Homosalate and ERK Knockdown in the Modulation of Aurelia coerulea Metamorphosis by Regulating the PI3K Pathway and ERK Pathway

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The title has not specific information. One is that what affects the metamorphosis of jellyfish, another is that through which pathway it is regulated?

line 28-29: There are contradictory statistical data cited.

An inconsistent description: There are 1000 species of jellyfish (line 27-28) and There are two types of jellyfish (line 47-48). How to discriminate the difference between them?

The authors described the positive and negative effects of homosalate, particularly emphasizing its harmfulness, but did not explain the reason for using the substance to study metamorphosis of jellyfish in terms of unwanted impairing on jellyfish. What function do the authors want to use this homosalate for? an agonist or an inhibitor?

Authors need to describe clearly what kind of shrimp species was used for the study, how to hatch shrimp eggs and what stage of shrimp larvae was used to cultivate the polyps. How do you know that polyps have ingested shrimp larvae? What is the sign of completing metamorphosis of polyps?

How to provide a stable supply of budless and hypertrophic polyps to ensure the normal development of metamorphosis? Please provide a detailed feeding protocol of polyps that can continuously provide budless and hypertrophic polyps.

You should clarify whether polyps are fed with larvae of shrimp during metamorphosis treatment with homosalate and YS-49. Please clarify whether homosalate and YS-49 are agonists or inhibitors. Please give an explanation for the simultaneous use of homosalate and YS-49 in that both of them are described as agonists.

You need to analyze changing trend of the metamorphosis with increasing dose of homosalate and SY-49, not just the trend over treatment time in order to achieve suitable dose.

We cannot see matamorphosis delay caused by homosalate or YS-49 treatment over days from Figure 1 and Figure 2. Figures 1-2 needs reconstruction.

See Figure 1, the replicate (n=3 ×10) is showed for each treatment, but this is an inappropriate expression. see in other results section.

Figure 1D should be placed in the method section.

Why do homosalate and YS-49 have different inhibiting effects on metamorphosis of jellyfish although they all regulate metamorphosis by activating PI3K/Akt pathway (Figure 4)?

This part (line 275-283) repeated and conflicted with the introduction.

This part (line 284-287) repeated the introduction.

line 304: The term dual stimulation appears but the practice was not performed in the present experiment.

The present study showed homosalate delays metamorphosis of jellyfish. Does this mean the occurrence of abnormal metamorphosis of jellyfish, which is beneficial for controlling outbreak of jellyfish? However, authors did not draw a distinct conclusion from the present results in terms of effect of homosalate treatment on metamorphosis of jellyfish in discussion section.

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

Recently, jellyfish outbreaks have occurred frequently worldwide, which brought a lot of troubles to the local residents. Metamorphosis is an important process during the life of jellyfish, which could play an important role in the control of the jellyfish outbreak. In the manuscript titled “Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K pathway and Inhibition of the ERK pathway”, the authors demonstrated how MAPK and PI3K pathways interacted and regulated the metamorphosis of Aurelia coerulea. It’s an interesting work. However, lacks of discussion on the other related pathways regulated the metamorphosis of Aurelia coerulea is the major flaw of the study. Therefore, minor revision is recommended before this manuscript could be accepted for publication in Current Issues Molecular Biology(CIMB). 

Specific comments:

1. In the discussion section, other related pathways regulated the metamorphosis of Aurelia coerulea could be added.

2. It’s suggested that “5. Conclusions” section should be provided according to the journal requirement.

Author Response

Dear Editor and Reviewers,

Thank you for your kind letter on September 2 and for the reviewers’ comments concerning our manuscript entitled “Homosalate and ERK Knockdown on the Modulation of Aurelia coerulea Metamorphosis by Regulating PI3K Pathway and ERK Pathway” (Manuscript ID: cimb-3179608). Those comments are all valuable and very helpful for improving our paper. We have studied comments carefully and have made corrections which we hope meet with approval. The revised portions have been highlighted with yellow color in the revised manuscript.

Here below are our point-by-point responses to the issues raised in the peer-review report(s).

