Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the potential associated infertility via regulating lipogenesis

Cherry Yin-Yi Chang; Li Yang; Joe Tse; Lun-Chien Lo; Chung-Chen Tseng; Li Sun; Ming-Tsung Lai; Ping-Ho Chen; Tritium Hwang; Chih-Mei Chen; Fuu-Jen Tsai; Jim Jinn-Chyuan Sheu

doi:10.1371/journal.pone.0271616

Peer Review History

Original SubmissionApril 13, 2022
11 May 2022 Decision Letter - Giovanni Nassa, Editor PONE-D-22-10884Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the associated infertility via regulating lipogenesisPLOS ONE Dear Dr. Sheu, 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. As you can see from comments appended below, while the reviewers find your work of interest, they have raised points that need to be addressed particularly concerning clinical samples and patient information. Please submit your revised manuscript by Jun 25 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. 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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 Reviewer #2: 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: The authors present a study about the role of genetic variations in UCA1, a lncRNA functioning as a miRNA sponge in endometriosis development. In common with a lot of other lncRNAs, UCA1 can regulate the transcription of genes via epigenetic modification. This is a topic of great interest , considering the genetic–epigenetic theory of the etiopathogenesis of endometriosis that is emerging in recent years. However, it is clear that the pathways involved in endometriosis are complicated, and the molecular mechanisms that underlie the process are largely elusive. The results reported want to add more information about miRNA sponge in endometriosis development Different concerns were raised that preclude acceptance in its current form. These are major limitations which should be addressed. TITLE 1) I suggest to evaluate to change the title in : “ Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the potential associated infertility via regulating lipogenesis” INTRODUCRION 2) The introduction should focus also on the genetic–epigenetic theory of the etiopathogenesis of endometriosis that is emerging in recent years. The authors could identify other literature on the topic and explain how the study relates to this previously published research on endometriosis. 3) Futhermore the introduction lacks some bibliographical references: - “For some patients (around 30-50%), they become infertile either due to distorting the fallopian tubes thus fail to pick up the egg after ovulation or due to constant inflammation that affect normal functions of the ovary, egg, fallopian tubes or uterus” - “ Hormone imbalance, a proposed cause for endometriosis, can be also a risk for several types of gynecological cancer” RESULTS 4) In the section about “ Functional SNPs in UCA1 and their associations with clinical features” The authors correlate the “a potent role of UCA1 in endometriosis development”. From the literature we now that there is not clear correlation with the extension of the disease and the pain score, so the results from this section that the UCA1 can have a role in endometriosis development according to the pain could be not completely correct. 5) Futherome could be interesting to analyze the correlation between the localization of endometriosis ( ovarian, peritoneal or deep endometriosis ) and the Functional SNPs in UCA1. DISCUSSION 6) The discussion should be focus more on the potential roles of UCA1 in endometriosis , because the results are not so consistent to support a certain involvement of novel haplotypes of UCA1 gene in endometriosis development and the associated clinical outcomes. Furthermore the result on association with including long-term pain and infertility are still not so consistent , but with a potential clinical relevance. METHODS Clinical samples and patient information 7) The authors should tell us how the control group were confirmed to be healthy . Minimal or mild endometriosis is difficult to detect with imaging and without a laparoscopy the presence of endometriosis can’t be confirmed 8) better specify the type of study ( prospective or retrospective study ) Reviewer #2: The authors provided a clear and well-written paper. The results and the statistical data provided are scientifically sound and the limitations of their findings were clearly assessed in the discussion section. For this reason, I cannot find major revisions. However, some minor revisions should be assessed: 1) Page 3 - "Those results confirmed the genetic vibrations in" --> variations? 2) Page 5 - "potent" is written 3 times in 3 consecutive sentences. Please, use a synonym 3) Page 5 - "Thermo-stability data, the haplotype with rs73005445 can strikingly change their full-length RNA structures (Fig. 3D and 3E)" - Is something missing? 4) Page 5 - Figure 3B is not cited in the text. 5) Page 7 - Figure 4 labels are very small. It should be better to increase a bit their size. 6) Page 10 - "Regulation of fatty acid... " --> I appreciate a good recap in biochemistry, but I don't understand the point of doing this in the discussion section. Is there some final sentence missing? What's the point of explaining all this and not connecting it with the text? I suggest adding a sentence or rephrasing the entire part. 7) Page 11 - "Blood samples were collected patients" --> "Blood samples were collected from patients" 8) Page 12 - "were further enriched by 5" --> A square appears after the 5. Some symbol is not recognized, please correct it. 9) Supplementary - I found many small typos in the pathways found: "Giloma" -> "Glioma"?, "Lysince degradation" -> "Lysine degradation"? etc... Please, check carefully the entire document and correct them all. ****** 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?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment 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. Registration is free. 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 PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0271616.r001
