多花黄精<i>ACY1</i>基因家族鉴定及其表达分析

张慧; 罗彬彬; 朱田圆; 班景洁; 张春渝; 李小利; 朱建明; 林玉玲; 赖钟雄

doi:10.19303/j.issn.1008-0384.2023.06.005

多花黄精ACY1基因家族鉴定及其表达分析

doi: 10.19303/j.issn.1008-0384.2023.06.005

1.
福建农林大学园艺植物生物工程研究所，福建　福州　350002
2.
福建金湖源生态农业有限公司，福建　邵武　354099

基金项目: 福建省高原学科建设基金项目(102/71201801101)；福建农林大学横向科技项目(102/ KH200373A)

详细信息

作者简介:
张慧（1998 —），女，硕士，研究方向：设施环境与农业工程（E-mail：zhanghuid0609@163.com）

通讯作者:
赖钟雄（1966 —），男，博士，研究员，研究方向：园艺植物生物技术与遗传资源（E-mail:laizx01@163.com）

中图分类号: R282
计量
- 文章访问数: 275
- HTML全文浏览量: 81
- PDF下载量: 18
- 被引次数: 0
出版历程
- 收稿日期: 2022-12-15
- 修回日期: 2023-04-12
- 刊出日期: 2023-06-28

Identification and Functions of ACY1 Family in Polygonatum cyrtonema

1.
Institute of Horticultural Biotechnology, Fujian Agriculture and Forestry University, Fuzhou, Fujian　350002, China
2.
Fujian Jinhuyuan Eco-agriculture Co., Ltd., Shaowu, Fujian　354099, China

摘要

摘要: 目的筛选鉴定多花黄精中氨基酰化酶（aminoacylase 1，ACY1）基因，为多花黄精抗逆基因功能研究奠定基础。方法基于多花黄精转录组数据，对多花黄精氨基酰化酶（PcACY1）基因家族进行鉴定，分析基本理化性质、基因结构、保守基序、进化关系等，并采用qRT-PCR验证ACY1家族成员在不同组织器官中和盐胁迫处理下的表达模式。结果多花黄精ACY1家族有2个成员，均为亲水蛋白，具有信号肽，无跨膜结构，亚细胞定位预测结果显示2个成员均定位于细胞质中。与植物其他物种ACY1构建进化树发现，其进化特性可分为2类。植物ACY1启动子顺式作用元件预测结果显示，ACY1存在多种激素、胁迫和生长发育相关的响应元件。qRT-PCR分析结果显示，PcACY1成员在多花黄精不同组织器官中的表达水平差异较大且受盐胁迫诱导。结论 PcACY1在多花黄精生长发育和抵御逆境过程中可能发挥重要作用，这为进一步研究多花黄精ACY1的功能及遗传改良等奠定基础。
- 多花黄精 /
- ACY1 /
- 家族鉴定 /
- 表达分析
Abstract: Objective Family of aminoacylase 1 genes (ACY1) in Polygonatum cyrtonema Hua was identified, and associated resistance functions analyzed. Methods Based on the transcriptome data, the family of P. cyrtonema Hua aminoacylase genes (PcACY1) was identified. Physicochemical properties, structure, conserved motifs, and evolutionary relationships of the members were analyzed. Expressions of PcACY1 in tissues/organs and under salt stress were detected by qRT-PCR. Results The PcACY1 family had two members located in the cytoplasm, both of which were hydrophilic proteins with signal peptides and no transmembrane structure. As shown in the phylogenetic tree, they exhibited two categories of evolutionary characteristics. The predicted cis-acting elements of the plant ACY1 promoter indicated the presence of multiple hormone-, stress-, and growth/development-related functions. PcACY1 displayed significantly differentiated expressions in different tissues and organs which could be induced by salt stress. Conclusion PcACY1 appeared to play an important role in the growth and development as well as stress resistance of P. cyrtonema .
- Polygonatum cyrtonema Hua /
- ACY1 /
- gene family identification /
- expression analysis

HTML全文

图 1 PcACY1氨基酸序列比对

Figure 1. Amino acid sequences of PcACY1

下载: 全尺寸图片幻灯片

图 2 多花黄精PcACY1蛋白的三级结构空间构象

图2中紫色部分为预测的结合位点。

Figure 2. Spatial conformation of PcACY1 protein tertiary structure

Purple-colored portion in Fig. 2 indicates predicted binding site.