Response to Reviewer 2 Comments
1. Summary
Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
2. Questions for General Evaluation	Reviewer’s Evaluation	Response and Revisions
Does the introduction provide sufficient background and include all relevant references?	Yes	Thank you for your positive comment.
Is the research design appropriate?	Yes	Thank you for your positive comment.
Are the methods adequately described?	Can be improved	Thank you. We have improved according to your comments.
Are the results clearly presented?	Yes	Thank you for your positive comment.
Are the conclusions supported by the results?	Can be improved	Thank you. We have improved according to your comments.
3. Point-by-point response to Comments and Suggestions for Authors
Comments 1: Recently, jellyfish outbreaks have occurred frequently worldwide, which brought a lot of troubles to the local residents. Metamorphosis is an important process during the life of jellyfish, which could play an important role in the control of the jellyfish outbreak. In the manuscript titled “Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K pathway and Inhibition of the ERK pathway”, the authors demonstrated how MAPK and PI3K pathways interacted and regulated the metamorphosis of Aurelia coerulea. It’s an interesting work. However, lacks of discussion on the other related pathways regulated the metamorphosis of Aurelia coerulea is the major flaw of the study. Therefore, minor revision is recommended before this manuscript could be accepted for publication in Current Issues Molecular Biology(CIMB).
Response 1: Thank you for your positive comments and suggestions. We have added some discussions on the other related pathways regulated the metamorphosis of Aurelia coerulea according to your comments in the Discussion section as follows: “The Wnt signaling pathway, known for orchestrating body axis formation, and the Hippo pathway, which regulates cell growth and tissue homeostasis, are also integral in jellyfish development [38,39]. Specifically, the ERK branch of the MAPK pathway is responsible for driving critical processes including cell proliferation and differentiation, alongside facilitating regeneration in hydroid species [40]. Additionally, specific inhibitors of PI3K can inhibit bud formation in Hydra [41]. The Notch target genes were necessary for budding in Hydra [42]”.
Comments 2: In the discussion section, other related pathways regulated the metamorphosis of Aurelia coerulea could be added. Response 2: Thank you for your suggestions. We have added some discussions on the other related signaling pathways regulated the metamorphosis of Aurelia coerulea according to your comments in the Discussion section as follows: “The Wnt signaling pathway, known for orchestrating body axis formation, and the Hippo pathway, which regulates cell growth and tissue homeostasis, are also integral in jellyfish development [38,39]. Specifically, the ERK branch of the MAPK pathway is responsible for driving critical processes including cell proliferation and differentiation, alongside facilitating regeneration in hydroid species [40]. Additionally, specific inhibitors of PI3K can inhibit bud formation in Hydra [41]. The Notch target genes were necessary for budding in Hydra [42]”.
Comments 3: It’s suggested that “5. Conclusions” section should be provided according to the journal requirement. Response 3: Thank you for your comments and suggestions. We are sorry for that, and we have added “5. Conclusions” section according to your comments and suggestions as follows: “Homosalate activates PI3K and ERK simultaneously to delay the process of metamorphosis of Aurelia coerulea, and YS-49 inhibits the metamorphosis of Aurelia coerulea by activating PI3K particularly. Additionally, after the ERK (evm.model.CTG_ 2.65) was knocked down, the process of metamorphosis of Aurelia coerulea also slowed down. These results indicate that activating PI3K and Knocking down the expression of ERK (evm.model.CTG_ 2.65) play important roles in delaying the process of metamorphosis of Aurelia coerulea. Both PI3K pathway and ERK pathway intervention may be as potential strategies for prevention and control on jellyfish blooms.”

Response to Comments on the Quality of English Language

Response: We had asked a professional team to polish our revised manuscript for the quality of English language.

 

Additional clarifications

Response to Editorial office’s comments：

Manuscript ID: cimb-3179608

Type of manuscript: Article

Title: Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K Pathway and Inhibition of the ERK Pathway

Authors: Jinhong Chen, Xiaoyu Geng, Bingbing Li, Jinyao Xie, Jieying Ma, Zhen Qin, Mingke Wang *, Jishun Yang *

Received: 11 Sep 2024

Response: Thank the reviewers and the editors again for the effort in shaping our manuscript. Please tell us if extensive editing is required.

 

Best Regards

Ji-Shun Yang, PhD
Naval Medical Center of PLA, Naval Medical University
No.338, Huaihai West Road, Shanghai 200052, China
E-mail: jasunyang@foxmail.com/wmke021@163.com

Reviewer 3 Report

Comments and Suggestions for Authors

 

In the manuscript titled, “Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K Pathway and Inhibition of the ERK Pathway”, aimed to to reveal new strategies for preventing and controlling jellyfish blooms. The paper, however, exhibit certain deficiencies in design and data processing.

