Free Amino Acids and Methylglyoxal as Players in the Radiation Hormesis Effect after Low-Dose γ-Irradiation of Barley Seeds

Round 1

Reviewer 1 Report

Radiation hormesis is a long-known problem that remains unsolved. Understanding of the mechanism of hormesis will provide valuable insights that contribute to increase in food production. The authors previously reported the morphological changes in the irradiated seven barley cultivars. The authors also previously performed HPLC analysis of metabolites in the irradiated another barley cultivar Nur. In this paper, the authors irradiated the seeds of seven different barley cultivars with gamma-rays and analyzed the correlation between the morphological changes and metabolic profiles. The research is well-organized and there is only a couple of minor issues as shown below.

• The Figure 1 and Figure 2 involve a panel of morphological changes of seven cultivars based on the ranking reported in previous paper. However, these seem to be rather confusing. I think that readers can understand the changes of the metabolites without the panels shown below the graphs.
• Some abbreviations, for example, UPR, GA, FC, CoA, SAM are not very common. It will be more reader-friendly to spell out at the first apparition in each section (i.e., Introduction, Materials and Methods, Results, and Discussion).
• I feel that the authors thought that shikimate was a top candidate, since it was shown in almost every panels. However, there is only little descriptions about shikimate in the text. The authors should explain about it even if it was negative result.
• In the Introduction, the authors described that the UPR is closely related to the hormesis. However, the cited paper, Wiegant et al 2013, focused mainly about HSR. The readers might wonder how the idea of UPR, but not HSR, came from. The authors should explain about this point.

Author Response

Dear Reviewer,

We are very grateful for your valuable comments and we improved the manuscript according to your recommendations. The point by point response is given below.

 

The Figure 1 and Figure 2 involve a panel of morphological changes of seven cultivars based on the ranking reported in previous paper. However, these seem to be rather confusing. I think that readers can understand the changes of the metabolites without the panels shown below the graphs.

We agree that the additional panels might cause confusion. We removed them and instead added the information on morphological changes to the Notes under the Figures 1 and 2.

 

Some abbreviations, for example, UPR, GA, FC, CoA, SAM are not very common. It will be more reader-friendly to spell out at the first apparition in each section (i.e., Introduction, Materials and Methods, Results, and Discussion).

Thank you, we spelled out abbreviations in each section to avoid confusion.

 

I feel that the authors thought that shikimate was a top candidate, since it was shown in almost every panels. However, there is only little descriptions about shikimate in the text. The authors should explain about it even if it was negative result.

Indeed, we considered shikimic acid as an interesting target and that might be evident from our text. We added more explanation to the beginning of Discussion section.

 

In the Introduction, the authors described that the UPR is closely related to the hormesis. However, the cited paper, Wiegant et al 2013, focused mainly about HSR. The readers might wonder how the idea of UPR, but not HSR, came from. The authors should explain about this point.

Thank you for this remark; we were totally unclear in this part. The introduction part related to UPR and HSR was improved.

Reviewer 2 Report

Please find the comments and suggestions for the manuscript entitled Potential "Free Amino Acids and Methylglyoxal as Players in the Radiation Hormesis Effect After Low-Dose γ-Irradiation of Barley Seeds".

1. Suggestion- Authors have results and discussion section separate, and then there is no need for citations in the results section. It is suggested to remove all the citations from the result section---

2. Line no- paper rolls were placed--- plants grew for seven days—please correct it. It can't be plants, it may be seedlings that grew---

3. Please also include humidity conditions for the growth of seedlings--. Also, line no-72 to 75 it is not much clear about what authors did out of 100, only 40 were used for analysis, then how many were used to get 1 gm biomass. And what happened to rest --- please elaborate about sampling and how many seedlings were used at what stage in a clear manner ---- As it's a bit confusing---

4. Table 3. – Authors were showing statistical differences by bold letters, but instead of it please use statistical letters to indicate the same and also include standard error in the table.---

5. In figure 1 author didn’t show any statistical differences between the treatments and cultivars. Is there a specific reason for the same—please explain and if required include the changes ---  similarly line no-183 authors say that in figure 2 and table 3 they found significant differences but in figures, it does not reflect at all—please change according to the claim--- 

Author Response

Dear Reviewer,

We are very grateful for your valuable comments and we improved the manuscript according to your recommendations. The point by point response is given below.

1. Suggestion- Authors have results and discussion section separate, and then there is no need for citations in the results section. It is suggested to remove all the citations from the result section

Thank you, we followed the suggestion and removed all citations from the Results section to the Discussion.

2. Line no- paper rolls were placed--- plants grew for seven days—please correct it. It can't be plants, it may be seedlings that grew

The mistake was corrected.

3. Please also include humidity conditions for the growth of seedlings. Also, line no-72 to 75 it is not much clear about what authors did out of 100, only 40 were used for analysis, then how many were used to get 1 gm biomass. And what happened to rest - please elaborate about sampling and how many seedlings were used at what stage in a clear manner. As it's a bit confusing.

Thank you for the suggestions, all required information was added to the Material and methods.

4. Table 3. – Authors were showing statistical differences by bold letters, but instead of it please use statistical letters to indicate the same and also include standard error in the table.

Thank you, we changed the bold letters to the asterisk sign. Standard error cannot be provided for these data, since they include only three replicates and are not normally distributed; therefore, non-parametrical data analysis was used. However, we provide additional information on sample in the Supplementary Table 1.

5. In figure 1 author didn’t show any statistical differences between the treatments and cultivars. Is there a specific reason for the same—please explain and if required include the changes ---  similarly line no-183 authors say that in figure 2 and table 3 they found significant differences but in figures, it does not reflect at all—please change according to the claim 

We included statistical differences in Figures, before they were only reflected in the Table.

Round 2

Reviewer 2 Report

The comments and suggestions on the manuscript entitled" Free Amino Acids and Methylglyoxal as Players in the Radiation Hormesis Effect After Low-Dose γ-Irradiation of Barley Seeds have been well addressed by the authors. The suggested changes in the figures and tables are satisfactory. The inclusions in the materials and methods are fine. I have no further suggestions and recommendations.

Thanks for the revised version.