Dear Eunjoon,

Thank you for submitting your revised manuscript entitled "Early correction of synaptic long-term depression improves abnormal anxiety-like behavior in adult GluN2B-C456Y-mutant mice" for consideration as a Research Article by PLOS Biology.

Your revision has now been evaluated by the PLOS Biology editorial staff, as well as by the original Academic Editor, and I am writing to let you know that we would like to send your submission out for external peer review.

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Kind regards,

Gabriel Gasque, Ph.D.,

Senior Editor

PLOS Biology

Dear Eunjoon,

Thank you for submitting your revised Research Article entitled "Early correction of synaptic long-term depression improves abnormal anxiety-like behavior in adult GluN2B-C456Y-mutant mice" for publication in PLOS Biology. I have now obtained advice from the original reviewers and have discussed their comments with the Academic Editor.

Based on the reviews, we will probably accept this manuscript for publication, assuming that you will modify the manuscript to address the remaining points raised by reviewer 2. Please also make sure to address the data and other policy-related requests noted at the end of this email.

We expect to receive your revised manuscript within two weeks. Your revisions should address the specific points made by reviewer 2. Please submit the following files along with your revised manuscript:

1. A 'Response to Reviewers' file - this should detail your responses to the editorial requests, present a point-by-point response to all of the reviewers' comments, and indicate the changes made to the manuscript.

*NOTE: In your point by point response to the reviewers, please provide the full context of each review. Do not selectively quote paragraphs or sentences to reply to. The entire set of reviewer comments should be present in full and each specific point should be responded to individually, point by point.

2. In addition to a clean copy of the manuscript, please also upload a 'track-changes' version of your manuscript that specifies the edits made. This should be uploaded as a "Related" file type.

In addition to the remaining revisions and before we will be able to formally accept your manuscript and consider it "in press", we also need to ensure that your article conforms to our guidelines. A member of our team will be in touch shortly with a set of requests. As we can't proceed until these requirements are met, your swift response will help prevent delays to publication.

*Copyediting*

Upon acceptance of your article, your final files will be copyedited and typeset into the final PDF. While you will have an opportunity to review these files as proofs, PLOS will only permit corrections to spelling or significant scientific errors. Therefore, please take this final revision time to assess and make any remaining major changes to your manuscript.

NOTE: If Supporting Information files are included with your article, note that these are not copyedited and will be published as they are submitted. Please ensure that these files are legible and of high quality (at least 300 dpi) in an easily accessible file format. For this reason, please be aware that any references listed in an SI file will not be indexed. For more information, see our Supporting Information guidelines:

https://journals.plos.org/plosbiology/s/supporting-information

*Published Peer Review History*

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https://blogs.plos.org/plos/2019/05/plos-journals-now-open-for-published-peer-review/

*Early Version*

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*Submitting Your Revision*

To submit your revision, please go to https://www.editorialmanager.com/pbiology/ and log in as an Author. Click the link labelled 'Submissions Needing Revision' to find your submission record. Your revised submission must include a cover letter, a Response to Reviewers file that provides a detailed response to the reviewers' comments (if applicable), and a track-changes file indicating any changes that you have made to the manuscript.

Please do not hesitate to contact me should you have any questions.

Sincerely,

Gabriel Gasque, Ph.D.,

Senior Editor

PLOS Biology

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DATA POLICY:

--Please update your S3 Data file to include Figure S2C.

-- Please also ensure that figure legends in your manuscript include information on where the underlying data can be found, and ensure your supplemental data file/s has a legend.

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BLOT AND GEL REPORTING REQUIREMENTS:

For manuscripts submitted on or after 1st July 2019, we require the original, uncropped and minimally adjusted images supporting all blot and gel results reported in an article's figures or Supporting Information files. We will require these files before a manuscript can be accepted so please prepare and upload them now. Please carefully read our guidelines for how to prepare and upload this data: https://journals.plos.org/plosbiology/s/figures#loc-blot-and-gel-reporting-requirements

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Reviewer remarks:

Reviewer #1: The authors have adequately addressed the majority of this reviewer's concerns. With the significant changes made to the manuscript during the revision, the paper is suitable for publication.

