Expression Pattern and Regulation Mechanism of ApSK3 Dehydrin （Agapanthus praecox）Response to Abiotic Stress and Hormone Signals

doi:10.16420/j.issn.0513-353x.2020-0660

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (8): 1565-1578.doi: 10.16420/j.issn.0513-353x.2020-0660

• Research Papers • Previous Articles Next Articles

Expression Pattern and Regulation Mechanism of ApSK₃ Dehydrin （Agapanthus praecox）Response to Abiotic Stress and Hormone Signals

YANG Tianchen, CHEN Xiaotong, LÜ Ke, ZHANG Di()

School of Design,Shanghai Jiao Tong University,Shanghai 200240,China

Received:2021-02-09 Revised:2021-06-03 Online:2021-08-25 Published:2021-09-06
Contact: ZHANG Di E-mail:zhangdi2013@sjtu.edu.cn

Abstract

Abstract:

This study cloned the dehydrin ApSK₃（Agapanthus praecox）2 195 bp promoter sequence of ApSK₃-P using real-time PCR technology to analyze tissue specificity of ApSK₃ gene,and gene expression pattern response to various abiotic stress and ABA signal. Expression vectors with five promoter deletion fragments of ApSK₃-P were constructed and transformed into Arabidopsis thaliana,in order to reveal the response pattern of ApSK₃ gene to different stress and hormone signals and the regulatory function of cis-acting elements. Bio-informatic analysis showed that the ApSK₃-P contained many typical cis-acting elements related to plant stress,hormone response and plant development. ApSK₃ gene was tissue-specific expression,the highest expression was found in fruits,followed by leaves and roots,and the lowest value was shown in flowers. ApSK₃ is more sensitive to ABA and salt signals,followed by drought and low temperature,and no obvious response to high temperature. ApSK₃-P::GUS were transformed into A. thaliana and it showed that ApSK₃-P presented strong activity in seedlings of A. thaliana,GUS activity of roots was stronger than that of leaves,and the activity gradually increased in the developing fruits. Different promoter deletion fragments of ApSK₃-P test results showed that-2 175 to-950 bp fragment played an important role in the regulation of promoter activity,multiple cis-acting elements responsed to drought stress,two tandem ABRE elements participated in response to ABA signals,and ApSK₃-P can be activated by ERE（-526 to-533 bp）and P-box（-561 to-567 bp）under ethylene and gibberellin signal,respectively. These results indicated that ApSK₃-P can be activated by drought,cold,salt,ABA,GA and ethylene,and ABRE,ERE and P-box elements of ApSK₃-P play important roles in response to abiotic stress and hormone signals.

Key words: Agapanthus praecox, dehydrin, promoter, abiotic stress, cis-acting element

CLC Number:

S682.1

YANG Tianchen, CHEN Xiaotong, LÜ Ke, ZHANG Di. Expression Pattern and Regulation Mechanism of ApSK₃ Dehydrin （Agapanthus praecox）Response to Abiotic Stress and Hormone Signals[J]. Acta Horticulturae Sinica, 2021, 48(8): 1565-1578.

Figures/Tables 9

References 40

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引物名称	序列（5′-3′）	退火温度/℃
Primer name	Primer sequence	T_m
Ap-Actin-S	CAGTGTCTGGATTGGAGG	50.0
Ap-Actin-A	TAGAAGCACTTCCTGTG	50.0
RT-ApSK₃-S	AAGAGCCAAGAGGAGGTT	55.0
RT-ApSK₃-A	CTTCTTCTCGCCGTCTTC	55.0
ApSK₃-SP1	TGTGGCCGGGGAGCTTCTGTTT	61.4
ApSK₃-SP2	ACGTCCTGTTCGGTTACCACGG	61.4
ApSK₃-SP3	CCACTTCCTCTTCGTCGCTCGA	61.4
ApSK₃-SP2-1	AGCTAGAGAAACGAATTATCACATCCCTC	59.6
ApSK₃-SP2-2	ACTTTCAAGCTCTCGACTCCGG	59.5
ApSK₃-SP2-3	CCCTCTCTCACTCACCTCCACA	61.4
Sp-2175-S	ATGACCATGATTACGCCAAGCTTGTGTTGCTATTTGTTAGAGAA	56.0
Sp-1167-S	ATGACCATGATTACGCCAAGCTTAAAGTAAAAAGAGCCAACACTTG	55.0
Sp-950-S	ATGACCATGATTACGCCAAGCTTCCACATCCGTCCATTAGTGC	57.0
Sp-646-S	ATGACCATGATTACGCCAAGCTTGGCAACATAATCATTAAGATACTGT	57.0
Sp-291-S	ATGACCATGATTACGCCAAGCTTGCTGATGGATAAGTAAAGAATAA	55.0
Sp-A	ACTGACCACCCGGGGATCCTTTTTTAATTAATTATAAACTTCAATG	59.0
pBI121-S	CGGCTCGTATGTTGTGTGGAATTG	60.0
pBI121-A	CGTTGGGGTTTCTACAGGACGTAA	58.0
Kana-S	TGGATTGCACGCAGGTTCTC	58.0
Kana-A	CTCGATGCGATGTTTCGCTT	58.0

