厚壳贻贝（<i>Mytilus coruscus</i>）视蛋白基因的鉴定、进化及表达分析

徐敏慧; 李继姬; 叶莹莹

厚壳贻贝（Mytilus coruscus）视蛋白基因的鉴定、进化及表达分析

浙江海洋大学国家海洋设施养殖工程技术研究中心，浙江舟山 316022

基金项目: 舟山市科技计划项目（2021C21017）。

详细信息

作者简介:
徐敏慧（1999—），女，浙江省衢州人，硕士研究生，主要研究方向为海洋生物学。E-mail：xuminhui@zjou.edu.cn

通讯作者:
叶莹莹（1986—），博士，副研究员，主要研究方向为海洋生物学。E-mail：yeyy@zjou.edu.cn

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出版历程
- 收稿日期: 2024-06-14
- 修回日期: 2025-03-24
- 网络出版日期: 2025-04-24

Analysis of opsin gene expression in Mytilus coruscus

Xu Minhui^1
,,
Li Jiji¹,
Ye Yingying^{1
, ,}

National Engineering Research Center for Marine Aquaculture, Zhejiang Ocean University, Zhoushan 316022, China

摘要

摘要: 厚壳贻贝是我国重要的经济养殖贝类，光环境可以影响其生活和行为，但其视蛋白基因在很大程度上是未知的。本研究基于厚壳贻贝全基因组数据，共鉴定出7个视蛋白，序列分析结果将其分为五类（r-opsin、c-opsin、Go-opsin、neuropsin和peropsins）。染色体定位结果显示同一亚家族的视蛋白基因位于同一染色体上。生物信息学结果显示，除r-opsin为亲水蛋白，其他蛋白质的均为疏水蛋白。保守基序结果显示视蛋白同一亚家族的视蛋白具有高度的保守性，同时也存在一定的差异。通过实时荧光定量PCR检测厚壳贻贝五个发育时期的视蛋白基因表达量变化。结果显示，不同发育时期的视蛋白基因表达量存在显著差异，c-opsin4和r-opsin在眼点期的表达量显著高于其他时期，表明这两个视蛋白在厚壳贻贝的眼点期中发挥了重要作用。本研究分析了厚壳贻贝视蛋白的分子特征，并初步探究厚壳贻贝发育过程中视蛋白基因的变化规律，为研究厚壳贻贝的视觉形成机制并对视蛋白调控厚壳贻贝变态过程关键分子机制提供科学依据。
- 厚壳贻贝 /
- 浮游幼虫 /
- 视蛋白 /
- 表达分析 /
- qPCR
Abstract: Mytilus coruscus is an economically significant shellfish cultivated in China, with its life and behavior greatly influenced by light conditions. However, much remains unknown about its opsin genes. In this study, seven opsins were identified based on whole-genome sequencing data of M. coruscus, and sequence analysis classified them into five types: r-opsin, c-opsin, Go-opsin, neuropsin, and peropsin. Chromosomal localization analysis revealed that opsin genes of the same subfamily are located on the same chromosome. Bioinformatics analysis showed that, except for r-opsin, all identified proteins are hydrophobic. The conserved motifs revealed high sequence conservation among opsin subfamily members, while inter-subfamily comparisons identified specific divergent residues. The expression profiles of opsin genes were examined across five developmental stages using real-time quantitative PCR, which demonstrated significant expression differences at various developmental stages. Notably, c-opsin4 and r-opsin were significantly upregulated during the eyespot stage, suggesting their crucial roles during this period.This study provides insights into the molecular characteristics of opsins in M. coruscus and preliminarily explores the expression patterns of opsin genes during its development. Additionally, it contributes to the understanding of visual formation mechanisms in M. coruscus and offers a scientific basis for further exploration of the regulatory role of opsins in the metamorphosis process.
- Mytilus coruscus /
- XXXX /
- opsin /
- expression analysis /
- qPCR