Reviewer #2, Marc Fuccillo: The authors have attempted to address many of my requests. In doing so, they have strengthened the idea that the C456Y point mutant functions like the NR2B heterozygous LOF mutation, both at the synaptic, protein and behavioral level. While perhaps expected, these data provide clarity to the role of ASD-associated NMDAR mutations. Another strong plus is the addition of data showing a similar plasticity effect in the mPFC. these data suggest that the data in hippo are a proxy for other areas (something that should be mentioned in the text).

Overall, I feel like this is a strong contribution to the asd pathophysiology literature. while not incredibly surprising, the data is clear and makes a strong point supporting this specific disease-associated mutation as GRIN2B haploinsufficiency. the reproducibility of DCS rescue across assays is also a strength.

A few short textual things should be added:

1. discuss the relationship between anxiogenic and anxiolytic behaviors as they relate to ASD. these are not part of the core behavioral phenotype but are the most clearly altered in this work. the literature on ASD and anxiety-related behaviors should be discussed.

2. please discuss that in light of the mPFC data, it is unclear which brain regions are contributing to the mutation-associated behavioral (and rescue-related) changes.

3. "suppressed recognition of the fact that a dark and closed place is generally safe" - this isn't any better than before - perhaps just describe the phenotype.

Reviewer #3: The authors performed extra experiments and gave explanations to address all the questions raised by the reviewers. The manuscript became even more data heavy and the conclusions are now better discussed to support the authors ideas. Although there are some technical issues, the authors do demonstrate GluN2B-C456Y haploinsufficiency decreases GluN2B protein levels, LTD, and anxiety-like behavior. The overall behavioral and physiological experiments provide an insight for the importance of early correction of pathophysiological deficits.

The findings are important in the field for the treatment of neurodevelopmental or psychiatric diseases caused by NMDAR mutations.

Reviewer #4: Satisfied with the manuscript as edited. Again, it is an important contribution to the literature as I stated in my initial review, though there are not major new insights here. Addition of the PFC data at least provides some additional evidence for more general deficits.

Dear Dr Kim,

On behalf of my colleagues and the Academic Editor, Thomas C Südhof, I am pleased to inform you that we will be delighted to publish your Research Article in PLOS Biology.

The files will now enter our production system. You will receive a copyedited version of the manuscript, along with your figures for a final review. You will be given two business days to review and approve the copyedit. Then, within a week, you will receive a PDF proof of your typeset article. You will have two days to review the PDF and make any final corrections. If there is a chance that you'll be unavailable during the copy editing/proof review period, please provide us with contact details of one of the other authors whom you nominate to handle these stages on your behalf. This will ensure that any requested corrections reach the production department in time for publication.

Early Version

The version of your manuscript submitted at the copyedit stage will be posted online ahead of the final proof version, unless you have already opted out of the process. 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.

PRESS

We frequently collaborate with press offices. If your institution or institutions have a press office, please notify them about your upcoming paper at this point, to enable them to help maximise its impact. If the press office is planning to promote your findings, we would be grateful if they could coordinate with biologypress@plos.org. If you have not yet opted out of the early version process, we ask that you notify us immediately of any press plans so that we may do so on your behalf.

We also ask that you take this opportunity to read our Embargo Policy regarding the discussion, promotion and media coverage of work that is yet to be published by PLOS. As your manuscript is not yet published, it is bound by the conditions of our Embargo Policy. Please be aware that this policy is in place both to ensure that any press coverage of your article is fully substantiated and to provide a direct link between such coverage and the published work. For full details of our Embargo Policy, please visit http://www.plos.org/about/media-inquiries/embargo-policy/.

Thank you again for submitting your manuscript to PLOS Biology and for your support of Open Access publishing. Please do not hesitate to contact me if I can provide any assistance during the production process.

Kind regards,

Vita Usova

Publication Editor,

PLOS Biology

on behalf of

Gabriel Gasque,

Senior Editor

PLOS Biology