大菱鲆MKK基因家族全基因组鉴定及其在生物和非生物应激下的表达分析

Genome-wide identification of the MKK gene family in Scophthalmus maximus and its involvement in abiotic and biotic stress responses

摘要:

为了阐明大菱鲆丝裂原活化蛋白激酶激酶 (mitogen-activated protein kinase kinases, MAPKKs或MKKs)基因家族在生物和非生物应激响应中的作用，本实验首先通过生物信息学方法对大菱鲆MKK基因家族进行了全基因组水平的鉴定，利用多个应激相关转录组数据集分析了大菱鲆MKK家族成员在不同组织及不同生物和非生物应激下的表达模式。结果显示，本研究在大菱鲆全基因组水平上共鉴定出9个MKK基因家族成员，它们不均匀地分布在7条染色体上，并分别对其编码蛋白的理化性质、蛋白二级结构和亚细胞定位进行了预测。基于系统发育分析，将SmMKKs划分为5个亚家族。内含-外显子结构、保守基序和多重序列比对分析结果不仅为大菱鲆MKK亚家族分类提供了证据，而且表明SmMKKs在进化上高度保守。SmMKKs在不同组织及不同生物和非生物应激下的基因表达模式分析表明，SmMKKs具有明显的组织特异性表达。另外，结果显示，粘孢子虫和肿大细胞病毒感染后，SmMKK6a呈极显著差异表达；热应激处理后，SmMKK6a呈极显著差异表达；高盐或低盐胁迫后，SmMKK4a、SmMKK4b、SmMKK6a和SmMKK7呈极显著差异表达。SmMKK6a在各种应激条件下均表现出极显著响应，表明其可能在综合抗应激中具有潜在的作用。这可能是第一个对大菱鲆MKK基因家族进行系统识别和功能分析的研究。以上研究结果不仅表明MKK基因家族在大菱鲆响应多种生物和非生物应激中发挥重要作用，而且也为大菱鲆综合抗逆分子选择育种研究提供了重要的理论支撑。

关键词:大菱鲆;丝裂原活化蛋白激酶激酶(MKK);基因家族;生物应激;非生物应激

Abstract:

Mitogen-activated protein kinase kinases (MAPKKs or MKKs) are essential components of the highly evolutionarily conserved mitogen-activated protein kinase (MAPK) cascade, which play crucial roles in response to a variety of biotic and abiotic stresses and immune responses. However, very little information is available about the MKKs in S. maximus. In order to detect the roles of the MKKs in response to biotic and abiotic stresses in Scophthalmus maximus, we first identified the MKKs of S. maximus at the whole genome level through bioinformatics methods, and then analyzed the expression patterns of MKKs in different tissues and under different biotic and abiotic stresses using multiple stress-related RNA-seq datasets. As a result, a total of 9 MKKs of S. maximus (SmMKKs) were identified, and they were unevenly distributed on 7 chromosomes. Physicochemical characteristics, secondary structure and subcellular localization of the proteins they encode were predicted, respectively. Phylogenetic analysis revealed that SmMKKs were classified into 5 subfamilies. Conserved motifs, intron-exon structure and multiple sequence alignment not only provided evidences for the classification of MKK subfamilies, but also revealed high levels of conservation in evolution within and between subfamilies. Expression patterns of SmMKKs in distinct tissues and under diverse abiotic and biotic stresses were examined using multiple published RNA-seq datasets. As a result, SmMKKs showed obviously tissue-specific expression. In addition, SmMKK6a was extremely significant differentially expressed after infection with both Enteromyxum scophthalmi and Megalocytivirus. After heat stress, SmMKK6a also showed extremely significant differential expression. Furthermore, SmMKK4a, SmMKK4b, SmMKK6a and SmMKK7 were extremely significant differentially expressed after high- or low- salinity stresses. Among these candidate stress-responsive MKK genes, SmMKK6a showed extremely significant response to both abiotic and biotic stresses, demonstrating its potential functions in comprehensive anti-stress. This study may be the first to systematically identify and analyze the MKK gene family in turbot. The results not only demonstrate that SmMKKs play crucial roles in response to various biotic and abiotic stresses, but also provide important theoretical support for the development of molecular selective breeding for comprehensive stress-resistance in S. maximus.

Key words:Scophthalmus maximus;mitogen-activated protein kinase kinases (MKK);gene family;biotic stress;abiotic stress