 

Specific comments

1. There are logic problems in the introduction of this article. Should the introduction of Metamorphosis be followed by the introduction of Aurelia coerulea, so as to elicit how MAPK and PI3K signaling pathways may affect Metamorphosis. In addition, cases and proportion of Aurelia coerulea stings should be supplemented and highlighted, so as to ensure the representativeness and practical significance of this experiment.

2. YS-49 is an activator of PI3K/Akt, but only homosalate is introduced in this article, and the introduction and research progress of YS-49 are lacking.

3. How to define the transverse cleavage between materials and methods? How is the strobilation rate calculated? Whether there is literature support.

4. ERK molecular knockdown and a different method of mammalian RNA extraction was used in the article, which required literature-based

5. Please briefly describe the process of sample collection and initial processing

6. Primer sequences of target gene and reference gene are missing in this paper, and it is not introduced whether RT-qPCR is repeated to avoid chance, and the specific calculation method is not mentioned.

7. Whether the original data was uploaded to NCBI, please provide the relevant database serial numbers.

8. The data analysis method used in the paper has some issues. The author directly applies one-way ANOVA to the data without considering whether it adheres to the normal distribution or conducting a secondary test for variance, which raises concerns about the accuracy of the results. In addition, it is recommended that a two-way ANOVA method be used to verify whether there is an interaction effect between time and concentration

9. In the article, the effects of homosalate and YS-49 on the metamorphosis of Aurelia Coerulea were verified separately, so why was it not verified whether there was a synergistic effect between the two?

10. Materials and Methods section 2.3 describes the treatment as selecting 1 μM and 10 μM of YS-49, while in Results section 2.3 it is described as 1,3,10, 30, and 50 μM, which is unconvincing!

11. The article is poorly presented, e.g. the samples used for transcriptome sequencing are not clearly described, which is a serious problem.

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 4 Report

Comments and Suggestions for Authors

I found this manuscript dispersive in some parts, with a research question unclear and satisfactory to the reader. Indeed, the scientific soundness and relapses of this study resulted in unclear at the end of the document, or not well enhanced. 

The first part of the introduction, as the first part of the discussion sections, starts with a focus on jellyfish threat to human health and activities. Hence, the focus moves to homosalate as a possible way to alternate the jellyfish life cycle. I think it could represent a rather dangerous speech from an ecological point of view and not anthropocentric.

The study model was not properly presented in the introduction section. In the present form, the reason for selecting this species for this study is unclear from a scientific point of view. Take always care to italicize all the scientific names and not only one of the studied species, which anyway should be better to complete with eponyms at least in the first mention and title.

The discussion section is very synthetic and does not report any real discussion of the data obtained in this study with similar/related organisms, to compare and validate/contrast some results. This led to unknown relapses in the study, strongly decreasing the soundness of this document.

More minor mistakes are present in the manuscript, but not very relevant at this stage.

 

Best regards

Comments on the Quality of English Language

I suggest that the Authors revise the manuscript with the help of an English language expert, to increase its richness and fluency.

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

no comments

Author Response

Dear Editor and Reviewers,

Thank you for your kind letter on September 18 and for the reviewers’ comments concerning our manuscript entitled “Homosalate and ERK Knockdown on the Modulation of Aurelia coerulea Metamorphosis by Regulating PI3K Pathway and ERK Pathway” (Manuscript ID: cimb-3179608). Those comments are all valuable and very helpful for improving our paper. We have studied comments carefully and have made corrections which we hope meet with approval. The revised portions have been highlighted with yellow color in the revised manuscript.

Here below are our point-by-point responses to the issues raised in the peer-review report(s).

Response to Reviewer 1 Comments
1. Summary
Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
2. Questions for General Evaluation	Reviewer’s Evaluation	Response and Revisions
Does the introduction provide sufficient background and include all relevant references?	Yes	Thank you for your comments.
Is the research design appropriate?	Yes	Thank you for your comments.
Are the methods adequately described?	Yes	Thank you for your comments.
Are the results clearly presented?	Yes	Thank you for your comments.
Are the conclusions supported by the results?	Yes	Thank you for your comments.
3. Point-by-point response to Comments and Suggestions for Authors
Comments 1: no comments.
Response 1: Thank you for your comment and effort in shaping our manuscript.

