马氏珠母贝PfAQP4免疫功能初探

doi:10.11978/2020050

热带海洋学报 ›› 2021, Vol. 40 ›› Issue (2): 83-89.doi: 10.11978/2020050CSTR: 32234.14.2020050

马氏珠母贝PfAQP4免疫功能初探

潘肖兰^1,²(), 刘惠茹^1,², 许濛^1,², 许瀚之^1,², 张华¹, 何毛贤¹()

1.中国科学院南海海洋研究所, 中国科学院热带海洋生物资源与生态重点实验室, 广东省应用海洋生物学重点实验室, 中国科学院南海生态环境工程创新研究院, 广东广州 510301
2.中国科学院大学, 北京 100049

收稿日期:2020-05-22 修回日期:2020-08-16 出版日期:2021-03-10 发布日期:2020-08-31
通讯作者: 何毛贤
作者简介:潘肖兰(1996—), 女, 广西壮族自治区河池市人, 硕士研究生, 研究方向为贝类分子遗传。email: panxiaolan17@mails.ucas.edu.cn
基金资助:
国家贝类产业技术体系岗位科学家项目(CARS-49);中国科学院南海生态环境工程创新研究院自主部署项目(ISEE2018PY03);以农产品为单元的广东省现代农业产业技术体系创新团队建设项目(贝藻产业)(2019KJ146);广东省科技计划项目(2017B0303014052)

Preliminary study on immunity function of aquaporin AQP4 in the pearl oyster Pinctada fucata martensii

PAN Xiaolan^1,²(), LIU Huiru^1,², XU Meng^1,², XU Hanzhi^1,², ZHANG Hua¹, HE Maoxian¹()

1. CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangdong Provincial Key Laboratory of Applied Marine Biology, Institution of South China Sea Ecology and Environmental Engineering, Chinese Academy of Sciences, Guangzhou 510301, China
2. University of Chinese Academy of Sciences, Beijing 100049, China

Received:2020-05-22 Revised:2020-08-16 Online:2021-03-10 Published:2020-08-31
Contact: HE Maoxian
Supported by:
Earmarked Fund for Modern Agro-industry Technology Research System(CARS-49);Innovation Academy of South China Sea Ecology and Environmental Engineering, Chinese Academy of Sciences(ISEE2018PY03);Shellfish and Large Algae Industry Innovation Team Project of Guangdong Province, China(2019KJ146);Science and Technology Planning Project of Guangdong Province, China(2017B0303014052)

摘要/Abstract

摘要：

文章对马氏珠母贝水通道蛋白基因PfAQP4进行免疫功能分析。利用实时荧光定量分析免疫刺激和RNA干扰后PfAQP4及免疫相关基因的表达模式, 旨在为马氏珠母贝自然免疫研究提供新的资料。结果表明, 注射脂多糖LPS后, 在外套膜组织中PfAQP4 mRNA表达量在24h时显著上调(p<0.05), 在消化腺中PfAQP4 mRNA表达量先在12h时显著下调(p<0.05), 后在24h、36h和48h时显著上调(p<0.05)。注射聚肌胞poly(I:C)后, 在外套膜组织中PfAQP4 mRNA表达量在36h时和48h时显著上调(p<0.05), 在消化腺中PfAQP4 mRNA表达量在48h时显著上调(p<0.05)。RNA干扰PfAQP4表达后, 免疫相关基因CuZn-SOD在外套膜中的表达量显著下降(p<0.05), Pearson相关性分析显示PfAQP4与CuZn-SOD的表达存在显著正相关(r=0.818, p<0.001)。综上所述, PfAQP4参与马氏珠母贝的免疫应答。

关键词: 马氏珠母贝, 水通道蛋白4基因, 免疫刺激, RNA干扰, 基因表达

Abstract:

To identify the immunity function of Aquaporin of Pinctada fucata martensii (PfAQP4), the mRNA expression patterns of PfAQP4 and immune-related genes were detected by qPCR after immunostimulation and RNA interference. The results showed that the mRNA expression of PfAQP4 in the mantle was significantly up-regulated at 24 h (p < 0.05), while that in the digestive gland was significantly down-regulated at 12 h, and increased at 24 h, 36 h and 48 h after LPS injection (p < 0.05). After poly (I : C) injection, the expression of PfAQP4 was significantly up-regulated in the mantle at 36 h and 48 h (p < 0.05), and up-regulated at 48 h in the digestive gland (p < 0.05). When PfAQP4 was knocked down by RNA interference, the mRNA expression of CuZn-SOD in the mantle was significantly down-regulated (p < 0.05); a significant positive correlation of expression level between CuZn-SOD and PfAQP4 was found (r = 0.818, p < 0.001). We showed that PfAQP4 plays an important role in the immune response of P. f. martensii, which provide new data for the innate immunity of P. f. martensii.

