Scientia Agricultura Sinica ›› 2023, Vol. 56 ›› Issue (24): 4854-4865.doi: 10.3864/j.issn.0578-1752.2023.24.005

• PLANT PROTECTION • Previous Articles     Next Articles

The Effect of 3-Oxyacyl ACP Reductase Gene FgOAR1 on the Growth, Development and Pathogenicity of Fusarium graminearum

GONG AnDong(), LEI YinYu, WU NanNan, LIU JingRong, SONG MengGe, ZHANG YiMei(), YANG Guang, YANG Peng   

  1. College of Life and Science, Xinyang Normal University, Xinyang 464000, Henan
  • Received:2023-07-28 Accepted:2023-09-29 Online:2023-12-21 Published:2023-12-21
  • Contact: ZHANG YiMei

RichHTML

14

PDF

159

Abstract:

【Background】Fusarium graminearum is the main pathogenic fungus which can infect wheat and result in Fusarium head blight. The infection of F. graminearum leads to huge reduction in crop quality and yield, and produces different types of mycotoxins that endanger grain production and human health. 3-oxoacyl ACP reductase (OAR1), catalyzing carboxyacyl carrier protein to oxyacyl carrier protein, which plays an important role in the fatty acid biosynthesis of F. graminearum.【Objective】To reveal the biological function of FgOAR1 in F. graminearum, the FgOAR1 deletion mutants were constructed. The phenotype, cell structure, conidia concentration, sexual reproduction and pathogenicity of mutant were analyzed and compared with wild-type strain PH-1. The results will uncover the function of FgOAR1 in the growth and pathogenic process of F. graminearum, and provide scientific evidences for the identification of novel antifungal target and control of Fusarium head blight.【Method】Wild-type strain F. graminearum PH-1 was used as material in this study. Split-Marker gene knockout technology was conducted to construct the ΔFgOAR1 mutants. The ΔFgOAR1-C reverting strain was obtained by the Gap-repair method. All strains were individually inoculated in common media (PDA, CM, YPG), and the stress selection medium containing Congo-Red, SDS, NaCl and H2O2, respectively. The mycelium phenotypes and fatty acid content were recorded. The strains were inoculated in CMC medium to analyze the conidia concentration. The carrot culture-medium was used to analyze the sexual reproduction of each strain. All strains were individually inoculated to the wheat coleoptile and spikelet to analyze the pathogenicity and mycotoxin content, respectively. The disease incidence of mutant strains was calculated and compared with PH-1 strain.【Result】Compared with the wild-type strain PH-1, the growth of ΔFgOAR1 mutant showed no difference in PDA, YEG and CM media. Whereas, under the pressure of 0.7 mol·L-1 NaCl, 0.03% H2O2, 0.01% SDS and 300 μg·mL-1 Congo-Red, the growth of ΔFgOAR1 mutant was significantly reduced compared with PH-1. The evidence demonstrated that FgOAR1 is related to the cell membrane and cell wall formation in F. graminearum. In CMC medium, the conidia concentration of ΔFgOAR1 mutant (8.1×105 conidia/mL) was less than PH-1 (1.26×106 conidia/mL) with significant difference; The pathogenicity analysis further demonstrated that the disease incidence of ΔFgOAR1 mutant was significantly lower than PH-1 in wheat coleoptile and spikelet inoculation tests.【Conclusion】FgOAR1 participates in the fatty acid biosynthesis, and plays an important role in cell membrane formation of F. graminearum. The deletion of FgOAR1 resulted in the reduced resistance to pressures, decreased conidia production and pathogenicity. Therefore, FgOAR1 is important for the growth, development and pathogenicity process of F. graminearum.

