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Dram1 confers resistance to Salmonella infection

Samrah Masud, Rui Zhang, Tomasz K. Prajsnar, Annemarie H. Meijer

Preprint posted on March 21, 2021 https://www.biorxiv.org/content/10.1101/2021.03.21.436194v1.full

A Dram(1)-atic response to Salmonella infection

Selected by Josie Gibson

Categories: immunology, microbiology

Background:

Autophagy is a cellular homeostatic process used to degrade and recycle unwanted or damaged cellular components. In addition, specialised autophagy processes can target invading pathogens. A key autophagy protein is Lc3, which marks autophagosomes in canonical autophagy, as well as phagosomes involved in a process known as Lc3-associated phagocytosis (LAP).

Dram1 is an autophagy modulator induced in infection scenarios. It has previously been demonstrated that dram1 promotes a beneficial host response against infection, in which dram1 is required for Lc3 co-localisation to the pathogen (1,2). In this study, the role of Dram1 in Salmonella enterica serovar Typhimurium (S. Typhimurium) zebrafish infection is examined. The authors have previously demonstrated that LAP is the primary autophagy response used to target Salmonella within macrophages, and that loss of LAP reduces host resistance (3). Therefore, the aim of this study was to investigate the role of dram1 in S. Typhimurium infection outcome and in activating an infection-associated LAP response.

Key findings:

The initial finding of the study is that morpholino-mediated knockdown of dram1 in zebrafish leads to increased susceptibility to S. Typhimurium infection, resulting in reduced zebrafish survival and higher bacterial burdens. This finding is later confirmed with the use of a dram1 mutant zebrafish line. In contrast, when dram1 mRNA was injected into larvae (leading to overexpression of dram1), zebrafish survival of S. Typhimurium infection was increased and was associated with a lower bacterial burden. This demonstrates a clear role of dram1 in promoting a beneficial host defence against S. Typhimurium.

Next, the authors determined that Dram1 is required for Lc3 recruitment during S. Typhimurium infection. In cases of dram1 knockdown, Lc3 recruitment to bacterial cells was significantly reduced. This finding was also confirmed using a dram1 mutant line. Furthermore, a significant increase in Lc3 recruitment to Salmonella was shown in larvae overexpressing dram1. Since LAP is known to be the primary autophagy response to Salmonella in macrophages (3), these data suggest that Dram1 promotes a LAP-mediated host response during S. Typhimurium zebrafish infection.

Finally, the likely role of LAP in S. Typhimurium infection was examined in further detail. LAP requires NADPH oxidase activity for phagosomal reactive oxygen species (ROS) production. Therefore, the level of ROS production was examined using a S. Typhimurium ROS biosensor strain, which becomes GFP fluorescent in the presence of ROS. In dram1 mutant larvae, significantly less ROS were detected at sites of S. Typhimurium in comparison to wild-type larvae, suggesting that Dram1 is required for a beneficial LAP response to S. Typhimurium infection.

Why I chose this Preprint:

I chose this manuscript because it focuses on host-pathogen interactions, specifically examining autophagy, an area I am personally interested in. The key finding is that dram1 is required for host resistance against S. Typhimurium infection. The study then examines the role of autophagic machinery, to show that Dram1 promotes a LAP (Lc3-associated phagocytosis) response against S. Typhimurium. This study deepens our understanding of host responses to Salmonella infection, suggesting that dram1 and LAP may be useful therapeutic targets.

Questions to the authors:

  1. The important role of LAP in S. Typhimurium control by macrophages has been previously determined (3). Do you think the beneficial role of Dram1 mediated LAP in S. Typhimurium infection, demonstrated here in zebrafish, is driven primarily through macrophage interactions?
  2. Considering that Salmonella is known to infect intestinal cells, do you think dram1 may play a similar role in autophagy responses in gut epithelial cells?


References

  1. Meijer AH, Van Der Vaart M. DRAM1 promotes the targeting of mycobacteria to selective autophagy. Vol. 10, Autophagy. Landes Bioscience; 2014. p. 2389–91.
  2. Zhang R, Varela M, Forn-Cuní G, Torraca V, van der Vaart M, Meijer AH. Deficiency in the autophagy modulator Dram1 exacerbates pyroptotic cell death of Mycobacteria-infected macrophages. Cell Death Dis. 2020 Apr 1;11(4):1–16.
  3. Masud S, Prajsnar TK, Torraca V, Lamers GEM, Benning M, Van Der Vaart M, et al. Macrophages target Salmonella by Lc3-associated phagocytosis in a systemic infection model. Autophagy. 2019 May 4;15(5):796–812.

 

Posted on: 12th April 2021 , updated on: 21st April 2021

doi: https://doi.org/10.1242/prelights.28195

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Author's response

Annemarie Meijer shared

Thank you for highlighting our preprint! I am pleased to answer your interesting questions.

  1. Yes, we believe that macrophages are the main drivers of Dram1-mediated host resistance in our model. In this preprint, we have not dissected the responses of macrophages and neutrophils separately, so we cannot exclude that Dram1 and LAP also play a role in neutrophils. However, we have previously shown that macrophages get infected and become Lc3-positive about three times more frequently than neutrophils, suggesting that macrophage-Salmonella interactions predominate during intravenous infection. Similarly, in Mycobacterium marinum infection, although not excluding a role for neutrophils, we have shown that the Dram1-mediated autophagy in macrophages is critical for preventing macrophage cell death and exacerbation of the infection.
  2. We know that Dram1 expression is enriched in leukocytes but are not sure about its abundance in gut epithelial cells. Nevertheless, it is interesting to speculate about the answer to this question. While we have found that LAP is the predominant degradation pathway in macrophages, there is much evidence that epithelial cells target Salmonella by selective autophagy. Dram1 could indeed have a similar role in this response. We hypothesize that Dram1 drives bacteria to lysosomal degradation by promoting the fusion of lysosomes with any type of bacteria-containing vesicles, so autophagosomes as well as LAPosomes.

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