PONE-D-20-34026

Wheat amino acid transporters highly expressed in grain cells regulate amino acid accumulation in grain

PLOS ONE

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Reviewers' comments:

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: No

Reviewer #2: Yes

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Reviewer #1: This MS reveals the important role of three amino acid transporters (TaAAP2, TaAAP13, TaAAP21) in wheat grains for amino acid transport, which deserves to be published. Several suggestions are as follows:

1 There are obvious deficiencies in the introduction of this MS. many important AAP genes in Arabidopsis and rice should be introduced, such as AtAAP2 and AtAAP8 in Arabidopsis, OsAAP3, OsAAP5 and OsAAP1 in rice. These genes have an important contribution to the grain yield of Arabidopsis or rice.

2 What is the sub-location of these three amino acid transporters in the plant cell? subcellular localization deserves consideration for supplementation.

3 why are there no overexpression lines and RNAi lines of the other two genes (TaAAP2 and TaAAP21)?

4 The sentences in the MS should be checked and organized, and there are many minor problems.

Reviewer #2: The authors use RNA-seq, RT-qPCR, and promoter-GUS assays to investigate the expression pattern of three amino acid transporters in different wheat tissues. They provide evidence to support that all three transporters can transport a broad spectrum of amino acids. Finally, manipulating the expression of amino acid transporters alters nitrogen accumulation in the wheat grains. The manuscript will be of interest to readers with foci on amino acid transporters as well as those exploring regulation of nitrogen accumulation in the crop grains.

Major comments:

1. The expressions of TaAAP2, TaAAP13, and TaAAP21 in different organs were shown in fig 1d-f. These genes all have three homologous genes, however, the pictures and article only generally show that TaAAP2, TaAAP13, and TaAAP21 are analyzed. Please note that the expression of all their homologous genes or one homologous gene was analyzed.

2. Lines 299-302, in order to determine the expression pattern of TaAAP2, TaAAP13, and TaAAP21, the promoters of TaAAP2b, TaAAP13d and TaAAP21a were selected for GUS staining experiment in Fig. 2. TaAAP2, TaAAP13, and TaAAP21 all have three homologous genes, but why did authors choose the promoter of TaAAP2b, TaAAP13d and TaAAP21a genes? Their high expression or other reasons should be explained.

3. Several yeast mutants were used to examine the transport activities of TaAAP2, TaAAP13, and TaAAP21. The genotypes of these mutants should be indicated or the references for the origin of the mutants should be provided.

4. In this paper, the transport activities of TaAAP2, TaAAP13, and TaAAP21 for several amino acids was examined. Why is histidine not detected?

5. Lines 443-444, which of the TaAAP13 homologous genes is overexpressed should be specified.

Mirror points

1. Line385, "whole grain four" is wrongly written, it should be "whole grain flour".

2. Line407, " flours " should not be plural.

3. Some gene names are not italicized in the paper, such as line260, 283. Please correct it.

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Reviewer #1: No

Reviewer #2: No

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Wheat amino acid transporters highly expressed in grain cells regulate amino acid accumulation in grain

PONE-D-20-34026R1

Dear Dr. Hawkesford,

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Kind regards,

Jin-Song Zhang, Ph.D.

Academic Editor

PLOS ONE

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