for this.

  7. eLife

    Reviewer #3 (Public Review):

    Summary:
    In this paper, Hajra et al have attempted to identify the role of Sirt1 and Sirt3 in regulating metabolic reprogramming and macrophage host defense. They have performed gene knockdown experiments in RAW macrophage cell lines to show that depletion of Sirt1 or Sirt3 enhances the ability of macrophages to eliminate Salmonella Typhimurium. However, in mice, inhibition of Sirt1 resulted in dissemination of the bacteria but the bacterial burden was still reduced in macrophages. They suggest that the effect they have observed is due to increased inflammation and ROS production by macrophages. They also try to establish a weak link with metabolism. They present data to show that the switch in metabolism from glycolysis to fatty acid oxidation is regulated by acetylation of Hif1a, and PDHA1.

    Strengths:
    The strength of the manuscript is that the role of Sirtuins in host-pathogen interactions has not been previously explored in-depth making the study interesting. It is also interesting to see that depletion of either Sirt1 or Sirt3 results in a similar outcome.

    Weaknesses:
    The major weakness of the paper is the low quality of data, making it harder to substantiate the claims. Also, there are too many pathways and mechanisms being investigated. It would have been better if the authors had focussed on either Sirt1 or Sirt3 and elucidated how it reprograms metabolism to eventually modulate host response against Salmonella Typhimurium. Experimental evidence is also lacking to prove the proposed mechanisms. For instance, they show correlative data that the knockdown of Sirt1-mediated shift in metabolism is due to HIF1a acetylation but this needs to be proven with further experiments.

  8. Version published to 10.1101/2022.11.21.517246v3 on bioRxiv
  9. Version published to 10.1101/2022.11.21.517246v2 on bioRxiv
  10. Version published to 10.1101/2022.11.21.517246v1 on bioRxiv