Scientia Agricultura Sinica ›› 2022, Vol. 55 ›› Issue (16): 3134-3143.doi: 10.3864/j.issn.0578-1752.2022.16.006

• PLANT PROTECTION • Previous Articles     Next Articles

Identification and Functional Analysis of StSCD Family in Setosphaeria turcica

HAO YuBin1,2(),LI HaiXiao1,2,ZHANG Sai1,LIU Ning1,2,LIU YingZi3,CAO ZhiYan1,2(),DONG JinGao1,2()   

  1. 1College of Plant Protection, Hebei Agricultural University, Baoding 071001, Hebei
    2State Key Laboratory of North China Crop Improvement and Regulation/Hebei Key Laboratory of Plant Physiology and Molecular Pathology, Baoding 071001, Hebei
    3Garden Management Centre of Chengde City, Hebei Province, Chengde 067000, Hebei
  • Received:2022-03-11 Accepted:2022-04-02 Online:2022-08-16 Published:2022-08-11
  • Contact: ZhiYan CAO,JinGao DONG E-mail:839970427@qq.com;caozhiyan@hebau.edu.cn;shmdjg@hebau.edu.cn

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Abstract:

【Objective】Scytalone dehydratase (SCD) is a key enzyme in melanin synthesis. The objective of this study is to identify the StSCD gene family of Setosphaeria turcica, and to analyze the differences in the expression of StSCD gene family during the appressorium development of S. turcica and the effect of SCD inhibitors on melanin synthesis, which will lay a foundation for further study on the important role of StSCD gene family in melanin synthesis and appressorium development. 【Method】Using the whole genome data of the wild-type strain 01-23 of S. turcica, the complete sequence of the StSCD gene family was obtained, and the protein sequences were compared with the SCD of Cochlibolus heterostrophus, Pyricularia oryzae, Colletotrichum lagenaria and other fungi. Materials from different appressorium developmental stages were collected for qRT-PCR analysis to obtain the expression levels of different StSCD genes at different stages, so as to determine the important dehydratase genes closely related to pathogen infection and appressorium melanization. Carpropamid, an inhibitor of SCD, was used to treat S. turcica, and the colony growth rate, melanin production and appressorium turgor pressure were measured, so as to determine the important function of SCD in appressorium development. 【Result】Four StSCD genes were found by searching the whole genome of S. turcica, the encoded protein of them has SCD conserved domain and conserved catalytic and substrate binding amino acid residues. StSCD3 protein has high homology with the functionally redundant SCD2 protein of Botrytis cinerea, and StSCD4 has high homology with SCD protein of Corynespora cassiicola. By analyzing the expression levels of StSCD genes at different stages during the growth and development of the pathogen, it was found that the expression levels of four StSCD genes were all up-regulated at the appressorium stage, and the expression levels of StSCD3 and StSCD4 were particularly up-regulated. The expression levels of StSCD3 and StSCD4 in the regenerated hyphae of the appressorium were significantly decreased, and the expression levels of StSCD4 were higher during the whole period of appressorium induction. The melanin synthesis was blocked and the appressorium turgor pressure was significantly reduced after carpropamid treatment. 【Conclusion】There are four StSCD genes in S. turcica. It is speculated that StSCD4 is involved in the synthesis of DHN melanin, and then affects the accumulation of appressorium turgor pressure.

Key words: Setosphaeria turcica, StSCD family, melanin, carpropamid

Table 1

The StSCD primers used for qRT-PCR"

引物名称
Primer name		 序列
Sequence (5′-3′)
StSCD1DL-F GCTCTCAAAGACGCTTGCAC
StSCD1DL-R TACCAAGCCATGACCGTGTC
StSCD2DL-F CATCCAGCCCCAAGTTCCTT
StSCD2DL-R CCAGCAAATTTCCACTCGCC
StSCD3DL-F GGTGTTCAGTCTTTCACGCG
StSCD3DL-R TCCATCGCTCTCCTCTCCAA
StSCD4DL-F AGCTTCAACACGCACTGGTA
StSCD4DL-R CTAGGCCTTTTCCGGGATCC
Tubulin-F GGGAACTCCTCACGGATGTTG
Tubulin-R TAACAACTGGGCAAAGGGTCA

Table 2

Sequence characterization of StSCD gene family"

基因名称
Gene name		 基因编号
Gene number		 基因长度
Gene length (bp)		 ORF长度
ORF length (bp)		 氨基酸数量
Number of amino acid (aa)		 基因组位置
Genomic location
StSCD1 gene_12136 688 525 175 scaffold_29:99576-100263 (-)
StSCD2 gene_9587 639 486 162 scaffold_14:439746-440384 (-)
StSCD3 gene_4774 590 531 177 scaffold_6:14708-15297 (-)
StSCD4 gene_41 865 540 180 scaffold_1:164410-165274 (+)

Fig. 1

StSCD protein domain composition"

Fig. 2

Bioinformatic analysis of StSCD proteins"

Fig. 3

Relative expression level of StSCDs at different stages The values are mean±SE, different lowercases above the bars represent significant differences (P<0.05). The same as Fig. 5"

Fig. 4

Effects of different concentrations of carpropamid on melanin synthesis and colony growth of S. turcica"

Fig. 5

Melanin production of S. turcica under different concentrations of carpropamid"

Table 3

Effect of carpropamid on the appressorium turgor pressure of S. turcica"

环丙酰菌胺浓度
<BOLD>C</BOLD>arpropamid concentration (μg·mL-1)
0 5.0 10.0
膨压
Turgor pressure (MPa)		 5.60±0a 5.32±0.16b 5.06±0.16c
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