下载: 全尺寸图片幻灯片

图 3 PcACY1蛋白磷酸化位点分析和糖基化位点分析

A：PcACY1-1中预测磷酸位点位置，B：PcACY1-1中预测糖基化位点位置，C：PcACY1-2中预测磷酸位点位置，D：PcACY1-2中预测糖基化位点位置。

Figure 3. Phosphorylation and N-glycosylation sites of PcACY1 proteins

A: Predicted phosphorylation sites in PcACY1-1; B: predicted N-glycosylation sites in PcACY1-1; C: predicted phosphorylation sites in PcACY1-2; D: predicted N-glycosylation sites in PcACY1-2.

下载: 全尺寸图片幻灯片

图 4 植物ACY1家族成员进化树

蓝色区域（A-ClassⅡ和B-ClassⅥ）为单子叶植物，红色区域（A-ClassⅢ和B-ClassⅤ）为双子叶植物，黄色区域（A-ClassⅠ）为苔藓植物，紫色区域（B-ClassⅣ）为蕨类植物。

Figure 4. Phylogenetic tree of ACY1 family members in plants

Blue-colored areas (A-Class II and B-Class VI) are monocotyledons; red-colored areas (A-Class III and B-Class V), dicotyledons; yellow-colored areas (A-Class I), mosses; and purple-colored areas (B-Class IV), ferns.

下载: 全尺寸图片幻灯片

图 5 植物ACY1家族的基因结构（A）及其蛋白保守基序（B）分布

Figure 5. Structure (A) and conserved motifs (B) of encoded ACY1 proteins

下载: 全尺寸图片幻灯片

图 6 ACY1基因家族启动子顺式作用元件分析

Figure 6. Cis-acting elements of ACY1 family promoter

下载: 全尺寸图片幻灯片

图 7 ACY1基因家族核心启动子分布

Figure 7. Core promoter map in ACY1 family

下载: 全尺寸图片幻灯片

图 8 多花黄精ACY1家族成员在不同组织器官中的表达热图（A）和盐胁迫下的表达热图（B）

Figure 8. Expression heat map (A) of PcACY1 family members in tissues and organs of P. cyrtonema and expression heat map (B) under NaCl treatment

下载: 全尺寸图片幻灯片

图 9 拟南芥ACY1家族成员在不同胁迫下的表达热图（A）水稻ACY1家族成员在不同胁迫下的表达热图（B）

Figure 9. Expression heat maps of PcACY1 family members of Arabidopsis thaliana (A) and of Oryza sativa (B) under abiotic stresses

下载: 全尺寸图片幻灯片

图 10 PcACY1 家族成员在多花黄精不同的组织器官（A）和不同浓度NaCl处理下（B）的表达

不同小写字母表示同一基因不同处理间差异显著（P＜0. 05）。

Figure 10. Expressions of PcACY1 family members in tissues and organs of P. cyrtonema (A) and under treatments of different concentration of NaCl (B)

Datas with different lowercase letters indicate significant differences between different treatments of the same gene at P＜0.05.

下载: 全尺寸图片幻灯片

表 1 qRT-PCR引物

Table 1. Primers for qRT-PCR

引物名称 Primer name	引物序列（5′-3′） Primer sequences( 5′-3′)
PcACY1-1F	CCTCCTCTCCCTCCTCGAAA
PcACY1-1R	GAGTTGAGGAGGAGGGAGGA
PcACY1-2F	CTCAACTCCCACACCGACTC
PcACY1-2R	GTACTGGATGGCGATGCACT
ACTIN-F	GGGCTCATAGACTGATCACAAG
ACTIN-R	TTGTGCTTACCGACCTAATGG

