Inborn Errors of Nucleoside Transporter (NT)-Encoding Genes (SLC28 and SLC29)
Round 1
Reviewer 1 Report
The paper is well written and to a comprehensive description of the biology of the nucleosides transporters, associates information on the clinical relevant characteristics due to mutated transporters, for clinician having to do with rare disorders.
Only a few suggestions: introductory remarks very detailed on the main disorders of nucleotides metabolism, with a look on the discoveries along the years: a figure with the timeline representation of the discovered disorders by year, would have more impact and facilitate reading.
Concluding remarks are very concise and mostly focused on the concept that nucleoside transporter proteins can play biological additional roles beyond salvage.
Authors should emphasize, particularly for clinicians reading the review, the concept of the high phenotype heterogeneity, the difficult diagnosis and the importance to suspect these disorders particularly in case of biochemical alteration highly suggestive for purine and pyrimidine metabolism alteration. Also, possible future treatment implications could be mentioned.
Author Response
We thank the reviewer for his/her suggestions.
- a) We have generated a new figure (Figure 1) which illustrates as a chronogram key discoveries within the field of rare diseases and nucleotide metabolism.
- b) The last section of the review (concluding remarks) has been modified according to the reviewer’s suggestions. The concepts of high phenotype heterogeneity, difficult diagnosis and the possibility that metabolic screening might be useful in this regard, have been emphasized. We have deleted the last paragraph of this section which referred to the roles of NT proteins beyond salvage. Regarding treatment implications, at this moment we think this is only relevant to the best known deficiency, ENT3, and therapeutic strategies are discussed in that particular section (3.2., lines 378-387).
Reviewer 2 Report
This is an important informative review article, by the group of Marcal Pastor-Anglada who have an excellent record of work on biomedically relevant aspects of nucleoside-nucleobase transport, focusing this time on rare diseases associated with mutations in human nucleoside transporters (SLC28/CNT and SLC29/ENT families). The text is very well written and structured and highlights recent evidence pointing to the gaps in knowledge implying that nucleoside transporter subtypes may have important roles beyond nucleotide homeostasis that are yet to be explored.
I have a few suggestions for improvement of the manuscript at specific points, mainly at the introductory parts (1 and 2).
(a) In the second paragraph of Introduction, starting at page 1 (line 38) and ending in line 76 at page 2, there are a multitude of abbreviated enzyme names (beginning from line 44 and running through all the remainder of the paragraph) that make the text difficult to follow. Although I understand that the full names of some of the enzymes might be awkwardly long, I would suggest that the authors write down the full name of each enzyme together with its abbreviation, as they did on other instances (for example, lines 40, 79-82). Alternatively, the authors might want to list all abbreviations of enzymes, metabolites, clinical syndromes, in one table as a Supplement.
(b) The sentence at page 3, lines 116-118 is somewhat confusing, as it stands, mainly because no mention has been made before on nucleobase transport. I would suggest to write the information more explicitly, as follows: ENT1 and ENT2 (but not CNT1, CNT2 or CNT3) have also been suggested to transport nucleobases, with ENT1 showing lesser kinetic efficiencies than ENT2 for these substrates [27]. The apparent affinities of hENT1 or hENT2 for nucleobases are lower than for nucleosides, although the transport efficiencies (Vmax/Km ratios) for nucleoside and nucleobase transport appear to be similar [27].
(c) The authors mention the recent structural evidence on hENT1 (lines 162-164, line 279) (refs 57 and 58 in the manuscript), but not the one on hCNT3, which is also recently solved, based on cryo-EM (Zhou et al. (2020), PLoS Biol. 18: e3000790). The authors should include this information, as well, either at the beginning (part 2, page 3) or at the conclusion (part 4), as a sentence pointing to the fact that the recent structural data on hENT1 and hCNT3 will facilitate better understanding of the structure-function relationships of NT proteins associated with their biological roles and the roles of particular mutants.
(d) In line 164, the reference (Wright and Lee, 2019) is [57] in the manuscript.
(e) Some references in the References list do not include the page numbers; some others do not include the corresponding doi number.
