PPATHOGENS-D-26-00464

p1/s1, a 3'-nucleotidase/nuclease, allows Leishmania major to circumvent host innate immune response mechanisms

PLOS Pathogens

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Dominique Soldati-Favre

Section Editor

PLOS Pathogens

Sumita Bhaduri-McIntosh

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0003-2946-9497

Michael Malim

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0002-7699-2064

Additional Editor Comments:

Based on the reviewers’ reports, the manuscript requires minor revisions before it can be accepted for publication.

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Reviewers' Comments:

Reviewer's Responses to Questions

Part I - Summary

Please use this section to discuss strengths/weaknesses of study, novelty/significance, general execution and scholarship.

Reviewer #1: This is a revised version of a manuscript that describes the role of p1/s1 nucleotidases/nucleases have in modulating the immune response to Leishmania. They use a cutting edge approach to delete a large region of the genome to delete the p1/s1 array and show that this array is important for digesting NETs. They additionally show that Leishmania utilizes adenosine to reduce the immune reponse.

I previously reveiwed this manuscript through Review Commons and they have responded well to my comments I had previously raised. Clarifications to the text and figures have been made and additional data has been included.

Reviewer #2: (No Response)

Reviewer #3: The authors responded to all my queries

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Part II – Major Issues: Key Experiments Required for Acceptance

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Reviewer #1: Generally, they have also responded appropriately to the comments from the other reviewers. However, I think there are some additional clarifications required:

- For the additional wording in the methodology (lines 99-101) in which they explain the composition of the stationary phase culture can a reference to figure S1 be included for the apoptotic-like cells and a representative set of PNA data shown for the metacyclic rate.

- For the comment about the compensatory mechanism, I looked at the data presented in the thesis the authors linked to and it did not seem to include RNAseq data from the KO cell lines. It just had expression levels for the different genes in the p1/s1 array in log, stationary phase and axenic amastigotes. The mechanism for compensation is not the focus of this manuscript so in my opinion it is not necessary to look at but I just wanted to clarify which data they were alluding to in that thesis.

- In response to Reviewer 3’s comment about the 50% of stationary phase that are dead or undergoing cell death, the authors state the in their culture conditions they see 50-60% dead parasites but in the new section of the methods they are calling this apoptotic-like. Similarly, in the response to Comment 5 that focuses on Figure 4F, the authors state that the stationary phase cultures have a comparable proportion of dead parasites (+/- 5%). In this context are the dead parasites the 50-60% apoptotic-like cells? Moreover, in the text (lines 371-381) the authors state that miltefosine induces 'early cell death processes' and stationary phase parasites 'undergo intrinsic cell death'. Can the authors clarify whether these are dead parasites or still alive but expressing apoptotic markers? There are a number of different phrases in the manuscript and the responses being used and I think it is important that there is consistency.

Finally, would the authors include an example gel to supplement Figure 5 which shows the endonuclease activity of the addback. They have it for the parentals and the KOs but not the addback. This could be included in a supplementary figure.

Reviewer #2: (No Response)

Reviewer #3: The authors responded to all my queries

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Part III – Minor Issues: Editorial and Data Presentation Modifications

Please use this section for editorial suggestions as well as relatively minor modifications of existing data that would enhance clarity.

Reviewer #1: (No Response)

Reviewer #2: (No Response)

Reviewer #3: The authors responded to all my queries

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Reviewer #2: No

Reviewer #3: No

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Dear Dr. van Zandbergen,

We are pleased to inform you that your manuscript 'p1/s1, a 3’-nucleotidase/nuclease, allows Leishmania major to circumvent host innate immune response mechanisms' has been provisionally accepted for publication in PLOS Pathogens.

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Best regards,

Dominique Soldati-Favre

Section Editor

PLOS Pathogens

Dominique Soldati-Favre

Section Editor

PLOS Pathogens

Sumita Bhaduri-McIntosh

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0003-2946-9497

Michael Malim

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0002-7699-2064

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Reviewer Comments (if any, and for reference):