https://orcid.org/0000-0002-7982-9764
Peer Review History
| Original SubmissionMay 26, 2021 |
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Dear Dr Przyborski, We received your recent submission to Reviews Commons, along with the reviewers' critiques, for consideration for publication in PLOS Pathogens. In light of the generally positive reviews and the appropriateness of the subject, we would like to invite the resubmission of a significantly-revised version that takes into account the reviewers' comments. We cannot make any decision about publication until we have seen the revised manuscript and your response to the reviewers' comments. Your revised manuscript is also likely to be sent to reviewers for further evaluation. When you are ready to resubmit, please upload the following: [1] A letter containing a detailed list of your responses to the review comments and a description of the changes you have made in the manuscript. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out. [2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file). Important additional instructions are given below your reviewer comments. Please prepare and submit your revised manuscript within 60 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email. Please note that revised manuscripts received after the 60-day due date may require evaluation and peer review similar to newly submitted manuscripts. Thank you again for your submission. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments. Sincerely, Kirk W. Deitsch Section Editor PLOS Pathogens Kirk Deitsch Section Editor PLOS Pathogens Kasturi Haldar Editor-in-Chief PLOS Pathogens orcid.org/0000-0001-5065-158X Michael Malim Editor-in-Chief PLOS Pathogens *********************** Figure Files: While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org. Data Requirements: Please note that, as a condition of publication, PLOS' data policy requires that you make available all data used to draw the conclusions outlined in your manuscript. Data must be deposited in an appropriate repository, included within the body of the manuscript, or uploaded as supporting information. This includes all numerical values that were used to generate graphs, histograms etc.. For an example see here on PLOS Biology: http://www.plosbiology.org/article/info%3Adoi%2F10.1371%2Fjournal.pbio.1001908#s5. Reproducibility: To enhance the reproducibility of your results, we recommend that you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols |
| Revision 1 |
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Dear Dr Przyborski, Thank you very much for submitting your manuscript "Co-chaperone involvement in knob biogenesis implicates host-derived chaperones in malaria virulence" for consideration at PLOS Pathogens. As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. The reviewers appreciated the attention to an important topic. Based on the reviews, we are likely to accept this manuscript for publication, providing that you modify the manuscript according to the review recommendations. As you will see from the written critiques, all three reviewers were largely satisfied with the revised version of the manuscript that you submitted. However, please address the minor suggestions and comments that the reviewers provided (no additional experiments are requested). Please prepare and submit your revised manuscript within 30 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email. When you are ready to resubmit, please upload the following: [1] A letter containing a detailed list of your responses to all review comments, and a description of the changes you have made in the manuscript. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out [2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file). Important additional instructions are given below your reviewer comments. Thank you again for your submission to our journal. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments. Sincerely, Kirk W. Deitsch Section Editor PLOS Pathogens Kirk Deitsch Section Editor PLOS Pathogens Kasturi Haldar Editor-in-Chief PLOS Pathogens orcid.org/0000-0001-5065-158X Michael Malim Editor-in-Chief PLOS Pathogens *********************** Reviewer Comments (if any, and for reference): 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: In this paper the function of Plasmodium falciparum exported protein PFA66, is investigated by replacing its functionally important dnaJ region with GFP. These modified parasites grew fine but produced elongated knob-like structures, called eknobs, at the surface of the parasites infected RBCs. Knobs are elevated platforms formed by exported parasite proteins at the surface of the infected RBC that are used to display PfEMP1 cytoadherance proteins which help the parasites avoid host immunity. The eknobs still display some PfEMP1 and contain exported proteins such as KAHRP but can no longer facilitate cytoadherence. Complementation of the truncated PFA66 with full length protein restored normal knob morphology however complementation with a non-functional HPD to QPD mutant did not restore normal morphology implying interaction of the PFA66 with a HSP70 possibly of host origin is important for function. The authors investigate the potential