内生真菌GG22蛋白质改善胁迫条件下红花幼苗的生长

doi:10.3969/j.issn.1004-1524.20240691

浙江农业学报 ›› 2025, Vol. 37 ›› Issue (6): 1193-1202.DOI: 10.3969/j.issn.1004-1524.20240691

内生真菌GG22蛋白质改善胁迫条件下红花幼苗的生长

周丹宁¹^,²(), 许姣¹, 白静³, 刘湘楠¹, 朱畇昊¹^,²^,^*()

1.河南中医药大学药学院,河南郑州 450046
2.呼吸疾病诊疗与新药研发河南省协同创新中心,河南郑州 450046
3.南阳职业学院,河南南阳 474550

收稿日期:2024-07-31 出版日期:2025-06-25 发布日期:2025-07-08
作者简介:周丹宁(2001—),男,河南新乡人,硕士,主要从事药用植物研究。E-mail:17837337981@163.com
通讯作者: *朱畇昊,E-mail:guxinhan123@163.com
基金资助:
国家自然科学基金(81603232)

Endophytic fungal GG22 protein improves growth of Carthamus tinctorius L. seedling under stress conditions

ZHOU Danning¹^,²(), XU Jiao¹, BAI Jing³, LIU Xiangnan¹, ZHU Yunhao¹^,²^,^*()

1. School of Pharmacy, Henan University of Chinese Medicine, Zhengzhou 450046, China
2. Respiratory Disease Diagnosis and Treatment and New Drug of Henan Collaborative Innovation Center, Zhengzhou 450046, China
3. Nanyang Vocational College, Nanyang 474550, Henan, China

Received:2024-07-31 Online:2025-06-25 Published:2025-07-08

摘要/Abstract

摘要：

为研究地黄内生真菌尖孢镰刀菌Fusarium oxysporum GG22的蛋白质(简称GG22蛋白质)对植物生长的作用,采用硫酸铵分级沉淀法得到不同组分的GG22蛋白质,分别命名为G20、G40、G60、G80;用G20、G40、G60、G80组的GG22蛋白质分别处理红花幼苗,初步探索3种逆境(高温、低温、盐)胁迫下GG22蛋白质对红花幼苗生长的影响。通过实时荧光定量PCR(qRT-PCR)分析胁迫条件下参与植物免疫过程的基因ARD1(激活的抗病基因1)、EDS1(疾病易感性增强基因1)、PAD4(植物抗毒素缺陷基因4)、NPR(病程相关基因非表达子基因)、CDKs(细胞周期蛋白依赖性激酶基因)的相对表达量。结果表明,不同胁迫条件下G60和G80组的GG22蛋白质对红花幼苗生长的影响较为明显。高温胁迫下G80组红花幼苗鲜重为对照组的1.41倍,G60组红花幼苗鲜重为对照组的1.39倍;低温胁迫下G80组和盐胁迫下G60组的红花幼苗芽长、鲜重、干重均较对照组显著增加。GG22蛋白质处理能够在一定程度上缓解逆境胁迫对红花幼苗根系的损伤,G60组和G80组效果较突出。在3种胁迫条件下,G60和G80处理组红花幼苗的ARD1、EDS1、PAD4、NPR、CDKs基因表达均不同程度地上调。蛋白质组鉴定得到与已知激活蛋白Aspf2、PEMG1和PeaT1相似度较高的3条蛋白质序列,可以将这3个蛋白质作为候选激活蛋白用于后续的分离、纯化与功能鉴定。综上,GG22蛋白质可以改善胁迫条件下红花幼苗的生长,GG22蛋白质中可能含有激活植物免疫反应的激活蛋白,推测其可以作为植物免疫诱抗剂。

关键词: 内生真菌, 尖孢镰刀菌, 胁迫, 激活蛋白, 红花

Abstract:

To investigate the effects of the protein from endophytic fungus Fusarium oxysporum GG22 in Rehmannia glutinosa(GG22 protein) on plant growth, different fractions of GG22 protein (designated as G20, G40, G60, and G80) were obtained using graded precipitation with ammonium sulphate. Safflower(Carthamus tinctorius L.) seedlings were treated with these GG22 protein fractions (G20, G40, G60 and G80) to preliminarily explore their impact on seedling growth under three stress conditions(high temperature, low temperature, and salinity). The relative expression levels of genes involved in plant immunity—ARD1 (activated disease resistance gene 1), EDS1 (enhanced disease susceptibility gene 1), PAD4 (plant antitoxin-deficient gene 4), NPR(non-expressors of pathogenesis related gene), and CDKs(cyclin-dependent kinase gene), under stress conditions were analyzed using quantitative real-time PCR (qRT-PCR). The results showed that the G60 and G80 groups significantly influenced safflower seedling growth under various stress conditions. Under high-temperature stress, the fresh weight of seedlings in the G80 group was 1.41 times that of the control group, while the G60 group was 1.39 times that of the control group. Under low-temperature stress (G80 group) and salt stress (G60 group), shoot length, fresh weight, and dry weight of seedlings significantly increased compared with the control. GG22 protein treatment alleviated stress-induced damage to seedling roots, with the G60 and G80 groups showing significant effects. Additionally, the expression of ARD1, EDS1, PAD4, NPR, and CDKs genes were upregulated to varying degrees in the G60 and G80 groups under all three stress conditions. Proteomic analysis identified three protein sequences with high similarity to known elicitor proteins—Aspf2, PEMG1 and PeaT1, which were selected as candidate elicitors for subsequent isolation, purification, and functional characterization. In conclusion, the protein from Fusarium oxysporum GG22 can improve safflower seedling growth under stress conditions and may contain elicitor proteins that activate plant immune responses, suggesting its potential as a plant immune elicitor.