引物名称	序列（5′-3′）	退火温度/℃
Primer name	Primer sequence	T_m
Ap-Actin-S	CAGTGTCTGGATTGGAGG	50.0
Ap-Actin-A	TAGAAGCACTTCCTGTG	50.0
RT-ApSK₃-S	AAGAGCCAAGAGGAGGTT	55.0
RT-ApSK₃-A	CTTCTTCTCGCCGTCTTC	55.0
ApSK₃-SP1	TGTGGCCGGGGAGCTTCTGTTT	61.4
ApSK₃-SP2	ACGTCCTGTTCGGTTACCACGG	61.4
ApSK₃-SP3	CCACTTCCTCTTCGTCGCTCGA	61.4
ApSK₃-SP2-1	AGCTAGAGAAACGAATTATCACATCCCTC	59.6
ApSK₃-SP2-2	ACTTTCAAGCTCTCGACTCCGG	59.5
ApSK₃-SP2-3	CCCTCTCTCACTCACCTCCACA	61.4
Sp-2175-S	ATGACCATGATTACGCCAAGCTTGTGTTGCTATTTGTTAGAGAA	56.0
Sp-1167-S	ATGACCATGATTACGCCAAGCTTAAAGTAAAAAGAGCCAACACTTG	55.0
Sp-950-S	ATGACCATGATTACGCCAAGCTTCCACATCCGTCCATTAGTGC	57.0
Sp-646-S	ATGACCATGATTACGCCAAGCTTGGCAACATAATCATTAAGATACTGT	57.0
Sp-291-S	ATGACCATGATTACGCCAAGCTTGCTGATGGATAAGTAAAGAATAA	55.0
Sp-A	ACTGACCACCCGGGGATCCTTTTTTAATTAATTATAAACTTCAATG	59.0
pBI121-S	CGGCTCGTATGTTGTGTGGAATTG	60.0
pBI121-A	CGTTGGGGTTTCTACAGGACGTAA	58.0
Kana-S	TGGATTGCACGCAGGTTCTC	58.0
Kana-A	CTCGATGCGATGTTTCGCTT	58.0

类别	名称	描述	核心序列	数量	位置/bp
Category	Name	Description	Core Sequence	Number	Position
结构元件 Structure elements	TATA-Box	转录起始-30核心启动子元件 Core promoter element around-30 of transcription start	TATATA (5)	16	123 ~-128,-1314 ~-1319, -1704 ~-1709, -1928 ~-1933, -1973 ~-1978
			TATAAA (1)		-595 ~-600
			TATA (7)		36 ~-39,-192~-195,-478 ~-481,-507 ~-510,-1208 ~-1211,-1753 ~-1756, -1778 ~-1781
			TTTATA (3)		-15 ~-20,-1385 ~ -1390,-1459 ~-1464
	CAAT-Box	启动子和增强子区域调控元件 Common cis-acting element in promoter and enhancer regions	CAAT (9)	15	-89 ~-92,-822 ~-825, -991 ~-994,-1002 ~ -1005,-1661 ~-1664, -1713 ~-1716,-1840 ~ -1843,-1965 ~-1968, -2010 ~-2013
	CAAT-Box		CAAAT (6)	15	-340 ~-344,-492 ~ -496,-1359 ~-1363, -1367 ~-1370,-1558 ~ -1561,-2068 ~-2072
胁迫响应元件Abiotic stress responsive elements	ARE	厌氧顺式调控元件 cis-Acting regulatory element essential for the anaerobic induction	AACCA	2	-584 ~-588, -1534 ~-1538
	MBS	干旱诱导MYB结合位点 MYB binding site involved in drought-inducibility	CAACTG CAACAG	2	-1512 ~-1517, -319 ~-324
	circadian	昼夜顺式调控元件 Circadian-control element	CAAAGTTATC	1	-1628 ~-1637
	TC-rich repeats	防卫和胁迫响应顺式调控元件 cis-Acting element involved in defense and stress responsiveness	GTTTTCTAA ATTCTCTAAC	2	-1127 ~-1135 -2159 ~-2168
激素响应元件Phytohor- mone responsive element	ABRE	脱落酸响应顺式调控元件 Abscisic acid responsive element	CACGTG	2	-977 ~-982, -881 ~-886
	CGTCA-motif	茉莉酸甲酯响应顺式调控元件 MeJA-responsive element	CGTCA	2	-165 ~-169, -1370 ~-1374
	ERE	乙烯响应顺式调控元件 Ethylene-responsive element	ATTTCAAA	1	-526 ~-533
	P-box	赤霉素响应顺式调控元件 Gibberellin-responsive element	CCTTTTG	1	-561 ~-567
	TGA-box	生长素响应顺式调控元件 Auxin-responsive element	CGGTGCAGT	1	-170 ~-178
生长发育调控元件 Development-related elements	Skn-1 motif	胚乳表达调控顺式作用元件 cis-Acting element involved in endosperm expression	GTCAT	5	-164 ~-168,-1005 ~-1009,-1785 ~-1789, -1956 ~-1960, -2023 ~-2027
生长发育调控元件 Development-related elements	GCN4 motif	胚乳表达调控顺式作用元件 cis-Acting element involved in endosperm expression	CAAGCCA	1	-1650 ~-1656