Key words: Fusarium graminearum, Fusarium head blight, gene knockout, FgOAR1, pathogenicity

Table 1

The primer sequences used in this study"

引物名称<BOLD>P</BOLD>rimer name 引物序列<BOLD>P</BOLD>rimer sequence (5′→3′)
FgOAR1_1F TCTCGATACCTCCTACCTACCT
FgOAR1_2R TTGACCTCCACTAGCTCCAGCCAAGCCTCAACACATGCTCTACACCCT
FgOAR1_3F GAATAGAGTAGATGCCGACCGCGGGTTCCAGGGTCTCAAAGCAAC
FgOAR1_4R ATCCCAGAAAATCCGCAA
HYG/F GGCTTGGCTGGAGCTAGTGGAGGTCAA
HY/R GTATTGACCGATTCCTTGCGGTCCGAA
YG/F GATGTAGGAGGGCGTGGATATGTCCT
HYG/R AACCCGCGGTCGGCATCTACTCTATTC
FgOAR1_5F ATCGGAGTTGAGGTGACAG
FgOAR1_6R GAAGAGGGGAGGAAAGGTT
FgOAR1_7F GGTCAATAGAGCAGGTCGC
FgOAR1_8R CAGGCGGTGGTAACTTCA
h850 TTCCTCCCTTTATTTCAGATTCAA
h852 ATGTTGGCGACCTCGTATTGG
h855R GTCGATGCGACGCAATCGT
h856F GCTGATCTGACCAGTTGC
FgOAR1_hbF CGACTCACTATAGGGCGAATTGGGTACTCAAATTGGTGACAGCGGCGTTAGGACA
FgOAR1_hbR CACCACCCCGGTGAACAGCTCCTCGCCCTTGCTCACGTACATCTCCAGTCCACCACT
insert_F TAACGCCAGGGTTTTCCCAGTCA
insert_R CGTGCTGCTTCATGTGGTCGG

Fig. 1

Multiple alignment of amino acid sequence of OAR1 in different fungi"

Fig. 2

PCR detection of FgOAR1 deletion mutant and complementary mutant"

Fig. 3

The colony morphology of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Table 2

The growth rate determination of three strains"

菌株
Strain		 生长速率Growth rate (cm·d-1)
PDA 5×YEG CM
PH-1 1.20±0.05 0.83±0.10 0.85±0.10
ΔFgOAR1 1.19±0.01 0.87±0.02 0.84±0.11
ΔFgOAR1-C 1.23±0.09 0.85±0.12 0.85±0.10

Table 3

The growth rate of PH-1、ΔFgOAR1 and ΔFgOAR1-C strains under different stress cultural conditions"

菌株
Strain		 生长速度Growth rate (cm·d-1)
CM Congo-Red NaCl SDS H2O2
PH-1 1.20±0.05 1.15±0.10 0.98±0.11 0.54±0.15 0.75±0.11
ΔFgOAR1 1.10±0.01 0.81±0.02* 0.62±0.17* 0.37±0.09* 0.60±0.09*
ΔFgOAR1-C 1.23±0.09 1.16±0.12 0.88±0.11 0.57±0.19 0.73±0.10

Fig. 4

The colony morphology of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains under different stress cultural conditions"

Fig. 5

Fatty acid content determination of PH-1 and ΔFgOAR1 strains"

Fig. 6

Conidia concentration determination and germination observation of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 7

Sexual reproduction of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 8

Pathogenicity of PH-1, ΔFgOAR1 and ΔFgOAR1-C strains"

Fig. 9

The expression of FgOAR1 under different growth conditions"

[1]
ZHANG J B, LI H P, DANG F J, QU B, XU Y B, ZHAO C S, LIAO Y C. Determination of the trichothecene mycotoxin chemotypes and associated geographical distribution and phylogenetic species of the Fusarium graminearum clade from China. Mycological Research, 2007, 111(8): 967-975.

doi: 10.1016/j.mycres.2007.06.008
[2]
陆维忠, 程顺和, 王裕中. 小麦赤霉病研究. 北京: 科学出版社, 2001: 2-39.
LU W Z, CHENG S H, WANG Y Z. The Study on Wheat Scab. Beijing: Science Press, 2001: 2-39. (in Chinese)
[3]
徐雍皋, 陈利锋. 小麦赤霉病防治理论研究与实践. 南京: 江苏科学技术出版社, 1993.
XU Y G, CHEN L F. Theoretical Research and Practice of Wheat Scab Control. Nanjing: Jiangsu Science and Technology Press, 1993. (in Chinese)
[4]
BAI G H, SHANER G. Management and resistance in wheat and barley to Fusarium head blight. Annual Review of Phytopathology, 2004, 42: 135-161.