Revision 1
23 Jun 2022 Author Response Comments from the Reviewer #1 The authors present a study about the role of genetic variations in UCA1, a lncRNA functioning as a miRNA sponge in endometriosis development. In common with a lot of other lncRNAs, UCA1 can regulate the transcription of genes via epigenetic modification. This is a topic of great interest, considering the genetic–epigenetic theory of the etiopathogenesis of endometriosis that is emerging in recent years. However, it is clear that the pathways involved in endometriosis are complicated, and the molecular mechanisms that underlie the process are largely elusive. The results reported want to add more information about miRNA sponge in endometriosis development Different concerns were raised that preclude acceptance in its current form. These are major limitations which should be addressed. 1. I suggest to evaluate to change the title in: “Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the potential associated infertility via regulating lipogenesis” Response: Thank you for this constructive suggestion. Due to limited sample size, we agree that our data in this study may not be able to fully support our conclusion. We have changed the title according to the suggestion. 2. The introduction should focus also on the genetic–epigenetic theory of the etiopathogenesis of endometriosis that is emerging in recent years. The authors could identify other literature on the topic and explain how the study relates to this previously published research on endometriosis. Response: Thank you for the invaluable suggestion. We have updated some proposed theories of endometriosis etiology into the introduction. In the first paragraph of the introduction, we inserted ‘‘With many biochemical changes in the endometriotic lesions, the genetic/epigenetic theory was purposed in recent years to explain the hereditary aspects, the predisposition, and the endometriosis-associated changes in the endometrium, immunology, and placentation during endometriosis development’’. Two references (Refs. 4, 5) were added. The references were reordered accordingly. 3. Furthermore the introduction lacks some bibliographical references: - “For some patients (around 30-50%), they become infertile either due to distorting the fallopian tubes thus fail to pick up the egg after ovulation or due to constant inflammation that affect normal functions of the ovary, egg, fallopian tubes or uterus” - “Hormone imbalance, a proposed cause for endometriosis, can be also a risk for several types of gynecological cancer” Response: Thank you for those helpful suggestion. We have inserted bibliographical references for endometriosis associated infertility (Refs. 2 and 3) and hormone imbalance induced gynecological cancers (Refs. 2, 6, 7). 4. In the section about “Functional SNPs in UCA1 and their associations with clinical features” The authors correlate the “a potent role of UCA1 in endometriosis development”. From the literature we know that there is not clear correlation with the extension of the disease and the pain score, so the results from this section that the UCA1 can have a role in endometriosis development according to the pain could be not completely correct. Response: Thank you for the insightful comment. Due to relatively small sample size, we agree that our clinical association data may not be completely correct. We have inserted the limitation of this study in the last paragraph in the discussion in page 13. The last conclusive sentence in this section was also removed. 5. Furthermore, it could be interesting to analyze the correlation between the localization of endometriosis (ovarian, peritoneal or deep endometriosis) and the Functional SNPs in UCA1. Response: Thank you for the insightful comment. We performed association study to know the correlation of lesion adhesion site with functional SNPs in UCA1. We found that patients with risk haplotypes (G-A of rs113579010-rs73003273, G-G of rs113579010-rs73005445, and A-G of rs73003273-rs73005445) show higher frequencies to harbor the lesions at cul-de-sac, an adhesion site that frequently associated with deeply infiltrating endometriosis and the associated pain or infertility (35, 36). However, such associations did not reach statistically significance. A new figure (Fig. 2 was added), thus the figures were reordered accordingly). Two additional references (Refs. 35, 36) were added to support the possible link of cul-de-sac infiltration with pain and infertility. We do not have clear clinical data about the DIE, thus failed to perform association study of risk haplotypes with DIE. 6. The discussion should be focus more on the potential roles of UCA1 in endometriosis, because the results are not so consistent to support a certain involvement of novel haplotypes of UCA1 gene in endometriosis development and the associated clinical outcomes. Furthermore, the result on association with including long-term pain and infertility are still not so consistent, but with a potential clinical relevance. Response: Thank you for the insightful comment. We removed some biochemistry parts on lipogenesis and focused more on the epigenetic roles of UCA1 variants in endometriosis development as shown in page 12-13. As what we have learnt from the new Fig.2, patients with risk haplotypes may easily get cul-de-sac infiltration, even though the data did not reach statistically significance. Such finding may partially explain why patients with risk haplotypes suffer from long-term pain and infertility. 