Dai Li, Liu Xue-Chen, Ye Sen, Li Hua-Wei, Chen Dian-Fu, Yu Xiao-Jian, Huang Xue-Ting, Zhang Li, Yang Fan, Yang Jin-Shu, Yang Wei-Jun
a Institute of Cell and Developmental Biology, College of Life Sciences , Zhejiang University , Hangzhou , Zhejiang , China.
b Institute of Basic Research in Clinical Medicine, School of Basic Medical Sciences , Zhejiang Chinese Medical University , Hangzhou , Zhejiang , China.
Stress. 2016 Nov;19(6):609-620. doi: 10.1080/10253890.2016.1244523. Epub 2016 Oct 26.
The most widespread type of RNA editing, conversion of adenosine to inosine (A→I), is catalyzed by two members of the adenosine deaminase acting on RNA (ADAR) family, ADAR1 and ADAR2. These enzymes edit transcripts for neurotransmitter receptors and ion channels during adaption to changes in the physical environment. In the primitive crustacean Artemia, when maternal adults are exposed to unfavorable conditions, they release diapause embryos to withstand harsh environments. The aim of the current study was therefore to elucidate the role of ADAR of Artemia diapause embryos in resistance to stress. Here, we identified Artemia ADAR (Ar-ADAR), which harbors a putative nuclear localization sequence (NLS) and two double-stranded RNA-binding motifs (dsRBMs) in the amino-terminal region and an adenosine deaminase (AD) domain in the carboxyl-terminal region. Western blot and immunofluorescence analysis revealed that Ar-ADAR is expressed abundantly in post-diapause embryos. Artemia (n = 200, three replicates) were tested under basal and stress conditions. We found that Ar-ADAR was significantly induced in response to the stresses of salinity and heat-shock. Furthermore, in vivo knockdown of Ar-ADAR (n = 100, three replicates) by RNA interference induced formation of pseudo-diapause embryos, which lack resistance to the stresses and exhibit high levels of apoptosis. These results indicate that Ar-ADAR contributes to resistance to stress in Artemia diapause embryos.
最普遍的RNA编辑类型，即腺苷向肌苷的转化（A→I），由作用于RNA的腺苷脱氨酶（ADAR）家族的两个成员ADAR1和ADAR2催化。这些酶在适应物理环境变化的过程中编辑神经递质受体和离子通道的转录本。在原始甲壳类动物卤虫中，当母体成虫暴露于不利条件时，它们会释放滞育胚胎以抵御恶劣环境。因此，本研究的目的是阐明卤虫滞育胚胎的ADAR在抗逆中的作用。在这里，我们鉴定了卤虫ADAR（Ar-ADAR），其在氨基末端区域含有一个推定的核定位序列（NLS）和两个双链RNA结合基序（dsRBMs），在羧基末端区域含有一个腺苷脱氨酶（AD）结构域。蛋白质免疫印迹和免疫荧光分析表明，Ar-ADAR在滞育后胚胎中大量表达。对卤虫（n = 200，三个重复）在基础条件和应激条件下进行了测试。我们发现，Ar-ADAR在盐度和热休克应激下显著诱导。此外，通过RNA干扰在体内敲低Ar-ADAR（n = 100，三个重复）诱导了假滞育胚胎的形成，这些胚胎缺乏抗逆性并表现出高水平的细胞凋亡。这些结果表明，Ar-ADAR有助于卤虫滞育胚胎的抗逆性。
