Identification of Pepper CaSYT1 and Its Function in Phytophthora capsici Infection

doi:10.16420/j.issn.0513-353x.2023-0113

Acta Horticulturae Sinica ›› 2024, Vol. 51 ›› Issue (3): 533-544.doi: 10.16420/j.issn.0513-353x.2023-0113

• Genetic & Breeding·Germplasm Resources·Molecular Biology • Previous Articles Next Articles

Identification of Pepper CaSYT1 and Its Function in Phytophthora capsici Infection

LIU Yanyan¹, DING Ying¹, LIU Xinghua¹, ZHENG Jiaqiu¹, LIU Zhiqin²^,^*()

¹ Jiangsu Coastal Area Institute of Agricultural Sciences，Yancheng，Jiangsu 224000，China
² College of Agriculture，Fujian Agriculture and Forestry University，Fuzhou 350002，China

Received:2023-08-30 Revised:2023-12-08 Online:2024-03-25 Published:2024-03-22
Contact: LIU Zhiqin

Abstract

Abstract:

In order to clarify whether the gene CaSYT1 encoding Synaptotagmin（SYT）protein in pepper may play an important role in pepper’s response to Phytophthora capsici infection. The expression pattern and function of CaSYT1 were preliminarily studied by protein conserved structure prediction，subcellular localization，P. capsici and exogenous hormone treatments，transient expression and virus-induced gene silencing. The results showed that CaSYT1 contained one transmembrane helix domain and two C2 domains，which were located to the plasma membrane. The transcript levels of CaSYT1 were induced by P. capsici infection and exogenous hormone SA，and the CaSYT1 promoter activities were activated and trigger the expression of downstream genes under P. capsici infection and exogenous SA treatment. The transient expression of CaSYT1 can induce the accumulation of H₂O₂ in pepper leaves and trigger obvious hypersensitive response（HR）-like cell death. CaSYT1 silencing enhanced pepper resistance to P. capsici infection，accompanied by up- regulated expression of resistance-related genes，including CaPR1，NPR1，and CaDEF1，suggesting CaSYT1plays a negative regulatory role in pepper response to P. capsici infection. CaSYT1 transient expression can trigger H₂O₂ accumulation and cell death，indicating that CaSYT1 plays a negative regulatory role in pepper response to P. capsici infection in a manner independent of cell death and H₂O₂ accumulation.

Key words: Capsicum annuum, CaSYT1, Phytophthora capsici, gene identification, signal passage, expression analysis

LIU Yanyan, DING Ying, LIU Xinghua, ZHENG Jiaqiu, LIU Zhiqin. Identification of Pepper CaSYT1 and Its Function in Phytophthora capsici Infection[J]. Acta Horticulturae Sinica, 2024, 51(3): 533-544.

Figures/Tables 9

Table 1 Primer sequences used in this study

用途 Usage	基因 Gene	正向序列（5′-3′） Forward sequence	反向序列（5′-3′） Reverse sequence
实时荧光定量 qRT-PCR	CaSYT1	ATGGGTTTTGTGAGTACGAT	AATTGGTGCCGCAATATCTT
	CaPR1（Rezaul et al.，2019）	CAGGATGCAACACTCTGGTGG	ATCAAAGGCCGGTTGGTC
	CaHIR1（Choi et al.，2012）	GACATGGTCCTGGTAACCCA	CCCAACAGAAGCCTGAAGAA
	CaNPR1（Dong，2004）	ACTTCTTCGCCGACGCCAAG	GCCAACACATTCACCAGAGCATC
	CaDEF1（Do et al.，2004）	GAGGAAGAAGTTTGAAAG	GAAATACACCACATGAAG
亚细胞定位载体构建Construction of subcellular localization vectors	CaSYT1	AAAAAGCAGGCTACATGGGTTTTGTGAGTACGAT	AGAAAGCTGGGTCGCTGCAATGGCGAACTGCATCT
瞬时表达载体构建Construction of transient overexpression vectors	CaSYT1	AAAAAGCAGGCTACATGGGTTTTGTGAGTACGAT	AGAAAGCTGGGTCTCAAGATGCAGTTCGCCATT
沉默载体构建 Construction of silent vectors	CaSYT1	AAAAAGCAGGCTACATGGGTTTTGTGAGTACGAT	AGAAAGCTGGGTCAATTGGTGCCGCAATATCTT

Fig. 1 Phylogenetic analysisof CaSYT1 and SYT1 from other plants The number represents credibility.

Fig. 2 Subcellular localization（A）of CaSYT1 and Western Blot（B）

Fig. 3 The transcriptional expression levels of CaSYT1 in pepper plant leaves inoculated with Phytophthora capsica and treated with different exogenous phytohormones * and ** indicate that there are significant difference at the levels of P < 0.05 and P < 0.01. The same below.

Fig. 4 CaSYT1 promoter cis-acting element

Fig. 5 GUS enzyme activity driven by CaSYT1 after inoculation with Phytophthora capsici and treatment with different exogenous plant hormones

Fig. 6 The effect of transient overexpression of CaSYT1 on hypersensitive response mimicked cell death（A）and conductivity（B）

Fig. 7 Effect of transient expression of CaSYT1 on expression of defense-related genes

Fig. 8 CaSYT1-silencing enhances pepper’s resistance to Phytophthora capsici inoculation A：The phenotype of CaPDS-silenced pepper plant three weeks post agro-infiltration. B：The relative expression of CaSYT1 was detected by real-time RT-PCR 24h after inoculation with Phytophthora capsici. C：Phenotype of pepper seedlings after inoculation with Phytophthora capsici. D：Diseased spots on isolated leaves of pepper after inoculation of Phytophthora capsici. E：Real-time RT-PCR detection of transcriptional levels of related resistance genes in pepper leaves.

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Identification of Pepper CaSYT1 and Its Function in Phytophthora capsici Infection

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