pmid: 15283663
[5]
MCMULLEN M, JONES R, GALLENBERG D. Scab of wheat and barley: A re-emerging disease of devastating impact. Plant Disease, 1997, 81(12): 1340-1348.

doi: 10.1094/PDIS.1997.81.12.1340 pmid: 30861784
[6]
PESTKA J J. Deoxynivalenol: Mechanisms of action, human exposure, and toxicological relevance. Archives of Toxicology, 2010, 84(9): 663-679.

doi: 10.1007/s00204-010-0579-8 pmid: 20798930
[7]
MAGAN N, HOPE R, COLLEATE A, BAXTER E S. Relationship between growth and mycotoxin production by Fusarium species, biocides and environment. European Journal of Plant Pathology, 2002, 108(7): 685-690.

doi: 10.1023/A:1020618728175
[8]
宫安东, 韩萌真, 孔宪巍, 魏彦博, 王磊, 程琳. 茶树内生菌的应用性研究进展. 信阳师范学院学报(自然科学版), 2017, 30(1): 168-172.
GONG A D, HAN M Z, KONG X W, WEI Y B, WANG L, CHENG L. Application analysis of endophytic microbes in Camellia sinensis. Journal of Xinyang Normal University (Natural Science Edition), 2017, 30(1): 168-172. (in Chinese)
[9]
GÜNENC A N, GRAF B, STARK H, CHARI A. Fatty acid synthase: Structure, function, and regulation. Sub-Cellular Biochemistry, 2022, 99: 1-33.

doi: 10.1007/978-3-031-00793-4_1 pmid: 36151372
[10]
HE M, SU J, XU Y P, CHEN J H, CHERN M, LEI M L, QI T, WANG Z K, RYDER L S, TANG B Z, et al. Discovery of broad-spectrum fungicides that block septin-dependent infection processes of pathogenic fungi. Nature Microbiology, 2020, 5(12): 1565-1575.

doi: 10.1038/s41564-020-00790-y pmid: 32958858
[11]
SMITH S, AGRADI E, LIBERTINI L, DILEEPAN K N. Specific release of the thioesterase component of the fatty acid synthetase multienzyme complex by limited trypsinization. Proceedings of the National Academy of Sciences of the United States of America, 1976, 73(4): 1184-1188.
[12]
CROSS E M, ADAMS F G, WATERS J K, ARAGAO D, EIJKELKAMP B A, FORWOOD J K. Insights into Acinetobacter baumannii fatty acid synthesis 3-oxoacyl-ACP reductases. Scientific Reports, 2021, 11(1): 7050.

doi: 10.1038/s41598-021-86400-1
[13]
YU Y H, MA J R, GUO Q Q, MA J C, WANG H H. A novel 3-oxoacyl-ACP reductase (FabG3) is involved in the xanthomonadin biosynthesis of Xanthomonas campestris pv.campestris. Molecular Plant Pathology, 2019, 20(12): 1696-1709.

doi: 10.1111/mpp.v20.12
[14]
SCHNEIDER R, BRORS B, BÜRGER F, CAMRATH S, WEISS H. Two genes of the putative mitochondrial fatty acid synthase in the genome of Saccharomyces cerevisiae. Current Genetics, 1997, 32(6): 384-388.

doi: 10.1007/s002940050292
[15]
WANG C F, ZHANG S J, HOU R, ZHAO Z T, ZHENG Q, XU Q J, ZHENG D W, WANG G H, LIU H Q, GAO X L, MA J W, KISTLER H C, KANG Z S, XU J R. Functional analysis of the kinome of the wheat scab fungus Fusarium graminearum. PLoS Pathogens, 2011, 7(12): e1002460.

