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Differential regulation of defense-related proteins in soybean during compatible and incompatible interactions between Phytophthora sojae and soybean by comparative proteomic analysis

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Abstract

Key message

Few proteomic studies have focused on the plant- Phytophthora interactions, our study provides important information regarding the use of proteomic methods for investigation of the basic mechanisms of plant- Phytophthora interactions.

Abstract

Phytophthora sojae is a fast-spreading and devastating pathogen that is responsible for root and stem rot in soybean crops worldwide. To better understand the response of soybean seedlings to the stress of infection by virulent and avirulent pathogens at the proteomic level, proteins extracted from the hypocotyls of soybean reference cultivar Williams 82 infected by P. sojae P6497 (race 2) and P7076 (race 19), respectively, were analyzed by two-dimensional gel electrophoresis. 95 protein spots were differently expressed, with 83 being successfully identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and subjected to further analysis. Based on the majority of the 83 defense-responsive proteins, and defense-related pathway genes supplemented by a quantitative reverse transcription PCR assay, a defense-related network for soybean infected by virulent and avirulent pathogens was proposed. We found reactive oxygen species (ROS) burst, the expression levels of salicylic acid (SA) signal pathway and biosynthesis of isoflavones were significantly up-regulated in the resistant soybean. Our results imply that following the P. sojae infection, ROS and SA signal pathway in soybean play the major roles in defense against P. sojae. This research will facilitate further investigation of the molecular regulatory mechanism of the defense response in soybean following infection by P. sojae.

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Abbreviations

ACC:

1-Aminocyclopropane-1-carboxylic acid

ACLY:

ATP citrate (pro-S)-lyase

ACO:

1-Aminocyclopropane-1-carboxylate oxidase

ACS:

1-Aminocyclopropane-1-carboxylate synthase

AOC:

Allene oxide cyclase

AOS:

Allene oxide synthase

APX2:

Ascorbate peroxidase 2

AsA:

Ascorbate

Avr:

Avirulence

CHI:

Chalcone isomerase

CHR:

Chalcone reductase

CHS:

Chalcone synthase

citF:

Citrate lyase subunit alpha/citrate CoA-transferase

COI1:

Coronatine insensitive 1

CTR1:

Constitutive triple response 1

DAB:

3,3′-diaminobenzidine

DFR:

Bifunctional dihydroflavonol 4-reductase/flavanone 4-reductase

EDS1:

Enhanced disease susceptibility 1

EDS5:

Enhanced disease susceptibility 5

EIL:

EIN3-like

ERF:

Ethylene response factor

EIN2:

Ethylene insensitive 2

EIN3:

Ethylene insensitive 3

EREBP:

Ethylene response element binding protein

ET:

Ethylene

ETI:

Effector-triggered immunity

ETR1:

Ethylene resistant 1

F3H:

Flavanone 3-hydroxylase

FLS:

Flavonol synthase

GR:

Glutathione reductase

GRP:

Glycine-rich protein

GSH:

Glutathione

GSSG:

Glutathione disulfide

GST24:

Glutathione S-transferase 24

HID:

2-Hydroxyisoflavanone dehydratase

Hpa :

Hyaloperonospora arabidopsidis

HR:

Hypersensitive response

I2′H:

Isoflavone 2′-hydroxylase

IFR:

Isoflavone reductase

IFS:

Isoflavone synthase

IOMT:

Isoflavone 7-O-methyltransferase

JA:

Jasmonic acid

JAZ:

Jasmonate ZIM domain-containing protein

MAMPs or PAMPs:

Microbial- or pathogen-associated molecular patterns

MnSOD:

Manganese superoxide dismutase

NAC:

Nascent polypeptide-associated complex

NPR1:

Nonexpresser of PR genes 1

OGDH:

2-Oxoglutarate dehydrogenase

OPDA:

Oxophytodienoic acid

OPR3:

12-Oxophytodienoate reductase 3-like

PAD4:

Phytoalexin deficient 4

PAL:

Phenylalanine ammonia lyase

PAPs:

Purple acid phosphatases

PCD:

Programmed cell death

PDF1.2:

Plant defensin gene

PRRs:

Pattern-recognition receptors

PTI:

PAMP-triggered immunity

rab-GDI:

Rab GDP dissociation inhibitor

ROS:

Reactive oxygen species

Rps genes:

Resistance to P. sojae

SA:

Salicylic acid

SAM:

S-adenosyl-l-methionine

SAMe:

S-adenosyl-l-methionine

SAR:

Systemic acquired resistance

SCPs:

Serine carboxypeptidases

SID2:

SA induction-deficient 2

UFD:

Ubiquitin fusion degradation

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Acknowledgments

This work was supported in part by grants from China National Funds for Distinguished Young Scientists (31225022) and Special Fund for Agro-Scientific Research in the Public Interest (201303018) of China to Yuanchao Wang.

