Genome-Wide Analysis and Expression Profiling of YUCCA Gene Family in Developmental and Environmental Stress Conditions in Tea Plant (Camellia sinensis)

Round 1

Reviewer 1 Report

The manuscript is devoted to the structural, phylogenetic and functional analysis of the YUCCA Gene Family in tea plant. The authors performed detailed research of the genes sequences and their role in plant development and stress response to various factors.

The article is quite similar to the previous author's study published in Plants 2022, 11, 2243:

Zhang, L.; Li, M.; Fu, J.; Huang, X.; Yan, P.; Ge, S.; Li, Z.; Bai, P.; Zhang, L.; Han, W.; Li, X. Genome‐wide identification and expression analysis of isopentenyl transferase family genes during development and resistance to abiotic stresses in tea plant (Camellia sinensis)

and the authors mentioned it in the references.

Although the study is complex and contains many data it has some minor disadvantages. 

p.3 (2.2. Multiple Sequence Alignment, Phylogenetic Analysis): maximum likelihood (ML) method is mention to construct the dendrogram, at the same time on the page 5 Figure 1  NJ method (neighbor-joining) is specified. What method was used?

Figures do not follow their mention in the text and sometimes they are put not in the order: for example, fig. 2A, fig. 3A, fig. 3B and then fig. 2B.

The manuscript contains too many figures (10) that should be reduced.

The font size and the sharpness should be increased (figures 2 and 3). 

Explanation of the blue and red circles and numbers should be added at least in the first figure (fig. 4, 5, 6, 9, 10).

Results are sometimes very descriptive and can be shorten / summarized to the one or two sentences, especially if there are figures (sections of chapter 3.5).

At least some numbers in differences of gene expression should be put in the text when the authors described the qRT-PCR results: the data on fig. 7 and 8 are hard to read and they are many.

p. 16,17 (3.5.3.3. The Roles of CsYUC Genes in Response to Drought Stress): there is some inconsistency - the authors defined that drought stress inhibited "the transcription of CsYUC11.8, -6, -7, and -2.3", "PEG treatment significantly inhibited the transcription of CsYUC1, -2.1, -2.3". But then they "speculated that the transcript of eleven CsYUC genes, including CsYUC2.2, -2.3, -6, -7, -8, -11.3, -11.4, -11.6, -11.7, -11.8, and -11.9, may involve in the positive regulation of drought stress resistance of the tea plant". The role of at least CsYUC 2.3 shoud be clarified.

There are also some small errors in the text: extra dots (p. 6 "3.2..", p.7 "3.3. . Gene Structure", p.9 "3.5. 1.. Expression" etc),  p.15: "wwere analyszed".

 

 

Author Response

The manuscript is devoted to the structural, phylogenetic and functional analysis of the YUCCA Gene Family in tea plant. The authors performed detailed research of the genes sequences and their role in plant development and stress response to various factors.

The article is quite similar to the previous author's study published in Plants 2022, 11, 2243:

Zhang, L.; Li, M.; Fu, J.; Huang, X.; Yan, P.; Ge, S.; Li, Z.; Bai, P.; Zhang, L.; Han, W.; Li, X. Genome‐wide identification and expression analysis of isopentenyl transferase family genes during development and resistance to abiotic stresses in tea plant (Camellia sinensis)

and the authors mentioned it in the references.

Although the study is complex and contains many data it has some minor disadvantages. 

1) p.3 (2.2. Multiple Sequence Alignment, Phylogenetic Analysis): maximum likelihood (ML) method is mention to construct the dendrogram, at the same time on the page 5 Figure 1  NJ method (neighbor-joining) is specified. What method was used?

Sorry. The ML method was used in this study. On the page 19, in the Figure legend of Figure 1, ‘N J method’ has been changed to ‘M L method’.

 

2) Figures do not follow their mention in the text and sometimes they are put not in the order: for example, fig. 2A, fig. 3A, fig. 3B and then fig. 2B.