interaction of PFA66 with two human RBC HSP70s and an exported HSP70x of parasite origin by showing that a recombinant PFA66 fused to a HSP70 substrate peptide can stimulate the ATPase activity of the 3 HSP70s. It has long been suspected that because parasites export many dnaJ domain proteins into their RBCs that they must be exploiting human HSP70s to promote parasite virulence and survival. This paper now not only shows this is likely but also alludes as to what specific functions ie, knob formation, that the PFA66 protein is involved in. Reviewer #2: This manuscript by Diehl et al reports on the function of the exported J-domain protein PFA66 in remodeling the infected RBC. The authors have largely resolved my earlier concerns and have also provided new data that indicate PFA66 is able to stimulate the ATPase activity of both HSP70s present in the host cytosol. This was carried out by fusing a known HsHSP70 substrate motif from HSF1 to the C-terminus of PFA66, providing both the J-domain protein and substate together so that HSP70-stimulation can be observed even when the J-domain itself is not a bona fide substrate. Using this system in single turn-over ATPase assays, a significant increase in activity for the parasite HSP70x as well as the two host HSP70s present in the RBC cytosol (HsHSP70 and HsHSC70) is observed over the substrate alone. Given that the PFA66 HPD motif is required to rescue the eKnob phenotype and that knockout of HSP70x (the only parasite HSP70 exported to the host cytosol) does not produce a knob phenotype, this result strengthens the implication that PFA66 performs its role through a partner HSP70 of host origin. Reviewer #3: The revised manuscript addresses a key knowledge gap in how Plasmodium parasites create knob-like structures on the surface of the host RBC. The knobs act as antigen-presenting platforms and play a critical role in the pathogenesis of malaria. The data are excellent, experiments are well controlled, and the conclusions are mostly sound. The authors should be commended on discovering an interesting mechanism for Plasmodium knob formation, one I suspect will greatly help understand malaria pathogenesis. The in vitro assays demonstrating the enhancement of human Hsp70’s APTase activity by PFA66 are very well done. In particular, the inclusion of the minimal substrate peptide shows that it is the tripartite complex that is essential for this activation. The assays are well controlled and add much to the discussion. ********** Part II – Major Issues: Key Experiments Required for Acceptance Please use this section to detail the key new experiments or modifications of existing experiments that should be absolutely required to validate study conclusions. Generally, there should be no more than 3 such required experiments or major modifications for a "Major Revision" recommendation. If more than 3 experiments are necessary to validate the study conclusions, then you are encouraged to recommend "Reject". Reviewer #1: I have already critiqued this paper for Review Commons where the major issue was a lack of physical evidence for the interaction of PFA66 with human and parasite HSP70s. Physical interactions of HSPs are highly transient and difficult to capture but the biochemical assays presented largely address the protein interaction issues. Other shortcomings with the PCR and western blot validations of the integration and expression of the GFP insertion into the PFA66 gene have been resolved. Reviewer #2: (No Response) Reviewer #3: (No Response) ********** 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: All the minor issues from the previous version appear to have been addressed. Reviewer #2: Minor comments -Line 122: Please state the expected fusion sizes for the skipped and unskipped as well as the PFA66-GFP fusion in the text and/or in the figure. I would also recommend providing a reference to the observation that these T2A fusions typically only skip ~50% of the time in Plasmodium, such as PMID 24160265 or 31164473. -Line 911: Given that ~50% of the protein is not skipped, I would change “ensured” to something like “mediated”. Also, the * and + designations on the WB should be described in the figure or figure legend. -Figure S1B: it appears the 43 kD marker is pointing to the top of the anti-GFP blot. I assume this should be the bottom of the Aldolase blot. Please adjust. -The complementation line is introduced as ∆PFA[PFA::HA] in line 132 but then this name is not used in lines 185-187 when it is introduced again as a control for the eKnob phenotype. I assume it’s the same line – if so I would refer to it consistently to avoid any confusion. -Were specific length or width thresholds use to quantify knob morphology categories in Figure 2C, 5D and S6B,D,E or was this done by eye? I didn’t find this in the methods or legends – sorry if I missed it but if it is not there, please add clarification on how the quantification was done. -Line 192: “the lumen of these eKnobs was often extremely dense” It appears that it is not the lumen (or core) but the tip of the eKnob that is electron dense by TEM while the lumen of the eKnobs shown in Fig 2D, 3E and S3 is generally less dense. The tomography seems to show a dense core but I’m not seeing it in