7. The authors should tell us how the control group were confirmed to be healthy. Minimal or mild endometriosis is difficult to detect with imaging and without a laparoscopy the presence of endometriosis can’t be confirmed Response: Thank you for the question. Yes, both patients and healthy women were confirmed by laparotomy or laparoscopy examines. We also rearranged the text in the “Clinical samples and patient information” section of M&M part in page 13-14 to make it clear. 8. better specify the type of study (prospective or retrospective study) Response: Thank you for the suggestion. It was a prospective study approved by the Institutional Review Board (IRB) at the CMUH (CMUH106-REC1-138) with informed consent from each participant. We have inserted the above information in the “Clinical samples and patient information” section of M&M part in page 13-14. Comments from the Reviewer #2 The authors provided a clear and well-written paper. The results and the statistical data provided are scientifically sound and the limitations of their findings were clearly assessed in the discussion section. For this reason, I cannot find major revisions. However, some minor revisions should be assessed: 1. Page 3 - "Those results confirmed the genetic vibrations in" --> variations? Response: Thank you for your correction, we have corrected the error in the revised version. 2. Page 5 - "potent" is written 3 times in 3 consecutive sentences. Please, use a synonym Response: Thank you for your suggestion, we have changed “those potent haplotypes” to “those novel haplotypes” in page 5. 3. Page 5 - "Thermo-stability data, the haplotype with rs73005445 can strikingly change their full-length RNA structures (Fig. 3D and 3E)" - Is something missing? Response: Thank you for your concern, we have revised the paragraph in page 6 to make it clear. 4. Page 5 - Figure 3B is not cited in the text. Response: Thank you for your concern, 3B is cited in page 6 (Fig. 3A to 3C). 5. Page 7 - Figure 4 labels are very small. It should be better to increase a bit their size. Response: Thank you for the constructive suggestion. We have increased the label size in Figure 4. We also revised the Fig. 5 and Fig. 6 accordingly to increase the label sizes 6. Page 10 - "Regulation of fatty acid... " --> I appreciate a good recap in biochemistry, but I don't understand the point of doing this in the discussion section. Is there some final sentence missing? What's the point of explaining all this and not connecting it with the text? I suggest adding a sentence or rephrasing the entire part. Response: Thank you for the insightful comment. We removed some biochemistry parts on lipogenesis and focused more on the epigenetic roles of UCA1 variants in endometriosis development as shown in page 12-13. 7. Page 11 - "Blood samples were collected patients" --> "Blood samples were collected from patients" Response: Thank you for your correction, we have corrected the error in the revised version in page 12. 8. Page 12 - "were further enriched by 5" --> A square appears after the 5. Some symbol is not recognized, please correct it. Response: Thank you for founding this error, we have fixed the “µ” sign in the revised version in page 13. 9. Supplementary - I found many small typos in the pathways found: "Giloma" -> "Glioma"?, "Lysince degradation" -> "Lysine degradation"? etc... Please, check carefully the entire document and correct them all. Response: Thank you for your corrections, we have checked the entire documents and made necessary corrections accordingly. Attachments Attachment Submitted filename: Rebutal letter UCA1 to Plos ONE.docx https://doi.org/10.1371/journal.pone.0271616.r002
5 Jul 2022 Decision Letter - Giovanni Nassa, Editor Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the potential associated infertility via regulating lipogenesis PONE-D-22-10884R1 Dear Dr. Sheu, 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. 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If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: All comments have been addressed Reviewer #2: All comments have been addressed ******** 2. 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 Reviewer #2: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: I Don't Know Reviewer #2: Yes ****** 4. 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 Reviewer #2: Yes ****** 5. 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 Reviewer #2: Yes ****** 6. 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: the authors have adequately addressed my comments raised in a previous round of review and I feel that this manuscript is now acceptable for publication Reviewer #2: The authors addressed all my comments and I have no other concerns. I hope they will try to continue on this topic with a bigger cohort. Good luck ****** 7. 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: No ******** https://doi.org/10.1371/journal.pone.0271616.r003
Formally Accepted
18 Jul 2022 Acceptance Letter - Giovanni Nassa, Editor PONE-D-22-10884R1 Genetic variations in UCA1, a lncRNA functioning as a miRNA sponge, determine endometriosis development and the potential associated infertility via regulating lipogenesis Dear Dr. Sheu: 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. Giovanni Nassa Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0271616.r004

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