Additional clarifications

Response to Editorial office’s comments：

Manuscript ID: cimb-3179608

Type of manuscript: Article

Title: Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K Pathway and Inhibition of the ERK Pathway

Authors: Jinhong Chen, Xiaoyu Geng, Bingbing Li, Jinyao Xie, Jieying Ma, Zhen Qin, Mingke Wang *, Jishun Yang *

Received: 28 Sep 2024

Response: Thank the reviewers and the editors again for the effort in shaping our manuscript. Please tell us if extensive editing is required.

 

Best Regards

Ji-Shun Yang, PhD
Naval Medical Center of PLA, Naval Medical University
No.338, Huaihai West Road, Shanghai 200052, China
E-mail: jasunyang@foxmail.com/wmke021@smmu.edu.cn

Reviewer 3 Report

Comments and Suggestions for Authors

I am still troubled by the two-way ANOVA conducted by the author. It cannot be seen from the results and images that the author conducted this analysis.The author can refer to the following papers:High-Fat-Diet-Induced Oxidative Stress in Giant Freshwater Prawn…………

https:// doi.org/10.3390/antiox11020228

 

 

Author Response

Dear Editor and Reviewers,

Thank you for your kind letter on September 18 and for the reviewers’ comments concerning our manuscript entitled “Homosalate and ERK Knockdown on the Modulation of Aurelia coerulea Metamorphosis by Regulating PI3K Pathway and ERK Pathway” (Manuscript ID: cimb-3179608). Those comments are all valuable and very helpful for improving our paper. We have studied comments carefully and have made corrections which we hope meet with approval. The revised portions have been highlighted with yellow color in the revised manuscript.

Here below are our point-by-point responses to the issues raised in the peer-review report(s).

Response to Reviewer 3 Comments
1. Summary
Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
2. Questions for General Evaluation	Reviewer’s Evaluation	Response and Revisions
Does the introduction provide sufficient background and include all relevant references?	Can be improved	Thank you. We have improved according to your comments.
Is the research design appropriate?	Can be improved	Thank you. We have improved according to your comments.
Are the methods adequately described?	Can be improved	Thank you. We have improved according to your comments.
Are the results clearly presented?	Must be improved	Thank you. We have improved according to your comments.
Are the conclusions supported by the results?	Can be improved	Thank you. We have improved according to your comments.
3. Point-by-point response to Comments and Suggestions for Authors
Comments 1: I am still troubled by the two-way ANOVA conducted by the author. It cannot be seen from the results and images that the author conducted this analysis.The author can refer to the following papers:High-Fat-Diet-Induced Oxidative Stress in Giant Freshwater Prawn.…………https:// doi.org/10.3390/antiox11020228.
Response 1: Thank you for your comments and suggestions. We have studied the reference you provided: “High-Fat-Diet-Induced Oxidative Stress in Giant Freshwater Prawn. https:// doi.org/10.3390/antiox11020228” carefully, and made some modifications in the ”2.8. Statistical analysis” section, the results and images section referring to your provided reference as follows: Firstly, we have revised the ”2.8. Statistical analysis” section as follows: “The parameter was calculated as follows: Strobilation rate (SR, %)= strobilation number/ total number ×100. Analyses were performed using the GraphPad Prism8.0.2 (GraphPad Software, USA) for Windows. All data are expressed as mean ±standard deviation (SD). The differences between groups were evaluated by a two-way ANOVA method. For all tests, P＜ 0.05 was considered statistically significant”. Secondly, we have added some descriptions in the results section including some analytical descriptions of significant differences in the time and concentration dimension as follows: “In the control group, early strobila appeared on the second day, late strobila appeared on the fourth day, and ephyra appeared on the seventh day; under the treatment of 1μM and 100 nM homosalate, late strobila appeared on the fifth day, and ephyra appeared on the seventh day, or later. There was no significant difference in inhibitory effects between 1 μM homosalate and 100 nM homosalate (Figure. 2)” (Line 253-258). “YS-49 solutions with a final concentration of 1 μM, 3 μM, 10 μM, 30 μM, and 50 μM with a chemical inducer configuration (5-methoxy-2-methylindole, salinity 2.6–3.2, artificial sea water, and YS-49) were selected to treat polyps. In the control group, early strobila appeared on the second day, late strobila appeared on the 5th day, and ephyra appeared on the 7th day; under five different concentrations of YS-49, no late strobila and ephyra appeared in the 10 μM YS-49 group on the 7th day. On the second day, the number of polyp in the control group was 20, and its percentage was 66.67%, while the number of polyp in 1 μM, 3 μM, 10 μM, 30 μM, and 50 μM YS-49 was 27, 27, 30, 27, and 30 respectively, and the percentage was 90%, 90%, 100%, 90%, and 100%, which was significantly higher than the control group. The results showed that, compared with the control group, the YS-49 solutions of five concentrations could delay the progress of metamorphosis and reduce the strobila rate of Aurelia coerulea from the second day. When compared with different doses of YS-49 in our study, the inhibitory effect of 10 μM YS-49 was the most significant, 3 μM YS-49 was the second (Figure. 3)” (Line 266-279). Thirdly, we have studied the reference you mentioned, and made some modifications in the Figure 2, Figure 3, and Figure 4. We hope these relevant revisions which we have made could meet with your approval, and the revised portions have been highlighted with yellow color in the revised manuscript.