Key words: Pinctada fucata martensii, aquaporin 4, immunoregulation, RNA interference, gene expression

中图分类号:

P735.4

潘肖兰, 刘惠茹, 许濛, 许瀚之, 张华, 何毛贤. 马氏珠母贝PfAQP4免疫功能初探[J]. 热带海洋学报, 2021, 40(2): 83-89.

PAN Xiaolan, LIU Huiru, XU Meng, XU Hanzhi, ZHANG Hua, HE Maoxian. Preliminary study on immunity function of aquaporin AQP4 in the pearl oyster Pinctada fucata martensii[J]. Journal of Tropical Oceanography, 2021, 40(2): 83-89.

图/表 4

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引物名称	引物序列(5' — 3')	用途
dsAQP4-T7F	GGATCCTAATACGACTCACTATAGGTTGCCTTTGGGCTATCGG	PfAQP4 RNAi含T7启动子序列正向引物
dsAQP4-T7R	GGATCCTAATACGACTCACTATAGGCGGAAGGAGCAGCAAAGAC	PfAQP4 RNAi含T7启动子序列反向引物
dsGFP4-T7F	GGATCCTAATACGACTCACTATAGGCGACGTAAACGGCCACAAGTT	GFP RNAi含T7启动子序列正向引物
dsGFP-T7R	GGATCCTAATACGACTCACTATAGGATGGGGGTGTTCTGCTGGTAG	GFP RNAi含T7启动子序列反向引物
dsAQP4-F	TTGCCTTTGGGCTATCGG	PfAQP4 RNAi正向引物
dsAQP4-R	CGGAAGGAGCAGCAAAGAC	PfAQP4 RNAi反向引物
dsGFP-F	CGACGTAAACGGCCACAAGTT	GFP RNAi正向引物
dsGFP-R	ATGGGGGTGTTCTGCTGGTAG	GFP RNAi反向引物
Faqp	CTGTGATGGCGTCAACTGATG	PfAQP4 qPCR正向引物
Raqp	CGCAATCGTGAGAAGATGACC	PfAQP4 qPCR反向引物
F18S	CGTTTCAACAAGACGCCAGTAG	18S qPCR正向引物
R18S	ACGAAAAAAAGGTTTGAGAGACG	18S qPCR反向引物
QT5Rel143	GATGGCAGAGGATGATTCTTCTT	NF-κB qPCR正向引物
QT3Rel332	TGATGGACCTTCACACTCATACC	NF-κB qPCR反向引物
MMPF	TCTGGCTCATGCGTTTTTCC	MMP qPCR正向引物
MMPR	AGGGCATGTCCAATCTCATGAG	MMP qPCR反向引物
QT5IL17seq460	CCAGTCCTCGTAATAAATGTGAACC	IL-17 qPCR正向引物
QT3IL17seq636	CGCTTCCGCTGCTAGATTCTT	IL-17 qPCR反向引物
LAMPF	TGCCTGTCACAATAATAACC	LAMP qPCR正向引物
LAMPR	AGACTCAAAGTAAGACCACCT	LAMP qPCR反向引物
CuZn-SODF	GAAACTGCAAACCCTATA	CuZn-SOD qPCR正向引物
CuZn-SODR	AGACATTCGTCAAACTCA	CuZn-SOD qPCR反向引物

基因名称	相关系数	p值
基质金属蛋白分解酶	0.356	0.212
核因子κB	0.655*	0.011
白细胞介素17	0.515	0.060
溶酶体膜相关糖蛋白	0.674**	0.008
铜锌超氧化物歧化酶	0.818***	0.000

马氏珠母贝PfAQP4免疫功能初探

Preliminary study on immunity function of aquaporin AQP4 in the pearl oyster Pinctada fucata martensii

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