下载: 导出CSV

表 2 多花黄精ACY1基因家族成员信息

Table 2. Information on ACY1 family of P. cyrtonema

基因名称 Gene name	转录组基因ID Accession No.	编码区 CDS/bp	长度 Length/aa	分子量 MW/kD	等电点 PI	不稳定指数 Instability index (II)	亲水性 Gravity	信号肽 Singal peptide	脂肪指数 Aliphatic Index	跨膜结构 Trans membrane domain	亚细胞定位 Subcellular localization
PcACY1-1	A21014918a_transcript_7940	1371	456	50.06	5.35	32.99	−0.173	YES	87.52	0	细胞质 Cytoplasm
PcACY1-2	A21014918a_transcript_9264	1317	438	48.02	5.42	50.75	−0.103	YES	86	0	细胞质 Cytoplasm

下载: 导出CSV

表 3 PcACY1蛋白保守结构域

Table 3. Conserved domain of PcACY1 protein

基因名称 Gene name	登录号 Accession	名称 Name	位置 Matches
PcACY1-1	IPR002933	Peptidase_M20
	IPR001261	ArgE/DapE_CS
	IPR010159	N-acyl_aa_amidohydrolase
PcACY1-2	IPR001261	ArgE/DapE_CS
	IPR002933	Peptidase_M20
	IPR036264	Bact_exopeptidase_dim_dom
	IPR011650	Peptidase_M20_dimer
	IPR010159	N-acyl_aa_amidohydrolase

下载: 导出CSV

表 4 植物ACY1家族成员分布和数量

Table 4. Distribution and number of ACY1 family members in plants

分类 Classification				物种 Species	数量 Number
门 Divisio	纲 Classis	目 Ordo	科 Quncs	物种 Species	数量 Number
苔藓植物门 Bryophyta	藓纲 Musci	真藓目 Eubryales	葫芦藓科 Funariaceae	小立碗藓 Pp	2
蕨类植物门 Pteridophyta	石松纲 Lycopodiinae	卷柏目 Selaginellales	卷柏科 Selaginellaceae	江南卷柏 Sm	1
被子植物门 Angiosperms	单子叶植物纲 Monocotyledons	禾本目 Poales	禾本科 Poaceae	水稻 Os	2
				小麦 Ta	2
				玉米 Zm	2
				二穗短柄草 Bd	2
				大麦 Hv	2
				双色高粱 Sb	2
		百合目 Liliflorae	百合科 Liliaceae	石刁柏 Ao	2
		粉状胚乳目 Farinosae	凤梨科 Bromeliaceae	菠萝 Ac	2
		芭蕉目 Scitamineae	芭蕉科 Musaceae	香蕉 Ma	2
		百合目 Liliflorae	薯蓣科 Dioscoreaceae	薯蓣 Dc	2
	双子叶植物纲 Dicotyledons	蔷薇目 Rosales	豆科 Leguminosae	大豆 Gm	2
				花生 Ad	2
				豇豆 Va	2
				鹰嘴豆 Cr	2
				菜豆 Pv	2
			蔷薇科 Rosaceae	野草莓 Fv	2
				日本梅 Pm	2
				苹果 Md	2
		十字花目 Brassicales	十字花科 Brassicaceae	拟南芥 At	2
				芜菁 Br	2
				甘蓝 Bo	2
		茄目 Solanales	茄科 Solanaceae	马铃薯 St	2
		茄目 Solanales	茄科 Solanaceae	番茄 Le	2
		葫芦目 Cucurbitales	葫芦科 Cucurbitaceae	黄瓜 Cu	2
		无患子目 Sapindales	芸香科 Rutaceae	甜橙 Cs	2
		葡萄目 Vitales	葡萄科 Vitaceae	葡萄 Vv	2
		桃金娘目 Myrtales	桃金娘科 Myrtaceae	巨桉 Eg	2
		伞形目 Apiales	伞形科 Apiaceae	胡萝卜 Da	3
		壳斗目 Fagales	胡桃科 Juglandaceae	山核桃 Ci	3
		菊目 Asterales	菊科 Asteraceae	莴苣 Ls	2

下载: 导出CSV

参考文献(29)