 

Additional clarifications

Response to Editorial office’s comments：

Manuscript ID: cimb-3179608

Type of manuscript: Article

Title: Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K Pathway and Inhibition of the ERK Pathway

Authors: Jinhong Chen, Xiaoyu Geng, Bingbing Li, Jinyao Xie, Jieying Ma, Zhen Qin, Mingke Wang *, Jishun Yang *

Received: 28 Sep 2024

Response: Thank the reviewers and the editors again for the effort in shaping our manuscript. Please tell us if extensive editing is required.

 

Best Regards

Ji-Shun Yang, PhD
Naval Medical Center of PLA, Naval Medical University
No.338, Huaihai West Road, Shanghai 200052, China
E-mail: jasunyang@foxmail.com/wmke021@smmu.edu.cn

Reviewer 4 Report

Comments and Suggestions for Authors

Dear Authors,

thanks for considered my previous comments during your revisions.

Author Response

Dear Editor and Reviewers,

Thank you for your kind letter on September 18 and for the reviewers’ comments concerning our manuscript entitled “Homosalate and ERK Knockdown on the Modulation of Aurelia coerulea Metamorphosis by Regulating PI3K Pathway and ERK Pathway” (Manuscript ID: cimb-3179608). Those comments are all valuable and very helpful for improving our paper. We have studied comments carefully and have made corrections which we hope meet with approval. The revised portions have been highlighted with yellow color in the revised manuscript.

Here below are our point-by-point responses to the issues raised in the peer-review report(s).

Response to Reviewer 4 Comments
1. Summary
Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
2. Questions for General Evaluation	Reviewer’s Evaluation	Response and Revisions
Does the introduction provide sufficient background and include all relevant references?	Can be improved	Thank you. We have improved according to your comments.
Is the research design appropriate?	Can be improved	Thank you. We have improved according to your comments.
Are the methods adequately described?	Yes	Thank you for your comments.
Are the results clearly presented?	Yes	Thank you for your comments.
Are the conclusions supported by the results?	Yes	Thank you for your comments.
3. Point-by-point response to Comments and Suggestions for Authors
Comments 1: Dear Authors, thanks for considered my previous comments during your revisions.
Response 1: Thank you for your comment and effort in shaping our manuscript.

Additional clarifications

Response to Editorial office’s comments：

Manuscript ID: cimb-3179608

Type of manuscript: Article

Title: Modulation of Aurelia coerulea Metamorphosis through Activation of the PI3K Pathway and Inhibition of the ERK Pathway

Authors: Jinhong Chen, Xiaoyu Geng, Bingbing Li, Jinyao Xie, Jieying Ma, Zhen Qin, Mingke Wang *, Jishun Yang *

Received: 28 Sep 2024

Response: Thank the reviewers and the editors again for the effort in shaping our manuscript. Please tell us if extensive editing is required.

 

Best Regards

Ji-Shun Yang, PhD
Naval Medical Center of PLA, Naval Medical University
No.338, Huaihai West Road, Shanghai 200052, China
E-mail: jasunyang@foxmail.com/wmke021@smmu.edu.cn

Round 3

Reviewer 3 Report

Comments and Suggestions for Authors

I cannot agree with the author's response.

Statistical analysis data with each level of the factor of interest (time and concentration of YS-49) and the interaction effect were missing.

I recommend the authors consult with a Professor of Statistics who can guide them through the Two-way ANOVA analysis.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf