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Acta Agronomica Sinica ›› 2023, Vol. 49 ›› Issue (1): 105-118.doi: 10.3724/SP.J.1006.2023.24004

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Transcriptional expression profiling of soybean genes under sulfur-starved conditions by RNA-seq

WANG Hui(), WU Zhi-Yi, ZHANG Yu-E, YU De-Yue()   

  1. National Center for Soybean Improvement / National Key Laboratory of Crop Genetics and Germplasm Enhancement / Key Laboratory of Biology and Genetic Improvement of Soybean, Ministry of Agriculture and Rural Affairs / Nanjing Agricultural University, Nanjing 210095, Jiangsu, China
  • Received:2022-01-04 Accepted:2022-03-25 Online:2023-01-12 Published:2022-04-20
  • Contact: YU De-Yue E-mail:wanghui0@njau.edu.cn;dyyu@njau.edu.cn
  • Supported by:
    National Natural Science Foundation of China(32072080)

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Abstract:

Soybean is an important grain and oil crop. Few researches focused on soybean sulfur utilization. Here, the tolerance of Yunmengliuyuehuaye (YM) and Qinyangdadou (QY) to low sulfur were evaluated. The gene expression profiles of roots and leaves of the two materials under the control (+S) and sulfur deficiency (-S) environments were analyzed via RNA resequencing. The results showed that YM was tolerance to low sulfur and QY was susceptibility to low sulfur. 9064 and 9795 differentially expressed genes (DEGs) were identified in leaves of YM and QY, and 3185 and 5006 DEGs were identified in roots of YM and QY, respectively. KEGG enrichment revealed that nine pathways were common in the two material leaves, of which MAPK signaling pathway—plant specially enriched more upregulated expressed genes. There were 18 common pathways in both roots. Nine of them responded consistently to low sulfur in YM and QY, of which four contained more up-regulated genes and five contained more downregulated genes. In the remaining nine pathways, YM contained more upregulated expressed genes. Soybean sulfate transporter genes were important for the absorption and transportation of sulfate. In the transcriptome, 27 soybean sulfate transporter genes were identified. These genes belonged to 4 subgroups respectively. Most of the genes in subgroups 1, 2, and 4 were induced by low sulfur, and the genes in subgroup 3 responded to low sulfur in a complex way. KEGG showed that the upregulated gene of GmEIL1 (ethylene-insensitive 3-like) in the MAPK signaling pathway-plant was clone. The gene was involved in the regulation of soybean sulfur utilization in the soybean chimeras with transgenic hairy root. These results provide a basis for deeply exploring the genetic mechanism of soybean sulfur utilization efficiency and candidate genes for soybean tolerance to low sulfate breeding in the future.

Key words: soybean, low sulfur, RNA-seq, sulfate transporter, EIN3/EIL

Table 1

Primers used in the study"

基因名称
Gene name		 上游引物
Forward primer (5°-3°)		 下游引物
Reverse primer (5°-3°)		 用途
Application
GmEIL1 TTCAGCAGCCTTCTTTCCCAA AGCAAAACCATGGCGAGTTTC 定量PCR
qRT-PCR
GmTubulin GGAGTTCACAGAGGCAGAG CACTTACGCATCACATAGC 内参基因
Reference gene
EFR1 (Glyma.13G123100) TCCAAAGCCACTAACAAAAA CTCAGAACCCAAGTCCTCCA 定量PCR
qRT-PCR
GmEIL1 ACCACGGTGACCCATTTCTGTATCT TGATAATGATGATGAGGTTTCCAGC 基因克隆
Gene cloning
OE-GmEIL1 CAGGTCGACTCTAGAGGATCCGCCACCATGATGATGATGCTTGAAGA GGGAAATTCGAGCTCGGTACCTCACTGATACCAAATAGAGA 构建过表达载体
Construction of
overexpression vectors
RI-GmEIL1 GGGGACAAGTTTGTACAAAAAAGCAGGCTCCCTTGAAGATATGGGATTTTG GGGGACCACTTTGTACAAGAAAGCTGGGTGAACCGAACTTTATCCTTCCA 构建干扰载体
Construction of RNAi vectors

Fig. 1

Phenotypic evaluation of YM and QY to low sulfate +S and -S represent the treatments of control and low sulfate, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou. *, **, and *** mean significant difference at the 0.05, 0.01, and 0.001 probability levels, respectively."

Fig. S1

qRT-PCR and RNA-seq results of 21 DEGs log2(FC): log2-transformed fold change value of DEGs between the control and low-sulfur treatments. L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Fig. 2

Number of differentially expressed genes (DEGs) in leaves and roots of YM and QY L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Fig. 3

KEGG pathways A: L-YM; B: R-YM; C: L-QY; D: R-QY. L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Fig. 4

Proportion of upregulated and downregulated DEGs in the common KEGG pathways The number of DEGs enriched in the pathway is list in bracket. L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Fig. 5

Relative expression profiles of DEGs encoding sulfate transmembrane transporters (SULTR) The number shows the log2-transformed fold change value of DEGs between the control and low-sulfur treatments. The gray blank represents no significant. L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Fig. 6

Phylogenetic tree based on the putative protein sequences of EILs (A) and the relative expression levels of soybean EILs (B) The number shows the log2-transformed fold change value of DEGs between the control and low-sulfur treatments. The gray blank represents no significant. L and R represent leaves and roots, respectively. YM: Yunmengliuyuehuaye; QY: Qinyangdadou."

Table S1

Sulfate transporter genes with differential expression between two soybean materials"

基因名
Gene name		 R-YM/R-QY L-YM/L-QY
对照 Control 缺硫 Low sulfate 对照 Control 缺硫 Low sulfate
GmSULTR1;1a — — 2.93 —
GmSULTR2;1b -2.50 — 1.57 —
GmSULTR2;1c -2.15 — 2.66 —
GmSULTR2;2c -1.38 — — —
GmSULTR3;1a — -1.45 1.38 —
GmSULTR3;2a -1.48 -1.97 1.86 —
GmSULTR3;3b -3.09 -2.25 — —
GmSULTR3;4b — 2.63 4.80 —
GmSULTR3;4c — — — 1.41
GmSULTR3;5b 6.17 — 6.92 —
GmSULTR4;1 -1.05 — — —
GmSULTR4;2 — — 1.31 —

Fig. 7

Relative expression levels of GmNIL1 genes in different tissues"

Fig. 8

Relative expression levels of GmEIL1 (A), the sulfate content of shoots (B), and hairy roots (C), and the relative expression level of ERF1 (Glyma.13G123100) (D) of transgenic soybean chimera +S: control; -S: low sulfate. OE-EV, OE-EIL1, RI-EV, and RI-EIL1 stand for transgenic soybean chimera containing gene expression empty vector of gene expression, GmEIL1 gene overexpression vector, empty vector of RNAi, and GmEIL1 gene RNAi vector, respectively. *, **, and *** mean significant difference at the 0.05, 0.01, and 0.001 probability levels, respectively."

[1] Li Q, Gao Y, Yang A. Sulfur homeostasis in plants. Int J Mol Sci, 2020, 21: 8926.
doi: 10.3390/ijms21238926
[2] Ding Y, Zhou X, Zuo L, Wang H, Yu D. Identification and functional characterization of the sulfate transporter gene GmSULTR1;2b in soybean. BMC Genomics, 2016, 17: 373.
doi: 10.1186/s12864-016-2705-3
[3] Phartiyal P, Kim W S, Cahoon R E, Jez J M, Krishnan H B. Soybean ATP sulfurylase, a homodimeric enzyme involved in sulfur assimilation, is abundantly expressed in roots and induced by cold treatment. Arch Biochem Biophys, 2006, 450: 20-29.
pmid: 16684499
[4] Phartiyal P, Kim W S, Cahoon R E, Jez J M, Krishnan H B. The role of 5’-adenylylsulfate reductase in the sulfur assimilation pathway of soybean: molecular cloning, kinetic characterization, and gene expression. Phytochemistry, 2008, 69: 356-364.
pmid: 17761201
[5] Krishnana H B, Jez J M. Review: the promise and limits for enhancing sulfur-containing amino acid content of soybean seed. Plant Sci, 2018, 272: 14-21.
doi: S0168-9452(18)30016-5 pmid: 29807584
[6] Chronis D, Krishnan H B. Sulfur assimilation in soybean (Glycine max [L.] Merr.): molecular cloning and characterization of a cytosolic isoform of serine acetyltransferase. Planta, 2004, 218: 417-426.
doi: 10.1007/s00425-003-1123-3
[7] Zhang C, Meng Q, Zhang M, Huang F, Gai J, Yu D. Characterization of O-acetylserine (thiol) lyase-encoding genes reveals their distinct but cooperative expression in cysteine synthesis of soybean [Glycine max (L.) Merr.]. Plant Mol Biol Rep, 2008, 26: 277-291.
doi: 10.1007/s11105-008-0047-2
[8] Chronis D, Krishnan H B. Sulfur assimilation in soybean: molecular cloning and characterization of O-acetylserine (thiol) lyase (cysteine synthase). Crop Sci, 2003, 43: 1819-1827.
doi: 10.2135/cropsci2003.1819
[9] 陈燕宁, 吴志医, 元文杰, 阚贵珍, 黄方, 喻德跃, 王慧. 核糖体基因GmRPL12对大豆低硫耐性的调控作用研究. 大豆科学, 2020, 39: 518-526.
Chen Y N, Wu Z Y, Yuan W J, Kan G Z, Huang F, Yu D Y, Wang H. Research on the regulation effect of ribosomal gene GmRPL12 on low sulfur tolerance in soybean. Soybean Sci, 2020, 39: 518-526. (in Chinese with English abstract)
[10] Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. Methods, 2001, 25: 402-408.
doi: 10.1006/meth.2001.1262 pmid: 11846609
[11] Pertea M, Pertea G M, Antonescu C M, Chang T C, Mendell J T, Salzberg S L. StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat Biotechnol, 2015, 33: 290-295.
doi: 10.1038/nbt.3122 pmid: 25690850
[12] Xie C, Mao X, Huang J, Ding Y, Wu J M, Dong S, Kong L, Gao G, Li C Y, Wei L P. KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res, 2011, 39: W316-W322.
doi: 10.1093/nar/gkr483
[13] Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol, 2013, 30: 2725-2729.
doi: 10.1093/molbev/mst197 pmid: 24132122
[14] Holsters M, de Waele D, Depicker A, Messens E, van Montagu M, Schell J. Transfection and transformation of Agrobacterium tumefaciens. Mol Gen Genet, 1978, 163: 181-187.
doi: 10.1007/BF00267408
[15] Kereszt A, Li D X, Indrasumunar A, Nguyen C D T, Nontachaiyapoom S, Kinkema M, Gresshoff P M. Agrobacterium rhizogenes-mediated transformation of soybean to study root biology. Nat Protoc, 2007, 2: 948-952.
pmid: 17446894
[16] Hirai M Y, Fujiwara T, Awazuhara M, Kimura T, Noji M, Saito K. Global expression profiling of sulfur-starved Arabidopsis by DNA macroarray reveals the role of O-acetyl-L-serine as a general regulator of gene expression in response to sulfur nutrition. Plant J, 2003, 33: 651-663.
doi: 10.1046/j.1365-313X.2003.01658.x
[17] Luo A, Zhan H, Zhang X, Du H, Zhang Y, Peng X. Cytoplasmic ribosomal protein L14B is essential for fertilization in Arabidopsis. Plant Sci, 2020, 292: 110394.
doi: 10.1016/j.plantsci.2019.110394
[18] Lin D, Jiang Q, Zheng K, Chen S, Zhou H, Gong X, Xu J, Teng S, Dong Y. Mutation of the rice ASL2 gene encoding plastid ribosomal protein L21 causes chloroplast developmental defects and seedling death. Plant Biol, 2015, 17: 599-607.
doi: 10.1111/plb.12271
[19] Ludwig A, Tenhaken R. Suppression of the ribosomal L2 gene reveals a novel mechanism for stress adaptation in soybean. Planta, 2001, 212: 792-798.
pmid: 11346953
[20] Kim K Y, Park S W, Chung Y S, Chung C H, Kim J I, Lee J H. Molecular cloning of low-temperature-inducible ribosomal proteins from soybean. J Exp Bot, 2004, 55: 1153-1155.
doi: 10.1093/jxb/erh125
[21] Dong X, Duan S, Wang H B, Jin H L. Plastid ribosomal protein LPE2 is involved in photosynthesis and the response to C/N balance in Arabidopsis thaliana. J Integr Plant Biol, 2020, 62: 1418-1432.
doi: 10.1111/jipb.12907
[22] Gururani M A, Upadhyaya C P, Strasser R J, Yu J W, Park S W. Evaluation of abiotic stress tolerance in transgenic potato plants with reduced expression of PSII manganese stabilizing protein. Plant Sci, 2013, 198: 7-16.
doi: 10.1016/j.plantsci.2012.09.014 pmid: 23199682
[23] Kang L, Kim H S, Kwon Y S, Ke Q, Ji C Y, Park S, Lee H, Deng X, Kwak S. IbOr regulates photosynthesis under heat stress by stabilizing IbPsbP in sweet potato. Front Plant Sci, 2017, 8: 989.
doi: 10.3389/fpls.2017.00989 pmid: 28642783
[24] Yabuta S, Ifuku K, Takabayashi A, Ishihara S, Ido K, Ishikawa N, Endo T, Sato F.Three PsbQ-like proteins are required for the function of the chloroplast NAD(P)H dehydrogenase complex in Arabidopsis. Plant Cell Physiol, 2018, 51: 866-876.
doi: 10.1093/pcp/pcq060
[25] Dietzen C, Koprivova A, Whitcomb S J, Langen G, Jobe T O, Hoefgen R, Kopriva S. The transcription factor EIL1 participates in the regulation of sulfur-deficiency response. Plant Physiol, 2020, 84: 2120-2136.
[26] Maruyama-Nakashita A, Nakamura Y, Tohge T, Saito K, Takahashi H. Arabidopsis SLIM1 is a central transcriptional regulator of plant sulfur response and metabolism. Plant Cell, 2006, 18: 3235-3251.
doi: 10.1105/tpc.106.046458
[27] Wawrzyńska A, Lewandowska M, Sirko A. Nicotiana tabacum EIL2 directly regulates expression of at least one tobacco gene induced by sulphur starvation. J Exp Bot, 2010, 61: 889-900.
doi: 10.1093/jxb/erp356 pmid: 20018902
[28] Zuber H, Davidian J C, Aubert G, Aimé D, Belghazi M, Lugan R, Heintz D, Wirtz M, Hell R, Thompson R, Gallardo K. The seed composition of Arabidopsis mutants for the group 3 sulfate transporters indicates a role in sulfate translocation within developing seeds. Plant Physiol, 2010, 154: 913-926.
doi: 10.1104/pp.110.162123
[29] Cao M J, Wang Z, Wirtz M, Hell R, Oliver D J, Xiang C B. The chloroplast-localized sulfate transporter SULTR3;1 affects ABA biosynthesis in Arabidopsis thaliana. Plant J, 2013, 73: 607-616.
doi: 10.1111/tpj.12059
[30] Kataoka T, Hayashi N, Yamaya T, Takahashi H. Root-to-shoot transport of sulfate in Arabidopsis: evidence for the role of SULTR3;5 as a component of low affinity sulfate transport system in the root vasculature. Plant Physiol, 2004, 136: 4198-4204.
doi: 10.1104/pp.104.045625
[31] Zhao H, Frank T, Tan Y, Zhou C, Jabnoune M, Arpat A B, Cui H, Huang J, He Z, Poirier Y, Engel K H, Shu Q. Disruption of OsSULTR3;3 reduces phytate and phosphorus concentrations and alters the metabolite profile in rice grains. New Phytol, 2016, 211: 926-939.
doi: 10.1111/nph.13969
[32] Yamaji N, Takemoto Y, Miyaji T, Mitani-Ueno N, Yoshida K T, Ma J F. Reducing phosphorus accumulation in rice grains with an impaired transporter in the node. Nature, 2017, 541: 92-95.
doi: 10.1038/nature20610
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