HTML全文

图 1 视蛋白基因多序列比对图

红色字体和蓝色字体分别代表本文鉴定的厚壳贻贝和深海偏顶蛤视蛋白序列。图中灰色虚线代表视紫红质的第Ⅲ、Ⅳ和Ⅶ跨膜区。依据牛视紫红质对其他物种中opsin序列的氨基酸位置进行编号，我们可以在其中观察到一些被用于区分opsin家族的保守的氨基酸序列位置，这些位置用红色三角箭头表示。同时，蓝色方框中还标注了r-opsin和c-opsin的特征序列“HPK”和“NKQ”

Fig. 1 Multiple sequence alignment of opsin proteins

The red and blue fonts represent the opsin sequences identified in this paper, respectively. The dashed gray lines in the figure represent the transmembrane regions Ⅲ, Ⅳ, and Ⅶ of the rhodopsin. In the numbering of amino acid positions of opsin sequences in other species by rhodopsin, we can observe some conserved amino acid sequence positions that are used to distinguish the opsin family, and these positions are indicated by red triangular arrows. At the same time, the feature sequences "HPK" and "NKQ" of r-opsin and c-opsin are also marked in the blue box

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图 2 视蛋白家族ML系统发育分析

厚壳贻贝的视蛋白序列名用橙色星号标出，深海偏顶蛤的视蛋白序列名用蓝色圆形标出。不同颜色的分枝代表不同的亚家族，树枝上的数字表示ML后验概率

Fig. 2 Phylogenetic relationship of opsin family based on a ML analysis of protein sequences

The opsin sequence names of Mytilus coruscus are marked with orange asterisks, and those of Bathymodiolus platifrons are marked with blue circles. Different colored branches represent different subfamilies, and the numbers on the branches represent ML posteriori probabilities

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图 3 厚壳贻贝视蛋白基因的染色体定位图

Fig. 3 Chromosomal mapping of the opsin genes of M.coruscus

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图 4 厚壳贻贝视蛋白保守基序结构图

Fig. 4 Analysis of conserved motif of M.coruscus opsin family members

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图 5 厚壳贻贝视蛋白家族基因在不同发育时期的表达模式

字母不同表示同一时间各实验组之间存在显著性差异(p <0.05)

Fig. 5 Expression of opsin family genes in different development stages in M.coruscus

Values with the same letters are not significantly different at the same time (p <0.05)