the TEMs being described here. -Line 219-220: Were these other exported proteins quantified in the same way? The main text here states “no significant difference” which implies a statistical test was applied but the legend for Fig S4 says “no drastic difference”. From the response comments, it appears that the authors were unable to identify a difference by eye in a blinded check of the images. If so, “no significant difference” is misleading and should be removed from the text. -Line 1005: The legend indicates GBP was detected in S1B but it is not there in the figure. -Line 1023-1025: Please state the expected size of the KAHRP-mCherry fusion in the legend. Line 326: references #43 and #44 are not right here. I think you mean Daniyan et al (#39) and Day et al 2019 (which appears to have been inadvertently deleted from the revised manuscript). -Lines 364-366: The authors should note that some NPP phenotypes are obscured by rich media (for example, Gupta et al 2020, PMID: 32069335). The co-chaperone function of PFA66 could play a role in supporting the establishment of PSAC/NPPs at a level that does not produce a fitness cost without nutrient limitation. Seems this warrants a comment in the discussion. Grammar -Line 238: should be “imaged by STED microscopy” -Line 469: “lead to misleading” is repetitive. Consider revising to “produce misleading” -Line 951: “remaining” and “remained” is repetitive. Consider revising. Reviewer #3: Lines 329-228: The JDS isn’t defined, what were the amino acid numbers? Line 655: The ‘S’ in JDS is lowercase unlike in the rest of the manuscript. Line 674: What was the source of ULP1 protease? Line 714: There’s a typo, ‘JDH’ instead of ‘JDS’ ********** PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: Yes: Josh Beck Reviewer #3: No Figure Files: While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org. Data Requirements: Please note that, as a condition of publication, PLOS' data policy requires that you make available all data used to draw the conclusions outlined in your manuscript. Data must be deposited in an appropriate repository, included within the body of the manuscript, or uploaded as supporting information. This includes all numerical values that were used to generate graphs, histograms etc.. For an example see here: http://www.plosbiology.org/article/info%3Adoi%2F10.1371%2Fjournal.pbio.1001908#s5. Reproducibility: To enhance the reproducibility of your results, we recommend that you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols References: Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. |
| Revision 2 |
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Dear Dr Przyborski, We are pleased to inform you that your manuscript 'Co-chaperone involvement in knob biogenesis implicates host-derived chaperones in malaria virulence' has been provisionally accepted for publication in PLOS Pathogens. Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests. Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated. IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript. Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS. Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Pathogens. Best regards, Kirk W. Deitsch Section Editor PLOS Pathogens Kirk Deitsch Section Editor PLOS Pathogens Kasturi Haldar Editor-in-Chief PLOS Pathogens orcid.org/0000-0001-5065-158X Michael Malim Editor-in-Chief PLOS Pathogens *********************************************************** Reviewer Comments (if any, and for reference): |
| Formally Accepted |
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Dear Dr Przyborski, We are delighted to inform you that your manuscript, "Co-chaperone involvement in knob biogenesis implicates host-derived chaperones in malaria virulence," has been formally accepted for publication in PLOS Pathogens. We have now passed your article onto the PLOS Production Department who will complete the rest of the pre-publication process. All authors will receive a confirmation email upon publication. The corresponding author will soon be receiving a typeset proof for review, to ensure errors have not been introduced during production. Please review the PDF proof of your manuscript carefully, as this is the last chance to correct any scientific or type-setting errors. Please note that major changes, or those which affect the scientific understanding of the work, will likely cause delays to the publication date of your manuscript. Note: Proofs for Front Matter articles (Pearls, Reviews, Opinions, etc...) are generated on a different schedule and may not be made available as quickly. Soon after your final files are uploaded, the early version of your manuscript, if you opted to have an early version of your article, will be published online. The date of the early version will be your article's publication date. The final article will be published to the same URL, and all versions of the paper will be accessible to readers. Thank you again for supporting open-access publishing; we are looking forward to publishing your work in PLOS Pathogens. Best regards, Kasturi Haldar Editor-in-Chief PLOS Pathogens orcid.org/0000-0001-5065-158X Michael Malim Editor-in-Chief PLOS Pathogens |
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