doi: 10.1371/journal.ppat.1002460
[16]
QIN J X, WU M C, ZHOU S Y. FgEaf6 regulates virulence, asexual/sexual development and conidial septation in Fusarium graminearum. Current Genetics, 2020, 66: 517-529.

doi: 10.1007/s00294-019-01043-0
[17]
ZHANG Y M, GAO X L, SUN M L, LIU H Q, XU J R. The FgSRP1 SR-protein gene is important for plant infection and pre-mRNA processing in Fusarium graminearum. Environmental Microbiology, 2017, 19(10): 4065-4079.

doi: 10.1111/emi.2017.19.issue-10
[18]
梁海东, 乔有明, 裴海昆, 段中华, 全小龙. 不同前处理分析植物脂肪酸的气相色谱-质谱法研究. 分析科学学报, 2015, 31(4): 525-528.
LIANG H D, QIAO Y M, PEI H K, DUAN Z H, QUAN X L. Comparison of different pre-processing methods to identify fatty acids in plants by gas chromatography-mass spectrometry. Journal of Analytical Science, 2015, 31(4): 525-528. (in Chinese)
[19]
GONG A D, LI H P, YUAN Q S, SONG X S, YAO W, HE W J, ZHANG J B, LIAO Y C. Antagonistic mechanism of iturin A and plipastatin A from Bacillus amyloliquefaciens S76-3 from wheat spikes against Fusarium graminearum. PLoS ONE, 2015, 10(2): e0116871.

doi: 10.1371/journal.pone.0116871
[20]
JIANG C, CAO S L, WANG Z Y, XU H J, LIANG J, LIU H Q, WANG G H, DING M Y, WANG Q H, GONG C, FENG C J, HAO C F, XU J R. An expanded subfamily of G-protein-coupled receptor genes in Fusarium graminearum required for wheat infection. Nature Microbiology, 2019, 4(9): 1582-1591.

doi: 10.1038/s41564-019-0468-8
[21]
LIU H Q, WANG Q H, HE Y, CHEN L F, HAO C F, JIANG C, LI Y, DAI Y F, KANG Z S, XU J R. Genome-wide A-to-I RNA editing in fungi independent of ADAR enzymes. Genome Research, 2016, 26(4): 499-509.

doi: 10.1101/gr.199877.115 pmid: 26934920
[22]
DEAN R, VAN KAN J A L, PRETORIUS Z A, HAMMOND- KOSACK K E, DI PIETRO A, SPANU P D, RUDD J J, DICKMAN M, KAHMANN R, ELLIS J, FOSTER G D. The top10 fungal pathogens in molecular plant pathology. Molecular Plant Pathology, 2012, 13(4): 414-430.

doi: 10.1111/mpp.2012.13.issue-4
[23]
SHARMA T, SRIDHAR P S, BLACKMAN C, FOOTE S J, ALLINGHAM J S, SUBRAMANIAM R, LOEWEN M C. Fusarium graminearum Ste3 G-protein coupled receptor: a mediator of hyphal chemotropism and pathogenesis. mSphere, 2022, 7(6): e0045622.

doi: 10.1128/msphere.00456-22
[24]
LI T, KIM D, LEE J. NADPH oxidase gene, FgNoxD, plays a critical role in development and virulence in Fusarium graminearum. Frontiers in Microbiology, 2022, 13: 822682.

doi: 10.3389/fmicb.2022.822682
[25]
CHAYAKULKEEREE M, SORRELL T C, SIAFAKAS A R, WILSON C F, PANTARAT N, GERIK K J, BOADLE R, DJORDJEVIC J T. Role and mechanism of phosphatidylinositol-specific phospholipase C in survival and virulence of Cryptococcus neoformans. Molecular Microbiology, 2008, 69(4): 809-826.

