Proteome Analysis of Different Resistant Apple Cultivars in Response to the Stress of Ring Rot Disease

doi:10.11983/CBB19204

Abstract

Abstract: The aim of this study was to investigate the differential expression of resistance related proteins in resistant and susceptible apple leaves in response to ring rot disease. The resistant and susceptible apple cultivars Huayue and Golden Delicious were analyzed by high-throughput isobaric tag (IBT) marker quantitative analysis combined with liquid chromatography-tandem mass spectrometry (LC-MS), proteome differential expression of resistant and susceptible apple leaves before and after pathogen treatment was analyzed, and 171 differential expression proteins (DEPs) were identified. GO enrichment and KEGG pathway analysis showed that 686 GO entries were annotated in cell components, molecular functions, and biological processes, of which 52 DEPs were annotated in 18 distinct pathways of KEGG pathway (P<0.05). Subcellular localization prediction analysis showed that 170 DEPs in 171 DEPs were located in 8 organelles. Functional annotation analysis showed that 46 DEPs were annotated in 7 classes of resistance-related proteins, including the thaumatin-like proteins, peroxidases, polyphenol oxidases, Mal d1 proteins, chitinases, endo-1,3-beta-glucosidases and MLP-like proteins. In addition, the expression characteristics and gene quantitative results of resistance-related proteins were analyzed. The results of this study provided a technical reference for further understanding the resistance mechanism of resistant and susceptible apples to the stress of ring rot disease.

Key words: apple, resistant and susceptible, IBT proteomics, ring rot disease, resistance-related proteins

Caixia Zhang,Gaopeng Yuan,Xiaolei Han,Wuxing Li,Peihua Cong. Proteome Analysis of Different Resistant Apple Cultivars in Response to the Stress of Ring Rot Disease[J]. Chinese Bulletin of Botany, 2020, 55(4): 430-441.

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URL: https://www.chinbullbotany.com/EN/10.11983/CBB19204

https://www.chinbullbotany.com/EN/Y2020/V55/I4/430

Figures/Tables 7

Table 1 Primers used in fluorescent quantitative PCR

Accession No.	Forward primer (5'-3')	Reverse primer (5'-3')
gi\|657977120	TCACCTTAGCCATCTTCTTCGC	TGCTAACTCGAACCCTGTGG
gi\|658027651	CTCCACTGTGCCTATTGCGA	GAGCCGGGTTAGCGAACAA
gi\|658061109	TTGATGCCAGCCCTGCAAAT	GGGCTTGAAGCTCGTTGTTG
gi\|661567324	GATTGCACCCCAGGCAATCA	TTGTGCTTCACGTAGCCGTA
gi\|658009573	AGGCATTCCCTCAGGACTAC	TTCCGACTTCATCCACTGC
MdActin	TGACCGAATGAGCAAGGAAATTACT	TACTCAGCTTTGGCAATCCACATC

Figure 1 Comparison of morphological characteristics of resistant and susceptible apple cultivars Huayue and Golden Delicious in response to ring rot disease R: Resistant cultivar Huayue; S: Susceptible cultivar Golden Delicious. Bars=1 cm

Figure 2 Distribution of differentially expressed proteins (DEPs) of apple leaves susceptible and resistant to ring rot disease, respectively R: Resistant cultivar Huayue; S: Susceptible cultivar Golden Delicious

Figure 3 Enriched Gene Ontology (GO) analysis of differentially expressed proteins (DEPs) in apple leaves resistant and susceptible to ring rot disease, respectively

Table 2 KEGG pathways analysis of differentially expressed proteins (DEPs)

No.	Pathway	Pathway ID	DEPs with pathway annotation (326 in total)	All proteins with pathway annotation (14916 in total)	P-value
1	Fatty acid biosynthesis	ko00061	2 (0.61%)	19 (0.13%)	0.011
2	Fatty acid elongation	ko00062	3 (0.92%)	9 (0.06%)	0.040
3	Arginine biosynthesis	ko00220	2 (0.61%)	16 (0.11%)	0.050
4	Alanine, aspartate and glutamate metabolism	ko00250	2 (0.61%)	23 (0.15%)	0.037
5	Cyanoamino acid metabolism	ko00460	2 (0.61%)	20 (0.13%)	0.022
6	Other glycan degradation	ko00511	2 (0.61%)	14 (0.09%)	0.011
7	Monoterpenoid biosynthesis	ko00902	5 (1.53%)	12 (0.08%)	0.035
8	Sesquiterpenoid and triterpenoid biosynthesis	ko00909	1 (0.31%)	2 (0.01%)	0.027
9	Nitrogen metabolism	ko00910	4 (1.23%)	28 (0.19%)	0.014
10	Sulfur metabolism	ko00920	1 (0.31%)	12 (0.08%)	0.036
11	Phenylpropanoid biosynthesis	ko00940	3 (0.92%)	47 (0.32%)	0.041
12	Isoquinoline alkaloid biosynthesis	ko00950	2 (0.61%)	16 (0.11%)	0.028
13	Biosynthesis of unsaturated fatty acids	ko01040	2 (0.61%)	9 (0.06%)	0.016
14	Fatty acid metabolism	ko01212	2 (0.61%)	32 (0.21%)	0.030
15	Ribosome	ko03010	13 (3.99%)	200 (1.34%)	0.005
16	RNA transport	ko03013	2 (0.61%)	33 (0.22%)	0.031
17	Homologous recombination	ko03440	1 (0.31%)	5 (0.03%)	0.039
18	Circadian rhythm-plant	ko04712	3 (0.92%)	9 (0.06%)	0.026

Figure 4 Subcellular localization of differentially expressed proteins (DEPs) in apple leaves resistant and susceptible to ring rot disease, respectively Chlo: Chloroplast; Cysk: Cytoskeleton; Cyto: Cytosol; Extr: Extracellular; Mito: Mitochondria; Nucl: Nucleus; Plas: Plasma membrane; Vacu: Vacuolar membrane

Figure 5 Quantitative real-time PCR analysis of genes encoding key resistance-related proteins in apple leaves resistance and susceptible to ring rot disease (A) A gene encoding a thaumatin (MD04G1018400); (B) The gene encoding Mal d1 (MD16G1160700); (C) A gene encoding a beta-1,3-glucosidase (MD11G1189000); (D) A gene encoding a polyphenol oxidase (MD10G1299100); (E) The gene encoding MLP423 (MD16G1088600). R: Resistant cultivars Huayue; S: Susceptible cultivars Golden Delicious; Different lowercase letters indicate significant differences between resistant and susceptible cultivars (P<0.05).

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