The Tomato Interspecific NB-LRR Gene Arsenal and Its Impact on Breeding Strategies
Round 1
Reviewer 1 Report
- Does the number of genes differ in wild tomatoes growing in climatic zones corresponding to their natural distribution and in places of growth?
- The cytoplasmic receptors NB LRRs for certain pathogens are known to interact with specific effector proteins. Activation of NB LRRs in most cases leads to the development of programmed cell death in a local area of the plant, which is believed to physically isolate the infection and prevent further spread of the pathogen.What genes in tomatoes are responsible for PCD during stress, besides NRC1?
- A large number of NB LRRs are encoded in the plant genome, providing specific protection against patogens. Thus, in Arabidopsis genome, 149 NB LRRs predicted by the structure are encoded, and in the rice and poplar genomes - about 400. How many NB LRRs are encoded in the wild tomato genome?
- It is known that when attacked by pathogens, ROS are generated, what happens at the gene level?
- Attention should be paid to the review by Vakhrusheva O.A. and articles by Lee H.A., Takken F.L., Rogozina E.V.
Comments for author File: 
Comments.doc
Author Response
We appreciated the time the reviewer has spent in reading and revising this manuscript and we are grateful to the reviewer for their valuable comments. We believe that their suggestions helped to improve the overall quality of our manuscript. All changes in the revised version are in red.
Please, find our responses to each of your suggestions/comments below.
Reviewer #1
Does the number of genes differ in wild tomatoes growing in climatic zones corresponding to their natural distribution and in places of growth?
Reply: The availability of wild tomato genome annotations is very limited. To date, the completed genome sequence of S. pennellii, S. pimpinellifolium and S. chilense is usable (Bolge et al. 2014; Wang et al. 2020; Stam et al. 2020). The vast majority of released genomic data for wild tomatoes are from re-sequencing and transcriptomic data (solgenomics.net). Therefore, due to limited annotated genomes, it is difficult to estimate the variability in the number of genes in wild tomatoes growing in climatic zones and in places of growth.
The cytoplasmic receptors NB LRRs for certain pathogens are known to interact with specific effector proteins. Activation of NB LRRs in most cases leads to the development of programmed cell death in a local area of the plant, which is believed to physically isolate the infection and prevent further spread of the pathogen. What genes in tomatoes are responsible for PCD during stress, besides NRC1?
Reply: The programmed cell death (PCD) plays a key role in various plant processes such as development and immunity (Hofius et al., 2007; Bozhkov and Lam, 2011; Coll et al., 2011). The hypersensitive response (HR) is a rapid and localized PCD reaction at the site of attempted pathogen invasion. A localized HR has been reported after activation of defence responses by extracellular and intracellular (NB-LRRs) receptors (Shen and Schulze-Lefert et al., 2007 and Naito et al., 2008). Genetic and morphological differences of PCD are emerged in response to activated TIR-NB-LRR and CC-NB-LRR proteins (Munch et al. 2015), but in both cases a localized HR has been reported (Hatsugai et al., 2004; Naito et al., 2008; Hatsugai et al., 2009). We added the sentence “Activation of NB-LRRs, during ETI response, induce a programmed cell death, as known as hypersensitive response (HR)” in the introduction section to precise that the NB-LRR genes are responsible for PCD during stress.
A large number of NB LRRs are encoded in the plant genome, providing specific protection against pathogens. Thus, in Arabidopsis genome, 149 NB LRRs predicted by the structure are encoded, and in the rice and poplar genomes - about 400. How many NB LRRs are encoded in the wild tomato genome?
Reply: We have indicated the amount of identified NB-LRRs in wild tomato species according to R-gene annotations released by Seong et al. (2020).
It is known that when attacked by pathogens, ROS are generated, what happens at the gene level?
Reply: Production of reactive oxygen species (ROS) is critical for successful activation of immune responses against pathogen infection (Lee et al. 2020). To provide resistance against pathogens, the plant innate immunity relies on NB-LRR genes and plasma membrane localized pattern recognition receptors (PRRs). PRRs recognize the pathogen-associated molecular patterns (PAMPs), resulting in pattern-triggered immunity (PTI) (Chisholm et al. 2006; Jones and Dangl 2011).
At line 58 we wrote “Plants own numerous non-self recognition receptors able to identify enemy molecules and induce a set of pathways and signaling cascades to repel attacks”. Usually, ROS occur very early in the defense response and are often important for the initiation of a hypersensitive response (HR) (Delledonne et al., 2001). In particular, the PRR complex leads to activation of downstream signaling including a rapid burst of ROS, activation of mitogen-activated protein kinase cascades, calcium influx, lipid peroxidation, regulation of calcium-dependent protein kinases, transcriptional reprogramming and phytohormone regulation (Chinchilla et al. 2007). We see the point raised by the reviewer. However, our review focuses on genome-wide arrangement of tomato NB-LRR genes, presence in wild species and evolution in the Solanaceae, and not on signaling cascade of plant immunity. We cannot discuss this issue here.
Attention should be paid to the review by Vakhrusheva O.A. and articles by Lee H.A., Takken F.L., Rogozina E.V.