Author Response
We thank the reviewer for his/her comments and suggestions.
a) Full names for all cited enzymes have been included in the text, as suggested.
b) Former sentence on page 3, lines 116-118 (now 128-135) has been re-written in line with the reviewer’s suggestions.
c) We did not cite that particular publication because CNT3 is not covered in the review. However, the reviewer is right and for sure this is a major breakthrough in the field and this publication is now cited at the end of the manuscript (page 13, lines 489-490) (new reference 89)
d and e) The reviewer is right. We missed that particular reference in the text and was not numbered. Now this is reference 40 and, therefore, numbering of other references has been done accordingly. We have re-checked the new list of references.
Reviewer 3 Report
Anglada et al. reviewed the current understanding of inheritable traits caused by variants in genes encoding nucleoside transporter (NT) proteins. As the authors mentioned, inborn errors in NT proteins are relatively under-recognized compared with those in nucleoside synthesis pathways. The authors clearly stated that manifestations among those disorders differ significantly. This review article could enhance the understanding of NT protein disorders for those who want to collect the information.
Major issues
This paper contains much information regarding nucleoside metabolism (chapter 1), basics of NT proteins (chapter 2), genetic alterations, and (potential) functional consequences in NT disorders (chapter 3). The reviewer feels that the manuscript requires revision so that the contents are presented more organized; this manuscript looks too rambling from the beginning to the end.
Chapter 1: Could the author present a brief graphical presentation of nucleoside metabolism pathways and related disorders?
Chapter 2: Fig. 2 should be more reader-friendly. Where are the plasma cell membrane and the mitochondrion? What part of the membrane proteins do reported gene variants affect (and potential functional consequences)?
Chapter 3: Same as the comment for Fig. 2.
L311: Brief introduction about remdesivir is required so that readers outside the research fields can comprehend easily.
L312-323: The authors bring up a cross-compensation theory among ENT subclasses, which is intriguing. However, this
part was particularly hard to follow. The reviewer recommends revisions to improve logic flow.
L361-363: The authors should explain in detail why taurine-dependent attenuation of ER stress could compromise
erythropoiesis.
Minor issues
The reviewer found typographical and grammatical errors.
L208: compensate for ENT1, not compensate ENT
L44, L279, L302, L340, L470: dysfunction, not disfunction.
Author Response
We thank the reviewer for his/her suggestions.
a) The aim of this manuscript is to provide an updated overview of rare diseases associated with genetic variability in genes encoding nucleoside transporter proteins. For this reason, we felt it made sense to include a first section on inborn errors of nucleotide metabolism to help the reader understand where we are in global terms and why it is now timely to write this review on NT proteins. After this introductory section, we thought the most suitable way of organizing the manuscript was, first, to briefly introduce what nucleoside transporter proteins are and do, to later focus on each particular transporter for which rare diseases have been associated or suspected. This restricts the review only to 3 out of the 7 genes known to encode nucleoside transporter proteins. This will be, when accepted, the first review ever published on this topic.
b) Graphical presentation of nucleoside metabolism pathways and related disorders. Based upon the reasons mentioned above, we respectfully believe this is out of the scope of our contribution. Moreover, a special issue of the journal “Molecular Genetics and Metabolism” has recently covered this topic (please, see references 1 and 2 in our manuscript). Transporter proteins had not been covered at that time and this prompted us to provide this review paper.
c) Following the reviewer’s suggestion, we have modified Figure 2 to make it more reader-friendly.
d) Several paragraphs in section 3 have been changed according to the reviewer’s comments and suggestions: we introduced what remdesivir is (lines 324-326), we have also re-written the hypothesis of cross compensation between ENT subtypes (former lines 312-323, now lines 327-337) and, finally, we provide a new reference to support the concept that bile acids attenuate ER stress (reference 73) and this action can be somehow impaired in ENT3 deficient hematopoietic stem cells.
Minor issues have been addressed accordingly.
Round 2
Reviewer 3 Report
The authors have appropriately addressed the issues raised by the reviewer, and the manuscript got substantial improvement.