[1]	于冰. ACY-1在结直肠癌中的表达及作用机制[D]. 济南: 山东大学, 2017. YU B. Expression and mechanism of ACY-1 in colorectal cancer [D]. Jinan: Shandong University, 2017. (in Chinese)
[2]	COOK R M, BURKE B J, BUCHHAGEN D L, et al. Human aminoacylase-1. Cloning, sequence, and expression analysis of a chromosome 3p21 gene inactivated in small cell lung cancer [J]. Journal of Biological Chemistry, 1993, 268(23): 17010−17017. doi: 10.1016/S0021-9258(19)85294-8
[3]	LINDNER H A, LUNIN V V, ALARY A, et al. Essential roles of zinc ligation and enzyme dimerization for catalysis in the aminoacylase-1/M20 family [J]. Journal of Biological Chemistry, 2003, 278(45): 44496−44504. doi: 10.1074/jbc.M304233200
[4]	FONES W S, LEE M. Hydrolysis of n-acyl derivatives of alanine and phenylalanine by acylase i and carboxypeptidase [J]. Journal of Biological Chemistry, 1953, 201(2): 847−856. doi: 10.1016/S0021-9258(18)66242-8
[5]	POLEVODA B, SHERMAN F. Nalpha-terminal acetylation of eukaryotic proteins [J]. The Journal of Biological Chemistry, 2000, 275(47): 36479−36482. doi: 10.1074/jbc.R000023200
[6]	SHI H J, HAYES M T, KIRANA C, et al. Overexpression of aminoacylase 1 is associated with colorectal cancer progression [J]. Human Pathology, 2013, 44(6): 1089−1097. doi: 10.1016/j.humpath.2012.09.015
[7]	PERRIER J, DURAND A, GIARDINA T, et al. Catabolism of intracellular N-terminal acetylated proteins: Involvement of acylpeptide hydrolase and acylase [J]. Biochimie, 2005, 87(8): 673−685. doi: 10.1016/j.biochi.2005.04.002
[8]	张露莹. ACY1调控mTOR信号通路的机制[D]. 广州: 广州医科大学, 2020. ZHANG L Y. Mechanism of ACY1 regulation of mTOR signaling pathway [D]. Guangzhou: Guangzhou Medical University, 2020. (in Chinese)
[9]	国家药典委员会. 中华人民共和国药典: 一部[M]. 北京: 中国医药科技出版社, 2010.
[10]	中国科学院中国植物志编辑委员会. 中国植物志: 第十五卷[M]. 北京: 科学出版社, 1978.
[11]	BIRNBAUM S M, LEVINTOW L, KINGSLEY R B, et al. Specificity of amino acid acylases [J]. Journal of Biological Chemistry, 1952, 194(1): 455−470. doi: 10.1016/S0021-9258(18)55898-1
[12]	CHEN Z M, GAO J H, SUN J, et al. Aminoacylase 1 (ACY-1) mediates the proliferation and migration of neuroblastoma cells in humans through the ERK/transforming growth factor β (TGF-β) signaling pathways [J]. Medical Science Monitor, 2020, 27: e928813.
[13]	TYLKI-SZYMANSKA A, GRADOWSKA W, SOMMER A, et al. Aminoacylase 1 deficiency associated with autistic behavior [J]. Journal of Inherited Metabolic Disease, 2010, 33(3): 211−214.
[14]	SASS J O, VAITHILINGAM J, GEMPERLE-BRITSCHGI C, et al. Expanding the phenotype in aminoacylase 1 (ACY1) deficiency: Characterization of the molecular defect in a 63-year-old woman with generalized dystonia [J]. Metabolic Brain Disease, 2016, 31(3): 587−592. doi: 10.1007/s11011-015-9778-6
[15]	ZHONG Y, ONUKI J, YAMASAKI T, et al. Genome-wide analysis identifies a tumor suppressor role for aminoacylase 1 in iron-induced rat renal cell carcinoma [J]. Carcinogenesis, 2009, 30(1): 158−164. doi: 10.1093/carcin/bgn255
[16]	CHENG Q A, GU S H, LIU Z W, et al. Expressional divergence of the fatty acid-amino acid conjugate-hydrolyzing aminoacylase 1 (L-ACY-1) in Helicoverpa armigera and Helicoverpa assulta [J]. Scientific Reports, 2017, 7: 8721. doi: 10.1038/s41598-017-09185-2