doi: 10.1111/mmi.2008.69.issue-4
[26]
ZHANG Y Z, CHEN Q, LIU C H, LIU Y B, YI P, NIU K X, WANG Y Q, WANG A Q, YU H Y, PU Z E, JIANG Q T, WEI Y M, QI P F, ZHENG Y L. Chitin synthase gene FgCHS8 affects virulence and fungal cell wall sensitivity to environmental stress in Fusarium graminearum. Fungal Biology, 2016, 120(5): 764-774.

doi: 10.1016/j.funbio.2016.02.002
[27]
DING M Y, LI J, FAN X Y, HE F, YU X Y, CHEN L, ZOU S S, LIANG Y C, YU J F. Aquaporin1 regulates development, secondary metabolism and stress responses in Fusarium graminearum. Current Genetics, 2018, 64(5): 1057-1069.

doi: 10.1007/s00294-018-0818-8
[28]
ZHU Q L, SUN L, LIAN J J, GAO X L, ZHAO L, DING M Y, LI J, LIANG Y C. The phospholipase C (FgPLC1) is involved in regulation of development, pathogenicity, and stress responses in Fusarium graminearum. Fungal Genetics and Biology, 2016, 97: 1-9.

doi: 10.1016/j.fgb.2016.10.004
[29]
BERMEJO C, RODRÍGUEZ E, GARCÍA R, RODRÍGUEZ-PENA J M, RODRÍGUEZ DE LA, CONCEPCIÓN M L, RIVAS C, ARIAS P, NOMBELA C, POSAS F, ARROYO J. The sequential activation of the yeast HOG and SLT2 pathways is required for cell survival to cell wall stress. Molecular Biology of the Cell, 2008, 19(3): 1113-1124.

doi: 10.1091/mbc.E07-08-0742 pmid: 18184748
[30]
RONCERO C, DURÁN A. Effect of Calcofluor white and Congo red on fungal cell wall morphogenesis: in vivo activation of chitin polymerization. Journal of Bacteriology, 1985, 163(3): 1180-1185.

doi: 10.1128/jb.163.3.1180-1185.1985
[31]
YANG K L, QIN Q P, LIU Y H, ZHANG L M, LIANG L L, LAN H H, CHEN C H, YOU Y C, ZHANG F, WANG S H. Adenylate cyclase AcyA regulates development, aflatoxin biosynthesis and fungal virulence in Aspergillus flavus. Frontiers in Cellular and Infection Microbiology, 2016, 6: 190.
[32]
YUAN M H, JIANG Z Y, BI G Z, NOMURA K, LIU M H, WANG Y P, CAI B Y, ZHOU J M, HE S Y, XIN X F. Pattern-recognition receptors are required for NLR-mediated plant immunity. Nature, 2021, 592(7852): 105-109.