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表 1 视蛋白家族序列及其NCBI登录号

Tab. 1 Sequence of opsin family and their NCBI accession number

物种名 Species	蛋白 Proten	登录号 Accession number	物种名 Species	蛋白 Protein	登录号 Accession number
文昌鱼 Branchiostoma belcheri	opsin6	BAC76024	光滑双脐螺 Biomphalaria glabrata	pineal opsin like	XP_055862616
	melanopsin	Q4R114		opsin5 like	XP_055889746
	opsin2	BAC76020	葡萄牙牡蛎 Crassostrea angulata	opsin VA like	XP_052683732
	peropsin	DAC74078		pineal opsin like	XP_052683447
	opsin1	BAC76019	戴河笠贝 Lottia peitaihoensis	r-opsin	WKW95581
	opsin3	BAC76023		neurospin	WKW95586
	opsin5	BAC76022		Go-opsin3	WKW95588
	opsin4	BAC76021		peropsin	WKW95585
	neuropsin	DAC74071	虎斑乌贼 Sepia pharaonis	RRH	CAE1253240
海湾扇贝 Argopecten irradians	opsin Gq2	ALO02515		opsin4	CAE1318583
	opsin Gq3	ALO02516	地中海贻贝 Mytilusgalloprovincialis	r-opsin	VDI13980
	opsin Gq4	ALO02517	虾夷扇贝 Mizuhopecten yessoensis	Go-coupled opsin	XP_021355729
	opsin Gq1	ALO02514	日本大鲍 Haliotis gigantea	rhodopsin	UVT38273
	Rhabdomeric opsin	AUR34014	欧洲扇贝 Pecten maximus	retinochrome	UUS63783
	Go-coupled opsin	AUR34012	夏威夷短尾乌贼 Euprymna scolopes	opsin	ACB05673
	Go-coupled opsin1	APB88014	乌贼 Sepia officinalis	rhodopsin	ADT78509
人 Homo sapiens	encephalopsin	NP055137	尖膀胱螺 Physella acuta	opsin 3 like	XP_059165397
	opsin3	AAH36773	硬壳蛤 Mercenaria mercenaria	opsin 5 like	XP_045202610
	blue opsin	P0399	砂海螂 Mya arenaria	opsin 5 like	XP_052784401
	rhodopsin	NP000530	Ambigolimax valentianus	rhodopsin	BBA21101
	green opsin	P04001	Ambigolimax valentianus	opsin 5B	BBH84662
	red opsin	P04000	Acanthopleura granulata	r-opsin	r-opsin
	melanopsin	NP150598	Ambigolimax valentianus	retinochrome	BBH84663
	opsin5	QU6736	Chiton virgulatus	opsin	APF30602
	peropsin	NP006574	Chiton tuberculatus	r-opsin	QKY89064
鼠 Mus musculus	melanopsin	AAF24979	Chiton marmoratus	r-opsin	QKY89065
	peropsin	AAC53344	Chaetopleura apiculata	r-opsin	QKY89066
	opsin5	NP861418	Elysia marginata	rhodopsin	GFS12646
褐家鼠 Rattus norvegicus	opsin5	NP861437	Leptochiton asellus	xenopsin	ASM47594
厚壳贻贝 Mytilus coruscus	opsin3	CAC5413267	Stenoplax conspicua	opsin	APF30603
	opsin5	CAC5413461	Ylistrum balloti	opsin 5 like	XP_060063722
紫贻贝 Mytilus edulis	opsin5	CAG2190128
	RRH	CAG2194598
	opsin4	CAG2193259

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表 2 本实验所用的引物序列

Tab. 2 Primer sequences used in this study

蛋白Protein	引物序列 Primer sequence（5'-3'）
c-opsin4	CATTAGTATGGGCGGTTCCTC
c-opsin4	TCGTGTTTCCTTATTGCTTTCC
r-opsin	TGAAAACCCCGAGCAACAT
r-opsin	CATCGCCCCACATCCATT
Go-opsin3	TAGGAAGGTAACATTGGCGTCTC
Go-opsin3	GCTGCTCTTGGCAAACATCG
opsin5	CAAACGCCGAGATGCATAA
opsin5	TGAACTGATACTATTAGAACACGGA
RRH	ATGCGGACTAAACTGGCTAAAA
RRH	CTCCTGACCACGGGTAACAAC
EF1a	CACCACGAGTCTCTCCCTGA
EF1a	GCTGTCACCACAGACCATTCC

下载: 导出CSV

表 3 厚壳贻贝视蛋白家族序列基本信息

Tab. 3 Basic information of opsin family members in M.coruscus

基因 Gene	氨基酸数目 Number of amino acids	等电点 Theoretical isoelectric point (PI)	分子量 Molecular weight (KD)	不稳定性指数 Instability index	脂肪族指数 Aliphatic index	GRAVY指数 GRAVY index
c-opsin4	416	9.05	46.18	45.90	101.42	0.316
c-opsin4	424	9.21	47.30	442.81	99.93	0.341
r-opsin	619	8.39	69.22	40.88	83.94	-0.145
Go-opsin3	326	9.05	37.22	29.62	114.42	0.165
opsin5	317	9.44	35.98	42.10	117.98	0.702
RRH	324	7.46	36.48	37.02	99.85	0.527
RRH	303	6.96	33.83	33.27	99.70	0.525

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