Abstract：
The exosomal miRNA plays a crucial role in the intercellular communication response to environmental stress and pathogenic stimulation. In the present study, the expression of exosomal miRNAs in the Pacific oyster Crassostrea gigas after high-temperature stress or Vibrio splendidus stimulation was investigated through high-throughput sequencing. The exosomes were identified to be teardrop-like vesicles with the average size of 81.7 nm by transmission electron microscopy. There were 66 known miRNAs and 33 novel miRNAs identified, of which 10 miRNAs were differentially expressed after both high-temperature stress and Vibrio stimulation compared to the control group. A total of 1868 genes were predicted as the putative targets of miRNAs, of which threonine aspartase 1-like was targeted by the highest number of related miRNAs. The robustness and reliability of miRNA expression from the sRNA sequencing data were verified by employing eight miRNAs for qPCR. GO and KEGG clustering analyses revealed that apoptosis was significantly enriched by the target genes of differentially expressed exosomal miRNAs after high-temperature stress, and autophagy and cytokine activity were significantly enriched after Vibrio stimulation. Energy metabolism was found to be significantly shared in the target gene enrichments after both high-temperature stress and Vibrio stimulation. These findings would improve our understanding of the regulatory mechanisms of exosomal miRNAs in C. gigas after high-temperature stress or Vibrio stimulation.
摘要：
外泌体miRNA在对环境应激和致病刺激的细胞间通讯反应中起着至关重要的作用。在本研究中,通过高通量测序对高温胁迫或脾弧菌刺激后太平洋牡蛎中外泌体miRNAs的表达进行了表征。通过透射电子显微镜鉴定外泌体为平均尺寸为81.7nm的泪滴状囊泡。有66个已知的miRNA和33个新的miRNA鉴定,其中10个miRNA在高温胁迫和弧菌刺激后与对照组相比差异表达。总共有1868个基因被预测为miRNA的推定靶标，其中苏氨酸天冬氨酸酶1样被最多数量的相关miRNA靶向。通过采用8种miRNA进行qPCR,验证了来自sRNA测序数据的miRNA表达的稳健性和可靠性。GO和KEGG聚类分析显示，高温胁迫后差异表达的外泌体miRNAs的靶基因显著富集了凋亡，弧菌刺激后细胞自噬和细胞因子活性显著富集。发现在高温胁迫和弧菌刺激后，靶基因富集中的能量代谢显着共享。这些发现将提高我们对高温胁迫或弧菌刺激后C.gigas中外泌体miRNA调控机制的理解。
