Alzheimer’s risk factor FERMT2 promotes the progression of colorectal carcinoma via Wnt/β-catenin signaling pathway and contributes to the negative correlation between Alzheimer and cancer

Wenzhen Xia; Zhaoyu Gao; Xia Jiang; Lei Jiang; Yushi Qin; Di Zhang; Pei Tian; Wanchang Wang; Qi Zhang; Rui Zhang; Nan Zhang; Shunjiang Xu

doi:10.1371/journal.pone.0278774

Peer Review History

Original SubmissionSeptember 10, 2022
12 Oct 2022 Decision Letter - Frédéric André, Editor PONE-D-22-25240Alzheimer’s risk factor FERMT2 promotes the progression of colorectal carcinoma via Wnt/β-catenin signaling pathway and contributes to the negative correlation between Alzheimer and cancerPLOS ONE Dear Dr.Xu, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Dear Doctor Xu Your manuscript has been evaluated by a reviewer and by me, as Academic Editor. It is a complete manuscript with a lot of data. However, some issues need to be addressed. (1) The authors should explain why - at least in the discussion part - some websites such as GEPIA (http://gepia.cancer-pku.cn/detail.php?gene=FERMT2) indicate that FERMT2 is under expressed in colorectal cancers compared to healthy tissue. These results seem to be in disagreement with your observations. (2) The authors should analyze the impact of FERMT2 knockdown on the expression or activation of beta1 integrin subunit. (3) The authors should respond to the reviewer's comment concerning the purity of the AB1-42 peptide (4) Collective cell migration plays an important role in the progression of colorectal cancer. However, the results presented in the manuscript only show single cell migration. The impact of FERMT2 in the collective migration could be assessed by the authors using a wound healing test.Dear Doctor Xu Please submit your revised manuscript by 26 November 2022. 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Additional Editor Comments : Dear Doctor Xu Your manuscript has been evaluated by a reviewer and by me, as Academic Editor. It is a complete manuscript with a lot of data. However, some issues need to be addressed. (1) The authors should explain why - at least in the discussion part - some websites such as GEPIA (http://gepia.cancer-pku.cn/detail.php?gene=FERMT2) indicate that FERMT2 is under expressed in colorectal cancers compared to healthy tissue. These results seem to be in disagreement with your observations. (2) The authors should analyze the impact of FERMT2 knockdown on the expression or activation of beta1 integrin subunit. (3) The authors should respond to the reviewer's comment concerning the purity of the AB1-42 peptide (4) Collective cell migration plays an important role in the progression of colorectal cancer. However, the results presented in the manuscript only show single cell migration. The impact of FERMT2 in the collective migration could be assessed by the authors using a wound healing test. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: This manuscript examine the relationship FERMT2, colorectal carcinoma (CRC) and Alzheimer disease (AD). FERMT2 levels are elevated in CRC and with poor prognostic outcomes. Knockdown of FERMT2 suppresses key tumorigenic responses in CRC cell lines, and these effects are linked to suppression of Wnt/β-catenin signaling. Treatment with the AD associated peptide, AB1-42 peptide, decreased FERMT2 levels in CRC levels and Wnt signaling, suggesting a basis for an inverse correlation between CRC and AD. Overall, the study is well-done, and an extensive body of data are presented. However, some issues need to be addressed: 1) By far, the most prominent role of FERMT2 is its regulation of integrin activity, much more so than its effects on Wnt/β-catenin signaling. Does knockdown of FERMT2 in CRC cells reduce the expression or activation of integrins on CRC cell lines? It would be sufficient to examine the expression and activity of β1 integrins on the cells by FACS using commercially available antibodies. 2) The AB1-42 is reported to be high purity but was the peptide tested for non-peptide contaminants. Was the peptide free of endotoxin or other non-peptide that could activate/inactivate cells? ****** 6. 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? 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Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0278774.r001
Revision 1
19 Nov 2022 Author Response Dear reviewers and editors, Thank you very much for giving us an opportunity to revise our manuscript entitled “Alzheimer’s risk factor FERMT2 promotes the progression of colorectal carcinoma via Wnt/β-catenin signaling pathway and contributes to the negative correlation between Alzheimer and cancer”. (PONE-D-22-25240). We really appreciate the editors and reviewers for your helpful and constructive comments and suggestions on our manuscript. We have studied the reviewer’s comments carefully and have made revision which marked in red in the paper. We have tried our best to revise our manuscript according to the comments. We would like to express our great appreciation to editors and reviewers for comments on our paper. Below the comments of the reviewers are response point by point and the revisions are indicated. We hope our revised manuscript will meet with the approval of editors to publish it in PLoS ONE. Thank you! Best regards. Yours sincerely, Shunjiang Xu Response to reviewers and editors Comment (1): The authors should explain why - at least in the discussion part - some websites such as GEPIA (http://gepia.cancer-pku.cn/detail.php?gene=FERMT2) indicate that FERMT2 is under expressed in colorectal cancers compared to healthy tissue. These results seem to be in disagreement with your