Dear Dr Michelot,

Thank you for submitting your manuscript entitled "Mechanical stiffness of reconstituted actin patches correlates tightly with endocytosis efficiency" for consideration as a Short Reports by PLOS Biology.

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Kind regards,

Di Jiang, PhD

Associate Editor

PLOS Biology

Dear Dr Michelot,

Thank you very much for submitting your manuscript "Mechanical stiffness of reconstituted actin patches correlates tightly with endocytosis efficiency" for consideration as a Short Reports by PLOS Biology. Your paper was evaluated by the PLOS Biology editors as well as by an Academic Editor with relevant expertise and by two reviewers.

Based on the reviews, we will probably accept this manuscript for publication, providing that you will modify the manuscript according to the review recommendations.

We expect to receive your revised manuscript within two weeks. Your revisions should address the specific points made by each reviewer. 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. One requirement is described below under 'DATA POLICY' and is marked with '***IMPORTANT: '. 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.

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Sincerely,

Di Jiang, PhD

Associate Editor

PLOS Biology

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Reviewer remarks:

Reviewer #1: see attached file

Reviewer #2: This manuscript addresses the role of mechanical stiffness of the endocytic actin network in the budding of endocytic vesicles in yeast cells. The mechanical properties of the endocytic protein machinery are a very poorly studied due to significant technical challenges in analyzing these tiny cellular structures. This question, however, is critical for mechanistic understanding of endocytosis, and for understanding other mechanical processes in cells, such as cell migration. The authors use a new and very innovative approach to tackle this question. They use in parallel an in vitro approach and an in vivo approach, and correlate the results from these two complementary approaches. The in vitro assay allows them to directly measure the mechanical stiffness of reconstituted endocytic actin patches with or without key actin filament crosslinkers. The in vivo approach reveals the endocytic functionality of the wildtype and comparable crosslinker mutant strains. The manuscript shows that crosslinker proteins increase the stiffness of the actin patch and that enough stiffness is essential for the proper budding of endocytic vesicles. The results and the approach are very interesting for the field of endocytosis, but also for the broader audience of people working on the cytoskeleton and cellular biomechanics.

The manuscript is very clear and concise. The experiments are well performed and controlled. I have only very minor suggestions for the authors.

- In the abstract use “actin crosslinkers” instead of just “crosslinkers”.

- Fig 1B: label the strains. There are no “black dots” in the fig 1B as stated in the legend.

- Fig 4: Although the model figure is quite clear, it would be helpful to have a figure legend that briefly describes the model.

Attachments

Attachment

Submitted filename: report.docx

Dear Dr Michelot,

On behalf of my colleagues and the Academic Editor, Laura Machesky, I am pleased to inform you that we will be delighted to publish your Short Reports 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.

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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,

Hannah Harwood

Publication Assistant,

PLOS Biology

on behalf of

Di Jiang,

Associate Editor

PLOS Biology