[17]	JIN G Z, DONG H, YU W L, et al. A novel panel of biomarkers in distinction of small well-differentiated HCC from dysplastic nodules and outcome values [J]. BMC Cancer, 2013, 13: 161. doi: 10.1186/1471-2407-13-161
[18]	STOCKER P, BRUNEL J M, DE REZENDE L, et al. Aminoacylase 1-catalysed deacetylation of bioactives epoxides mycotoxin-derived mercapturates; 3, 4-epoxyprecocenes as models of cytotoxic epoxides [J]. Biochimie, 2012, 94(8): 1668−1675. doi: 10.1016/j.biochi.2012.01.006
[19]	KOREISHI M, NAKATANI Y, OOI M, et al. Purification, characterization, molecular cloning, and expression of a new aminoacylase from Streptomyces mobaraensis that can hydrolyze N-(middle/long)-chain-fatty-acyl-L-amino acids as well as N-short-chain-acyl-L-amino acids [J]. Bioscience, Biotechnology, and Biochemistry, 2009, 73(9): 1940−1947. doi: 10.1271/bbb.90081
[20]	CHEN D B, LI J H, JIAO F C, et al. ZmACY-1 antagonistically regulates growth and stress responses in Nicotiana benthamiana [J]. Frontiers in Plant Science, 2021, 12: 593001. doi: 10.3389/fpls.2021.593001
[21]	NAKANO M, NISHIHARA M, YOSHIOKA H, et al. Silencing of DS2 aminoacylase-like genes confirms basal resistance to Phytophthora infestans in Nicotiana benthamiana [J]. Plant Signaling & Behavior, 2014, 9(2): e28004.
[22]	CHEN C J, CHEN H, ZHANG Y, et al. TBtools: An integrative toolkit developed for interactive analyses of big biological data [J]. Molecular Plant, 2020, 13(8): 1194−1202. doi: 10.1016/j.molp.2020.06.009
[23]	BAUMEIER C, KAISER D, HEEREN J, et al. Caloric restriction and intermittent fasting alter hepatic lipid droplet proteome and diacylglycerol species and prevent diabetes in NZO mice [J]. Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids, 2015, 1851(5): 566−576. doi: 10.1016/j.bbalip.2015.01.013
[24]	LINDNER H A, TÄFLER-NAUMANN M, RÖHM K H. N-acetylamino acid utilization by kidney aminoacylase-1 [J]. Biochimie, 2008, 90(5): 773−780. doi: 10.1016/j.biochi.2007.12.006
[25]	CAIRA S, IANNELLI A, SCIARRILLO R, et al. Differential representation of liver proteins in obese human subjects suggests novel biomarkers and promising targets for drug development in obesity [J]. Journal of Enzyme Inhibition and Medicinal Chemistry, 2017, 32(1): 672−682. doi: 10.1080/14756366.2017.1292262
[26]	KUHNS E H, SEIDL-ADAMS I, TUMLINSON J H. A lepidopteran aminoacylase (L-ACY-1) in Heliothis Virescens (Lepidoptera: Noctuidae) gut lumen hydrolyzes fatty acid-amino acid conjugates, elicitors of plant defense [J]. Insect Biochemistry and Molecular Biology, 2012, 42(1): 32−40. doi: 10.1016/j.ibmb.2011.10.004
[27]	KUTMON M, RIUTTA A, NUNES N, et al. WikiPathways: Capturing the full diversity of pathway knowledge [J]. Nucleic Acids Research, 2016, 44(D1): D488−D494. doi: 10.1093/nar/gkv1024
[28]	WU G, MORRIS S. Arginine metabolism: nitric oxide and beyond [J]. Biochemical Journal, 1998: 331-336.
[29]	LINDNER H, HÖPFNER S, TÄFLER-NAUMANN M, et al. The distribution of aminoacylase I among mammalian species and localization of the enzyme in porcine kidney [J]. Biochimie, 2000, 82(2): 129−137. doi: 10.1016/S0300-9084(00)00191-7