observations. Response: We are grateful to you for this suggestion. Indeed, the results of FERMT2 expression in colorectal cancers are inconsistent from different databases. The reasons may be related to pathological classification, progress, patient population, detection window and other factors. In this study, we firstly analyzed the expression of FERMT2 protein using the public database (Oncomine), and then we have verified the results in patients with colorectal cancer from our cohort, as well as in colorectal cancer cells. Both the bioinformatics analysis results and our detection indicated that the FERMT2 expression was up-regulated in CRC. We have discussed this disagreement in our revised manuscript. Comment (2): The authors should analyze the impact of FERMT2 knockdown on the expression or activation of beta1 integrin subunit. (By far, the most prominent role of FERMT2 is its regulation of integrin activity, much more so than its effects on Wnt/β-catenin signaling. Does knockdown of FERMT2 in CRC cells reduce the expression or activation of integrins on CRC cell lines? It would be sufficient to examine the expression and activity of β1 integrins on the cells by FACS using commercially available antibodies.) Response: Thank you very much for your valuable comments. Integrins, composed of α- and β-subunits, mediate recognition and adhesion between cells or between cells and extracellular matrix. Several studies have shown that β1 integrins can regulate the proliferation, migration and invasion of tumor cells through different signal pathways. FERMT2 has been shown to modulate β1 integrins activity in cancers. According to the reviewer’s suggestion, we have examined the expression of β1 integrins on SW1116 and SW1463 cells after knockdown of FERMT2 by FACS. The results indicated that knockdown of FERMT2 reduced the expression of β1 integrin in CRC cell lines, suggesting that β1 integrin subunit is involved in the FERMT2 mediated progression in CRC. We have provided the results as supplementary data (S3 Fig A-D) and discussed it in our revised manuscript. S3 Fig. Knockdown of FERMT2 inhibited the expression of β1 integrins in SW1116 or SW1463 cells. (A-D) The expression of β1 integrins in SW1116 cells (A, B) or SW1463 cells (C, D) after FERMT2 knockdown was measured by FACS. P < 0.05 vs shNC. Comment (3): The authors should respond to the reviewer's comment concerning the purity of the Aβ1-42 peptide. (The Aβ1-42 is reported to be high purity but was the peptide tested for non-peptide contaminants. Was the peptide free of endotoxin or other non-peptide that could activate/inactivate cells?) Response: Thank you for your question. Firstly, we didn’t test the non-peptide contaminants of the Aβ1-42 peptide we used in this study. According to the certificate of analysis, the purity of Aβ1-42 is 98.93 %. In addition, we have contacted the biosynthesis company and they said that the Aβ1-42 peptide was synthesized by chemical synthesis method. The materials used for chemical synthesis include amino acids, resins, and some other chemical reagents, but do not include Escherichia coli, which can produce endotoxin. Therefore, the Aβ1-42 peptide is free of endotoxin or other non-peptide that could activate/inactivate cells. Comment (4): Collective cell migration plays an important role in the progression of colorectal cancer. However, the results presented in the manuscript only show single cell migration. The impact of FERMT2 in the collective migration could be assessed by the authors using a wound healing test. Response: Thank you for your constructive comment. According to the suggestion of the reviewers, we have detected the ability of collective migration in SW1116 and SW1463 cells after knockdown of FERMT2 by wound healing assay. The results indicated that silencing FERMT2 remarkably inhibited the collective cell migration in SW1116 and SW1463 cells (Fig 6A-B). The cell migration ability detected by transwell assay or wound healing assay showed the same decreased trend in CRC cells after knockdown of FERMT2. In order to control the experimental error, we used transwell assay to detect the cell migration ability in the subsequent experiments. Fig 6. Knockdown of FERMT2 inhibited the migration, invasion and EMT in SW1116 or SW1463 cells. (A-D) The migration ability of SW1116 or SW1463 cells after FERMT2 knockdown were detected by wound healing assay (A, B) or transwell assay (C, D). Scale bars: 100 μm. (E-H) The expression of E-cadherin, N-cadherin and Vimentin in SW1116 (E, F) or SW1463 (G, H) cells after FERMT2 knockdown were measured by western blot assay. All the tests were repeated at least three times. P < 0.05, *P < 0.01vs shNC. Attachments Attachment Submitted filename: Respone to Reviewers.docx* https://doi.org/10.1371/journal.pone.0278774.r002
23 Nov 2022 Decision Letter - Frédéric André, Editor Alzheimer’s risk factor FERMT2 promotes the progression of colorectal carcinoma via Wnt/β-catenin signaling pathway and contributes to the negative correlation between Alzheimer and cancer PONE-D-22-25240R1 Dear Dr. Xu, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Frédéric André Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0278774.r003
Formally Accepted
28 Nov 2022 Acceptance Letter - Frédéric André, Editor PONE-D-22-25240R1 Alzheimer’s risk factor FERMT2 promotes the progression of colorectal carcinoma via Wnt/β-catenin signaling pathway and contributes to the negative correlation between Alzheimer and cancer Dear Dr. Xu: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Frédéric André Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0278774.r004

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