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The authors declare that they have no conflict of interest.

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    Authors and Affiliations

    1. Department of Plant Pathology, Nanjing Agricultural University, Nanjing, 210095, China

      Maofeng Jing, Hongyu Ma, Haiyang Li, Baodian Guo, Xin Zhang, Wenwu Ye, Haonan Wang, Qiuxia Wang & Yuanchao Wang

    2. Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, 210095, China

      Maofeng Jing, Hongyu Ma, Haiyang Li, Baodian Guo, Xin Zhang, Wenwu Ye, Haonan Wang, Qiuxia Wang & Yuanchao Wang

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    1. Maofeng Jing
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    Correspondence to Yuanchao Wang.

    Additional information

    Communicated by H. S. Judelson.

    M. Jing and H. Ma contributed equally to this work.

    Electronic supplementary material

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    299_2015_1786_MOESM1_ESM.tif

    Fig. S1 The phenotypes of the infection processes during incompatible and compatible interactions between P. sojae and soybean. (TIFF 1791 kb)

    299_2015_1786_MOESM2_ESM.tif

    Fig. S2 Representative protein spot maps of seedling hypocotyls of soybean cultivar Williams 82 which infected by Phytophthora sojae P6497 and P7076. 2-DE was performed using 1200 µg soluble protein, nonlinear 24 cm IPG strips (pH 4-7) and 12% SDS-PAGE gels for second dimension electrophoresis. Gels were stained with CBB G-250. (TIFF 692 kb)

    299_2015_1786_MOESM3_ESM.tif

    Fig. S3 Identified proteins involved in the metabolic pathway network. SOD, superoxide dismutase; GSSG, glutathione disulfide; GSH, glutathione; ACS, 1-aminocyclopropane-1-carboxylate synthase; OGDH, 2-oxoglutarate dehydrogenase; ACLY, ATP citrate (pro-S)-lyase; and citF, citrate lyase subunit alpha / citrate CoA-transferase. Red italics indicate the protein spots identified in this study. (TIFF 422 kb)

    299_2015_1786_MOESM4_ESM.tif

    Fig. S4 Histochemical identification of H2O2 by DAB staining in soybean hypocotyls. A, C: soybean hypocotyls at 12 and 24 h after inoculation with zoospores were stained by DAB; the incompatible interaction; B, D: soybean hypocotyls at 12 and 24 h after inoculation with zoospores were stained by DAB; the compatible interaction. Bar, 50 μm. (TIFF 5929 kb)

    299_2015_1786_MOESM5_ESM.tif

    Fig. S5 The protein–protein interaction in seedling hypocotyls from the soybean cultivar Williams 82 infected by P. sojae P6497 and P7076. Analysis of a predicted protein–protein interaction network using STRING 9.1 (http://string-db.org). Arabidopsis thaliana and a confidence level of 0.4 were used as analysis parameters. Different-colored lines represent the types of evidence used to predict the associations: gene fusion (red), neighborhood (green), co-occurrence across genomes (blue), co-expression (black), experimental (purple), association in curated databases (light blue) or co-mentioned in PubMed abstracts (yellow). Five clusters of highly interacting protein nodes are marked with circles and include the proteins involved in amino acid and nitrogen metabolism, signal transduction, redox homeostasis, carbohydrate metabolism, and secondary metabolism. (TIFF 2125 kb)

    299_2015_1786_MOESM6_ESM.docx

    Table S1 The descriptions of the 83 differentially expressed proteins identified by MS/MS in the seedling hypocotyls from soybean cultivar Williams 82. (DOCX 47 kb)

    Table S2 Primers used in this work. (DOCX 18 kb)

    299_2015_1786_MOESM8_ESM.rar

    Supplementary material 2 All 2-DE maps of seedling hypocotyls of soybean cultivar Williams 82, which infected by Phytophthora sojae P6497 and P7076. (RAR 17807 kb)

    Supplementary material 3 The HTML Document of Mascot search results. (RAR 623 kb)

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    Jing, M., Ma, H., Li, H. et al. Differential regulation of defense-related proteins in soybean during compatible and incompatible interactions between Phytophthora sojae and soybean by comparative proteomic analysis. Plant Cell Rep 34, 1263–1280 (2015). https://doi.org/10.1007/s00299-015-1786-9

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