母源性长链非编码RNA Six2a-OT参与斑马鱼胚胎发育调控  

Maternal Long Noncoding RNA Six2a-OT Involved in Embryonic Development of Zebrafish

作　　者：景新星 宋佳坤[1,2,3,4] 范纯新 JING Xin-xing;SONG Jia-kun;FAN Chun-xin(Institute for Marine Biosystem and Neurosciences,Shanghai Ocean University,Shanghai 201306,China;Shanghai Universities Key Laboratory of Marine Animal Taxonomy and Evolution,Shanghai 201306,China;National Demonstration Center for Experimental Fisheries Science Education,Shanghai 201306,China;Shanghai Collaborative Innovation for Aquatic Animal Genetics and Breeding,Shanghai 201306,China)

机构地区：[1]上海海洋大学海洋生物系统和神经科学研究所,中国上海201306 [2]海洋动物系统分类与进化上海高校重点实验室,中国上海201306 [3]水产科学国家级实验教学示范中心,中国上海201306 [4]水产动物遗传育种中心上海市协同创新中心,中国上海201306

出　　处：《生命科学研究》2018年第5期363-369,共7页Life Science Research

基　　金：上海海洋大学国际海洋研究中心资助项目(A-0209-15-0802);国家自然科学基金资助项目(31772406)

摘　　要：Six基因家族进化高度保守,在脊椎动物个体发育中起着重要作用。但目前对其调控机制尚不完全清楚。在UCSC (The University of California Santa Cruz) Genome Browser的斑马鱼基因组数据库中发现, Six2a-Six3a基因簇周围存在一条长链非编码RNA——Six2a-OT。首先,利用反转录PCR (RT-PCR)结合测序证实了该RNA确实存在。然后,利用荧光半定量PCR (qPCR)和整胚原位杂交(whole-mount in situ hybridization, WISH)确定了其特异性的表达模式:Six2a-OT在卵巢中的表达远高于精巢;在受精卵和早期原肠胚中高表达,随后显著下降;受精后48 h (48 hpf)集中表达于耳囊(otic vesicle)、躯干中线(trunk midline)和鳃弓(branchial archs)。随后,进一步利用反义吗啡啉(morpholino)特异性阻断Six2a-OT剪接,观察到63%的48 hpf胚胎表现出躯干变短、侧弯,或心包积水,同时Six2a和Six3a的表达显著下降。以上结果表明:Six2a-OT作为母源性因子通过调节Six2a和Six3a的表达参与斑马鱼胚胎早期发育。这为揭示Six基因簇的表达调控机制提供了重要依据。The Six gene family is highly conserved. It plays an important role in vertebrate development, but its regulatory mechanism is not fully understood. A long noncoding RNA around Six2a-Six3a gene cluster was found in the zebrafish genome database from UCSC （The University of California Santa Cruz） Genome Browser. The existence of this RNA was confirmed by reverse transcription PCR （RT-PCR） and sequencing. The specific expression pattern of Six2a-OT was revealed by real-time quantitative PCR （qPCR） and whole- mount in situ hybridization （WISH）： the expression of Six2a-OT in ovary was much higher than in testis, and it was expressed strongly in fertilized eggs and at early gastrula stage, and then decreased significantly. Its expression product was concentrated in the otic vesicle, midline of trunk and branchial archs 48 hours post- fertilization （48 hpf）. Furthermore, after Six2a-OT splicing was blocked with antisense morpholino oligos, 63% of the 48 hpf embryos showed shortened trunk, lateral curvature or edema, and the expression of Six2a and Six3a decreased significantly. Taken together, Six2a-OT was involved in the early embryonic development of zebrafish through regulating the expression of Six2a and Six3a. This study will help us understand the regu- lation of Six gene cluster expression.

关 键 词：斑马鱼 Six基因簇 长链非编码RNA(lncRNA) RNA剪接 胚胎发育 

分 类 号：Q291[生物学—细胞生物学]