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三重基序蛋白(the tripartite motif-containing protein,TRIM)家族是一类结构保守、进化快速的E3泛素蛋白连接酶,参与诸多重要的生命过程。TRIM36是TRIM家族的一员,trim36编码E3泛素蛋白连接酶。本研究在已完成的半滑舌鳎(Cynoglossus semilaevis)全基因组测序及转录组测序的基础上,利用RACE技术克隆得到半滑舌鳎trim36基因(Gen Bank登录号:KU244470),序列全长为2 899 bp,其中ORF为2 214 bp,编码737个氨基酸,5’非翻译区(untranslated regions,UTR)为289 bp,3’UTR为396 bp。生物信息学分析显示,半滑舌鳎TRIM36蛋白包含1个锌指结构,2个B-BOX类型锌指蛋白结构域(B-Box-type zinc finger,BBOX),1个卷曲螺旋结构,1个羧基末端的纤连蛋白Ⅲ(fibronectin typeⅢ,FN3)结构域和1个双特异性蛋白激酶spla和兰尼碱受体(spla kinase and ryanodine receptor,SPRY)结构域。此外,在不同脊椎动物中,trim36基因与其邻近的基因具有保守的共线性关系。半定量RT-PCR及q RT-PCR结果显示,在半滑舌鳎不同组织中,trim36基因表达存在差异。trim36在卵巢中表达量最高,其次为雌、雄鱼的脑中;不同发育时期中,trim36基因在120日龄半滑舌鳎性腺中开始表达,且卵巢表达量显著高于精巢(P<0.05)。原位杂交结果显示,trim36基因杂交信号主要出现在Ⅰ期和Ⅱ期卵母细胞。trim36时空表达特征表明,其在卵巢腔的形成及Ⅰ期、Ⅱ期卵母细胞的形成中起重要作用。结果表明,trim36在性别发生方面有一定的作用,这为进一步研究鱼类trim36基因功能及TRIM家族基因功能提供线索和参考。
The tripartite motif-containing protein (TRIM) family is a class of structurally conserved and rapidly evolving E3 ubiquitin protein ligases involved in many important life processes. TRIM36 is a member of the TRIM family and trim36 encodes E3 ubiquitin protein ligase. In this study, based on the complete genome sequencing and transcriptome sequencing of Cynoglossus semilaevis, the trim36 gene (Gen Bank accession number: KU244470) was cloned by RACE technology and the full length of the sequence was 2 899 bp. The ORF was 2 214 bp, encoding 737 amino acids. The 5 ’untranslated region (UTR) was 289 bp and the 3’ UTR was 396 bp. Bioinformatics analysis showed that the TRIM36 protein of the half-smooth tongue trumpet contains one zinc finger structure, two B-Box type zinc fingers (BBOX), one coiled-coil structure, one carboxy-terminal Fibronectin type Ⅲ (FN3) domain and a double-specific protein kinase spla and spla kinase and ryanodine receptor (SPRY) domain. In addition, trim36 has a conserved collinearity with its neighboring genes in different vertebrates. The results of semi-quantitative RT-PCR and q RT-PCR showed that the expression of trim36 gene was different in different tissues of the soleus. The expression of trim36 was the highest in ovary, followed by that in female and male. The trim36 gene began to express in the 120-day-old gonadal gonads at different developmental stages, and the ovarian expression was significantly higher than that in testis (P <0.05). In situ hybridization results showed that the signal of trim36 gene hybridization mainly appeared in stage I and stage II oocytes. The temporal and spatial expression of trim36 indicated that it plays an important role in ovarian cavity formation and stage I and stage II oocyte formation. The results showed that trim36 played a certain role in genital occurrence, which provided clues and reference for further study of trim36 gene function and TRIM family gene function.