doi: 10.1038/s41586-021-03316-6
[1] GAO XiaoXiao, TU LiQin, YANG Liu, LIU YaNan, GAO DanNa, SUN Feng, LI Shuo, ZHANG SongBai, JI YingHua. Construction of an Infectious Clone of Tobacco Mild Green Mosaic Virus Isolate Infecting Pepper from Jiangsu Based on Genomic Clone [J]. Scientia Agricultura Sinica, 2023, 56(8): 1494-1502.
[2] LI HuiXin, SONG WenPing, HAN ZongXi, LIU ShengWang. Isolation and Pathogenicity of Fowl Adenovirus Serotype 8a Strain [J]. Scientia Agricultura Sinica, 2023, 56(16): 3226-3236.
[3] MA HongXiang, WANG YongGang, GAO YuJiao, HE Yi, JIANG Peng, WU Lei, ZHANG Xu. Review and Prospect on the Breeding for the Resistance to Fusarium Head Blight in Wheat [J]. Scientia Agricultura Sinica, 2022, 55(5): 837-855.
[4] YANG ShiMan, XU ChengZhi, XU BangFeng, WU YunPu, JIA YunHui, QIAO ChuanLing, CHEN HuaLan. Amino Acid of 225 in the HA Protein Affects the Pathogenicities of H1N1 Subtype Swine Influenza Viruses [J]. Scientia Agricultura Sinica, 2022, 55(4): 816-824.
[5] HUANG JiaQuan,LI Li,WU FengNian,ZHENG Zheng,DENG XiaoLing. Proliferation of Two Types Prophage of ‘Candidatus Liberibacter asiaticus’ in Diaphorina citri and their Pathogenicity [J]. Scientia Agricultura Sinica, 2022, 55(4): 719-728.
[6] ZHANG JinLong,ZHAO ZhiBo,LIU Wei,HUANG LiLi. The Function of Key T3SS Effectors in Pseudomonas syringae pv. actinidiae [J]. Scientia Agricultura Sinica, 2022, 55(3): 503-513.
[7] LI ZhengGang,TANG YaFei,SHE XiaoMan,YU Lin,LAN GuoBing,HE ZiFu. Molecular Characteristics and Pathogenicity Analysis of Youcai Mosaic Virus Guangdong Isolate Infecting Radish [J]. Scientia Agricultura Sinica, 2022, 55(14): 2752-2761.
[8] ZHANG ChengQi,LIAO LuLu,QI YongXia,DING KeJian,CHEN Li. Functional Analysis of the Nucleoporin Gene FgNup42 in Fusarium graminearium [J]. Scientia Agricultura Sinica, 2021, 54(9): 1894-1903.
[9] CAO YuHan,LI ZiTeng,ZHANG JingYi,ZHANG JingNa,HU TongLe,WANG ShuTong,WANG YaNan,CAO KeQiang. Analysis of dsRNA Carried by Alternaria alternata f. sp. mali in China and Identification of a dsRNA Virus [J]. Scientia Agricultura Sinica, 2021, 54(22): 4787-4799.
[10] CAI Ni,YAN DuoZi,NONG XiangQun,WANG GuangJun,TU XiongBing,ZHANG ZeHua. Adhesin Gene mad2 Knockout and Functional Effects on Biological Characteristics and Inducing Plant Responses in Metarhizium anisopliae [J]. Scientia Agricultura Sinica, 2021, 54(22): 4800-4812.
[11] ZHANG Li,TANG YaFei,LI ZhengGang,YU Lin,LAN GuoBing,SHE XiaoMan,HE ZiFu. Molecular Characteristic of Squash Leaf Curl China Virus (SLCCNV) Infecting Cucurbitaceae Crops in Guangdong Province [J]. Scientia Agricultura Sinica, 2021, 54(19): 4097-4109.
[12] TAO Bu, QI YongZhi, QU Yun, CAO ZhiYan, ZHAO XuSheng, ZHEN WenChao. Construction and Verification of Fusarium Head Blight Prediction Model in Haihe Plain Based on Boosted Regression Tree [J]. Scientia Agricultura Sinica, 2021, 54(18): 3860-3870.
[13] WANG ChengLi,YIN ZhiYuan,NIE JiaJun,LIN YongHui,HUANG LiLi. Identification and Virulence Analysis of CAP Superfamily Genes in Valsa mali [J]. Scientia Agricultura Sinica, 2021, 54(16): 3440-3450.
[14] ZHAO JingYa,XIA HuiQing,PENG MengYa,FAN Zhuo,YIN Yue,XU SaiBo,ZHANG Nan,CHEN WenBo,CHEN LinLin. Identification and Functional Analysis of Transcription Factors FpAPSES in Fusarium pseudograminearum [J]. Scientia Agricultura Sinica, 2021, 54(16): 3428-3439.
[15] ZHANG ChengQi,WANG XiaoYan,CHEN Li. The Actin Binding Protein FgAbp1 is Involved in Growth, Development and Toxisome Formation in Fusarium graminearum [J]. Scientia Agricultura Sinica, 2021, 54(13): 2759-2768.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
京ICP备09089781号-30
Copyright 2018 © Editorial office of Scientia Agricultura Sinica
Tel: 86-010-82106279    E-mail: zgnykx@mail.caas.net.cn
Supported by Beijing Magtech Co.Ltd