Key words: endophytic fungi, Fusarium oxysporum, stress, activator protein, Carthamus tinctorius L.

中图分类号:

S567.21

周丹宁, 许姣, 白静, 刘湘楠, 朱畇昊. 内生真菌GG22蛋白质改善胁迫条件下红花幼苗的生长[J]. 浙江农业学报, 2025, 37(6): 1193-1202.

ZHOU Danning, XU Jiao, BAI Jing, LIU Xiangnan, ZHU Yunhao. Endophytic fungal GG22 protein improves growth of Carthamus tinctorius L. seedling under stress conditions[J]. Acta Agriculturae Zhejiangensis, 2025, 37(6): 1193-1202.

图/表 6

表1 引物信息

Table 1 Primer information

名称 Name	正向引物(5'→3') Forward primer(5'→3')	反向引物(5'→3') Reverse primer(5'→3')	扩增长度 Amplification length/bp
CtCDKs	ATGGCTGCCTCTCTTTCCTC	ATGTTGAAAGGTTGGGGCAG	249
CtNPR	TCCCGCCAAAATCTTGAAAT	ATGCACCTATGAATGCCCAC	137
CtARD1	ATCTCAGCCTTCGCCCACCT	GGTCGGATTTGTCGGATGTG	133
CtEDS1	GAAGGTGCCAGTTCCGAGAC	AGCAAGATTCCGACCTGTGT	213
CtPAD4	CGCTATGGCTTCTCTCCTATC	GAGCGAACAACAGCCGTG	182
Ct60S	GGCAACACTGGTCTTGATCCAT	CTGCGATTGTTGGATAATGGATG	92

图1 不同组分GG22蛋白质对胁迫条件下红花幼苗生长的影响 CK表示空白对照,G20、G40、G60、G80分别表示对应的蛋白质处理。*代表P<0.05,**代表P<0.01,***代表P<0.001,****代表P<0.000 1。

Fig.1 Effects of various fractions of GG22 protein on growth of safflower seedlings under stress conditions CK indicates blank control, G20, G40, G60 and G80 indicate treatments with the corresponding proteins. * represents P<0.05, ** represents P<0.01, *** represents P<0.001, **** represents P<0.000 1.

表2 不同组分GG22蛋白质对胁迫条件下红花幼苗根系活力的影响

Table 2 Effects of various fractions of GG22 protein on root activity of safflower seedlings under stress conditions μg·g-1·h-1

处理 Treatment	根系活力Root activity
处理 Treatment	正常处理Normal	高温胁迫Heat-temperature stress	低温胁迫Low-temperature stress	盐胁迫Salt stress
CK	53.83±2.348 c	41.93±1.24 b	37.31±7.32 d	47.39±1.71 b
G20	59.91±3.061 b	37.85±1.52 b	35.77±1.73 d	49.51±4.37 b
G40	54.17±2.919 c	39.94±1.35 b	48.14±1.48 c	50.57±0.64 b
G60	62.45±0.859 ab	69.87±3.27 a	59.07±0.53 b	53.63±0.36 ab
G80	66.10±0.703 a	42.42±3.64 b	82.59±1.04 a	58.32±0.85 a

图2 不同组分GG22蛋白质对胁迫条件下红花幼苗免疫相关基因表达的影响 A1、A2、A3、A4分别表示高温胁迫下使用G20、G40、G60、G80处理;B1、B2、B3、B4分别表示低温胁迫下使用G20、G40、G60、G80处理;C1、C2、C3、C4分别表示盐胁迫下使用G20、G40、G60、G80处理。

Fig.2 Effects of various fractions of GG22 protein on the expression of immunity-related genes of safflower seedlings under stress conditions A1, A2, A3, and A4 indicate G20, G40, G60 and G80 treatments under high temperature stress, respectively; B1, B2, B3 and B4 indicate G20, G40, G60 and G80 treatments under low-temperature stress, respectively; C1, C2, C3 and C4 indicate the treatment with G20, G40, G60, and G80 under salt stress, respectively.

图3 不同组分GG22蛋白质的电泳结果

Fig.3 Electrophoresis results of different GG22 protein fractions

图4 蛋白质组定性鉴定 A,表观相对分子量分布;B,不同组分蛋白质鉴定的韦恩图。

Fig.4 Qualitative identification of proteomes A, Distribution of apparent relative molecular weight; B, Venn diagram for identification of different fractions of proteins.

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内生真菌GG22蛋白质改善胁迫条件下红花幼苗的生长

Endophytic fungal GG22 protein improves growth of Carthamus tinctorius L. seedling under stress conditions

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