Reply: We thank the reviewer for their valuable suggestions. We have now cited the suggested works.
Reviewer 2 Report
This manuscript reviews NB-LRR genes of tomato and related Solanaceae species regarding their genome-wide arrangement, presence in wild species, evolution in the Solanaceae. Furthermore, how such accumulated extensive information and diverse resources can be potentially used for efficiently generating new tomato varies with novel resistance. The content of manuscript covers extensive and updated information regarding all the above topics and thus would be helpful for the Solanaceae scientific communities as well as private sectors. Some additional question, comments and suggestions: ※ Line 35: Ref. 2 & 3 mainly focus on fruit and metabolism of tomato, and thus are not appropriate to cover the provided information. ※ Line 37: Ref. 4 is a report specifically describing a fungal disease of tomato, and thus is not appropriate to cover the provided information. ※ The first paragraph in page 3: Is it true that NB-LRR genes of tomato are absent in chromosomes 7, 8, 9, 12? If so, what could be the possible reasons for it? ※ Line 140-144: The sentences need to be checked and revised. ※ Fig. 1: It is difficult to get the information provided in this figure, particularly the “B” circle. And, is the number of genes in each group positively correlated with the length of the circle of each group? In the figure legend, the wording “B) ….the number of the numbers” is unclear. The total number of predicted NB-LRR genes in each of the subclasses should be provided. ※ Line 153-154 & Fig. 1: The wording “three large expansions of regions” is not clear. Besides, the Prf/R1 and Gpa2/Bs2/Rx/Rx2 subfamilies only have 14 and 6 genes, respectively. How could these fit the description? How about the subfamily which has 24 uncharacterized genes? ※ Line 166: The format of this subtitle should be revised. ※ Line 230: The reference citation should be revised. ※ Line 284-288: Ref. 74 & 75 are cited wrongly. ※ The format of references is inconsistent and the information in some references is incomplete. All the details should be carefully checked and revised.
Comments for author File: Comments.docx
Author Response
We are grateful to the reviewer for their valuable comments. We believe that their suggestions helped to improve the overall quality of our manuscript. All changes in the revised version are in red.
Please, find our responses to each of your suggestions/comments below.
Reviewer #2
This manuscript reviews NB-LRR genes of tomato and related Solanaceae species regarding their genome-wide arrangement, presence in wild species, evolution in the Solanaceae. Furthermore, how such accumulated extensive information and diverse resources can be potentially used for efficiently generating new tomato varies with novel resistance. The content of manuscript covers extensive and updated information regarding all the above topics and thus would be helpful for the Solanaceae scientific communities as well as private sectors.
We acknowledge the reviewer for their useful comments and suggestions, that, in our opinion, helped to improve the overall quality of our manuscript. All changes in the revised version are highlighted in red.
We have gone through the concerns and tried to clarify murky points. Please, find our responses to each of your suggestions/comments below.
Some additional question, comments and suggestions:
※ Line 35: Ref. 2 & 3 mainly focus on fruit and metabolism of tomato, and thus are not appropriate to cover the provided information.
Reply: We thank the reviewer for pointing out this inconsistency. We have replaced the references with a more appropriate one.
※ Line 37: Ref. 4 is a report specifically describing a fungal disease of tomato, and thus is not appropriate to cover the provided information.
Reply: We thank the reviewer for pointing out this inconsistency. The reference was removed.
※ The first paragraph in page 3: Is it true that NB-LRR genes of tomato are absent in chromosomes 7, 8, 9, 12? If so, what could be the possible reasons for it?
Reply: We see the point raised by the reviewer, and we corrected the text accordingly.
※ Line 140-144: The sentences need to be checked and revised.
Reply: We have rephrased the sentence and we hope it is clearer now.
.
※ Fig. 1: It is difficult to get the information provided in this figure, particularly the “B” circle. And, is the number of genes in each group positively correlated with the length of the circle of each group? In the figure legend, the wording “B) ….the number of the numbers” is unclear. The total number of predicted NB-LRR genes in each of the subclasses should be provided.
Reply: The correct wording was “the number of members”. However, to avoid confusion we replaced “the number of members” with “the amount of members”. Furthermore, we revised Fig.1 and its legend. We confirm that, the amount of NB-LRRs in each group is positively correlated with the size of the annular segment of each group.
※ Line 153-154 & Fig. 1: The wording “three large expansions of regions” is not clear. Besides, the Prf/R1 and Gpa2/Bs2/Rx/Rx2 subfamilies only have 14 and 6 genes, respectively. How could these fit the description? How about the subfamily which has 24 uncharacterized genes?
Reply: We have rephrased the sentence.
※ Line 166: The format of this subtitle should be revised.
Reply: Apologies for the formatting error. The subtitle is now properly formatted.
※ Line 230: The reference citation should be revised.
Reply: We correct it.
※ Line 284-288: Ref. 74 & 75 are cited wrongly.
Reply: We correct it.
※ The format of references is inconsistent and the information in some references is incomplete. All the details should be carefully checked and revised.
Reply: We went through all the references and we have standardized the formatting and corrected all errors.