Thank you. Done.

 

3) The manuscript contains too many figures (10) that should be reduced. The font size and the sharpness should be increased (figures 2 and 3). 

Thank you. Done.

 

4) Explanation of the blue and red circles and numbers should be added at least in the first figure (fig. 4, 5, 6, 9, 10).

Thank you. The explanation has been added in the first figure-Fig.4.

 

5) Results are sometimes very descriptive and can be shorten / summarized to the one or two sentences, especially if there are figures (sections of chapter 3.5).

Thank you. Done.

 

6) At least some numbers in differences of gene expression should be put in the text when the authors described the qRT-PCR results: the data on fig. 7 and 8 are hard to read and they are many.

Sorry. In fig. 6 and 7 of the revised manuscript, some percentages in differences of gene expression have been supplemented.

 

7) p. 16,17 (3.5.3.3. The Roles of CsYUC Genes in Response to Drought Stress): there is some inconsistency - the authors defined that drought stress inhibited "the transcription of CsYUC11.8, -6, -7, and -2.3", "PEG treatment significantly inhibited the transcription of CsYUC1, -2.1, -2.3". But then they "speculated that the transcript of eleven CsYUC genes, including CsYUC2.2, -2.3, -6, -7, -8, -11.3, -11.4, -11.6, -11.7, -11.8, and -11.9, may involve in the positive regulation of drought stress resistance of the tea plant". The role of at least CsYUC 2.3 shoud be clarified.

Sorry. The expression levels of CsYUC2.2 and -2.3 both were decreased by drought or PEG treatment. So the conclusion in the text is wrong and is changed to ‘it can be speculated that the transcript of nine CsYUC genes, including CsYUC6, -7, -8, -11.3, -11.4, -11.6, -11.7, -11.8, and -11.9, may involve in the positive regulation of drought stress resistance of the tea plant’. Accordingly, in the section ‘4. Conclusions’, lines 557-558, the sentence has been changed into ‘There are 9 and8 CsYUC genes which ……’.

 

8) There are also some small errors in the text: extra dots (p. 6 "3.2..", p.7 "3.3. . Gene Structure", p.9 "3.5. 1.. Expression" etc),  p.15: "wwere analyszed".

 Thank you. They have been revised.

 

Author Response File: Author Response.docx

Reviewer 2 Report

Manuscript: Genome-Wide Analysis and Expression Profiling of YUCCA Gene Family in Developmental and Environmental Stress Conditions in Tea Plant (Camellia sinensis)

The manuscript explains important research on an important gene family in tea.

The scientific merit of the manuscript is high.

However the followings need to be improved.

1. Writing in certain sections are too long and these sections can be summarized to read better.

For example,  1.      Under 2.6.2 ‘ the lengthy discussion can be summarized in a table and reduce the text.

 

 

2. Overall language editing is required, There are some spelling mistakes which need to be corrected. Basically some long discussions can be summarized but without removing the facts.

 

 

1. Writing in certain sections are too long and these sections can be summarized to read better.

For example,  1.      Under 2.6.2 ‘ the lengthy discussion can be summarized in a table and reduce the text.

 

 

 

 

2. Overall language editing is required, There are some spelling mistakes which need to be corrected. Basically some long discussions can be summarized but without removing the facts.

 

Author Response

Author reply in attachment

Author Response File: Author Response.docx

Reviewer 3 Report

In this work authors present the search for  CsYUC Genes in the tea plant. the search is done by a simple Blast matching, but no significance parameters are given, therefore it is impossible to estimate the quality and significance of blast matches. 

The major drawback of the work is the absence of functional tests of the identified genes. The test must show functional relevance of the identified genes to the YUC genes. Overall the work represents pure predictions and the attracted third party data on expression etc in no way supports either blast predictions or prove that predictions were made correctly. 