类别	名称	描述	核心序列	数量	位置/bp
Category	Name	Description	Core Sequence	Number	Position
结构元件 Structure elements	TATA-Box	转录起始-30核心启动子元件 Core promoter element around-30 of transcription start	TATATA (5)	16	123 ~-128,-1314 ~-1319, -1704 ~-1709, -1928 ~-1933, -1973 ~-1978
			TATAAA (1)		-595 ~-600
			TATA (7)		36 ~-39,-192~-195,-478 ~-481,-507 ~-510,-1208 ~-1211,-1753 ~-1756, -1778 ~-1781
			TTTATA (3)		-15 ~-20,-1385 ~ -1390,-1459 ~-1464
	CAAT-Box	启动子和增强子区域调控元件 Common cis-acting element in promoter and enhancer regions	CAAT (9)	15	-89 ~-92,-822 ~-825, -991 ~-994,-1002 ~ -1005,-1661 ~-1664, -1713 ~-1716,-1840 ~ -1843,-1965 ~-1968, -2010 ~-2013
	CAAT-Box		CAAAT (6)	15	-340 ~-344,-492 ~ -496,-1359 ~-1363, -1367 ~-1370,-1558 ~ -1561,-2068 ~-2072
胁迫响应元件Abiotic stress responsive elements	ARE	厌氧顺式调控元件 cis-Acting regulatory element essential for the anaerobic induction	AACCA	2	-584 ~-588, -1534 ~-1538
	MBS	干旱诱导MYB结合位点 MYB binding site involved in drought-inducibility	CAACTG CAACAG	2	-1512 ~-1517, -319 ~-324
	circadian	昼夜顺式调控元件 Circadian-control element	CAAAGTTATC	1	-1628 ~-1637
	TC-rich repeats	防卫和胁迫响应顺式调控元件 cis-Acting element involved in defense and stress responsiveness	GTTTTCTAA ATTCTCTAAC	2	-1127 ~-1135 -2159 ~-2168
激素响应元件Phytohor- mone responsive element	ABRE	脱落酸响应顺式调控元件 Abscisic acid responsive element	CACGTG	2	-977 ~-982, -881 ~-886
	CGTCA-motif	茉莉酸甲酯响应顺式调控元件 MeJA-responsive element	CGTCA	2	-165 ~-169, -1370 ~-1374
	ERE	乙烯响应顺式调控元件 Ethylene-responsive element	ATTTCAAA	1	-526 ~-533
	P-box	赤霉素响应顺式调控元件 Gibberellin-responsive element	CCTTTTG	1	-561 ~-567
	TGA-box	生长素响应顺式调控元件 Auxin-responsive element	CGGTGCAGT	1	-170 ~-178
生长发育调控元件 Development-related elements	Skn-1 motif	胚乳表达调控顺式作用元件 cis-Acting element involved in endosperm expression	GTCAT	5	-164 ~-168,-1005 ~-1009,-1785 ~-1789, -1956 ~-1960, -2023 ~-2027
生长发育调控元件 Development-related elements	GCN4 motif	胚乳表达调控顺式作用元件 cis-Acting element involved in endosperm expression	CAAGCCA	1	-1650 ~-1656

Expression Pattern and Regulation Mechanism of ApSK₃ Dehydrin （Agapanthus praecox）Response to Abiotic Stress and Hormone Signals

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