Feng Eryan, Chen Huan, Li Yan, Jiang Wei, Wang Zhongkang, Yin Youping
School of Life Science, Chongqing University, Chongqing Engineering Research Center for Fungal Insecticide, 400030, China.
Institute of Plant Physiology and Ecology, Chinese Academy of Sciences Key Laboratory of Insect Developmental and Evolutionary Biology, CAS, Shanghai, 200032, China.
Comp Biochem Physiol B Biochem Mol Biol. 2014 Jun-Jul;172-173:49-56. doi: 10.1016/j.cbpb.2014.04.003. Epub 2014 Apr 16.
The 14-3-3 proteins, a highly evolutionarily conserved and ubiquitous protein family in eukaryotic cells, have a range of biological functions including regulation of signal transduction, stress response, apoptosis, and control of the cell cycle. To investigate the function of 14-3-3 in Spodoptera litura, the full length of 14-3-3ζ was cloned from S. litura on the basis of an expressed sequence tag of 14-3-3ζ from the S. litura fat body suppression subtractive hybridization library, and named SpL14-3-3ζ. SpL14-3-3ζ cDNA was 1196 bp with an open reading frame of 744 bp, encoding 247 amino acids. Multiple alignment analysis revealed the putative amino acids shared >80% homology with 14-3-3ζ from other organisms and shared typical conservative structures. Phylogenetic analysis confirmed SpL14-3-3ζ was closely related to other available Lepidoptera 14-3-3ζ. Real-time PCR analysis indicated SpL14-3-3ζ was expressed throughout the developmental stages of S. litura, with a relatively high expression level in pre-pupa, and was expressed constitutively in all examined tissues with relatively high levels in hemocytes and midgut. Moreover, the transcription level of SpL14-3-3ζ could be induced by Nomuraea rileyi infection, up-regulated in hemocytes, followed by head, fat body and midgut. Knocking down SpL14-3-3ζ transcripts by RNAi significantly increased S. litura sensitivity to fungal infection, and resulted in higher mortality of S. litura during the larval development. These results provide novel insights into the 14-3-3ζ signal regulation which may be related to host defense as well as larval development in S. litura.
14-3-3蛋白是真核细胞中高度进化保守且普遍存在的蛋白家族，具有一系列生物学功能，包括信号转导调节、应激反应、细胞凋亡以及细胞周期控制。为研究14-3-3在斜纹夜蛾中的功能，基于斜纹夜蛾脂肪体抑制性消减杂交文库中14-3-3ζ的表达序列标签，从斜纹夜蛾中克隆了14-3-3ζ的全长，并命名为SpL14-3-3ζ。SpL14-3-3ζ cDNA为1196 bp，开放阅读框为744 bp，编码247个氨基酸。多序列比对分析显示，推测的氨基酸与其他生物的14-3-3ζ具有>80%的同源性，并具有典型的保守结构。系统发育分析证实SpL14-3-3ζ与其他已有的鳞翅目14-3-3ζ密切相关。实时荧光定量PCR分析表明，SpL14-3-3ζ在斜纹夜蛾的整个发育阶段均有表达，在预蛹期表达水平相对较高，在所有检测组织中组成性表达，在血细胞和中肠中表达水平相对较高。此外，莱氏野村菌感染可诱导SpL14-3-3ζ的转录水平，在血细胞中上调，其次是头部、脂肪体和中肠。通过RNA干扰敲低SpL14-3-3ζ转录本显著增加了斜纹夜蛾对真菌感染的敏感性，并导致斜纹夜蛾幼虫发育期间的死亡率更高。这些结果为14-3-3ζ信号调节提供了新的见解，这可能与斜纹夜蛾的宿主防御以及幼虫发育有关。