Within the paper there are many contradictions either to itself or to known facts, some are listed below:

Anaerobic induction in plant leaves this is a real nonsense  fig 2.

Expression levels in leaves at the lowest of all organs which is nonsense assuming the functionality of YUC (fig 4)

Xia  et  al.  [28]  reported  that  AtYUC2  promoter  activity  was observed in mature leaves of Arabidopsis. Consistent with this report,  CsYUC-2.1, -2.3 and -11.4 were expressed in mature tea leaves - Your data exactly contradicts that !!!

No significance values are presented in figures 4 and 5 

Short-Read Archive (SRA) - this shows the complete ignorance of authors in the gene expression topic

17 unique CsYUC members with confirmed conserved domains - how did you confirm this? It is not in the paper 

Fig 1: Phylogenetic tree is a complete nonsense - all branches are around a hundred that means the distances are all almost the same, the tree makes no sense in this case. Try to make bootstrapping analysis and you will see a different tree each time 

“The results showed that the expression profiles of CsYUCs varied  widely  in  five  tissues.” - This is exactly the opposite of what is expected from a group of similar genes -  they must show a correlated pattern of expression or correlated behavior. This is a fundamental property of a group which differs it from a random set. As in your case a random set of genes will have expression varied across tissues etc. This is because your prediction of CsYUCs genes is a full nonsense.

Data on Figure 8 shows there is no coordinated pattern in expression and it majorly contradicts your previous study [22].

 

 

English:

for deeply studying

Author Response

Author reply in attachment

Author Response File: Author Response.docx

Reviewer 4 Report

The tea tree is one of the popular cultural plants. Therefore, all research aimed at improving the quality of tea leaves and increasing productivity are priority areas. The presented study is a continuation of a series of studies and is devoted to the study of genes of the YUCCA family associated with auxin biosynthesis. The results obtained are of undoubted interest.

There are some minor comments, mainly related to the design.

Section 2.6.2. - provide only a description of the methodology. Move the entire explanation to the Results and Discussion section.

2.7.-Remove the underlining, indicate a space between 250 mM.

Move the description of treatment with stress factors to a separate subsection.

Table 1 - instead of molecular weight, indicate molecular mass

GRAVY - provide a reference for the method or describe it in the Materials and Methods section.

Figure 2 - it is advisable to make the picture brighter, like Figure 3.

Reference 11 - remove underlining

References 35,36 - use a dot, not a dash

Author Response

Author reply in attachment

Author Response File: Author Response.docx

Round 2

Reviewer 3 Report

In the revised version and the reply letter authors did not respond adequately to a single question. Authors must reply with good objectives and proofs on their view in case they do not agree. But in no way with “I don 't completely agree with you.” Other replies they only repeat what was already in the text. 

Ignoring the reviewer's comments is not a right strategy to publish the manuscript. 

 

Right in the first point the concerns about the Blast search significance and the search parameters were ignored, taking that this is a central point of the entire work. 

Point 2: Restrictions on time is not an excuse not to do the work. You must do the experiments 

3: This is exactly what should not be done - you are hiding results that you do not like - this is a falsification of the work! 

 

Point 9 – very critical - ignored.

 

Author Response

Dear Reviewer, 

Right in the first point the concerns about the Blast search significance and the search parameters were ignored, taking that this is a central point of the entire work. 

Sorry. In this study, the protein sequences of tea CsYUC family numbers were extracted using Blast Wrapper including in TBtools software based on Arabidopisis AtCsYUC protein sequences. The E-value was set to 1e-5. The sentences ‘The E-value was set to 1e-5’ was added in line 157 in the revised version.

Point 2: Restrictions on time is not an excuse not to do the work. You must do the experiments 

Sorry. When the plant materials of tea plants can be used, we will do the experiment of functional tests immediately.

Best wishes!     
Sincerely yours,  
Liping Zhang

Author Response File: Author Response.docx