Studies on the novel 5MP-ATF-4 pathway using insects as the model system

分子生物学的中心教条指出，当基因被激活时，DNA会产生制造蛋白质的RNA。目前对基因如何开启和关闭的理解集中在这些步骤的第一步：改变DNA基因组不同区域产生的RNA的数量(转录控制)。通过改变RNA上合成的蛋白质的数量来进行翻译控制的作用还不是很清楚。这个项目将研究控制RNA上蛋白质合成的第一步的细胞机制。研究人员将研究这种翻译起始机制某些部分的突变是如何损害神经细胞的生长并导致动物，特别是昆虫的早期死亡的。尤卡里亚特有的翻译因子复合体之一是由四个真核起始因子eIF1、eIF2、eIF3、eIF5和Met-tRNAiMet组成的多起始因子复合体(MFC)。它在8月密码子的严格翻译启动过程中起着至关重要的作用。人eIF5模拟蛋白1(5MP1)通过模拟eIF5的C末端结构域结合eIF2和eIF3，并抑制翻译。因此，这是第一个被确定为MFC功能的潜在调节蛋白。MFC调节(eIF2抑制)的一个重要结果是上调ATF4的翻译，ATF4是控制氨基酸合成和抗逆性的转录因子。在本项目中，将利用培养的昆虫细胞系，包括神经细胞系，研究5MP与翻译机制的相互作用以及5MP对ATF4翻译的调控。在果蝇中，遗传证据表明5MP参与轴突引导。利用第二种昆虫模型生物赤粉甲虫，旨在检测5MP或ATF4基因敲除是否导致轴突引导缺陷，并确定5MP的哪个蛋白质伙伴可以调节这一生物学过程，以及这是否通过ATF4完成。

The central dogma of molecular biology states that when genes are turned on, DNA makes RNA that makes proteins. The current understanding of how genes are turned on and off centers on the first of these steps: changing the amount of RNA produced from various regions of DNA genome (transcriptional control). The role of translational control through changing the amount of proteins made on RNA is less understood. This project will investigate the cell machinery that controls the first steps of protein synthesis on RNA. The researchers will investigate how mutations in some parts of this translation initiation machinery impair growth of neural cells and cause early death in animals, in particular insects.One of the Eukarya-specific translation factor complex is the multi-initiation factor complex (MFC) made of four eukaryotic initiation factors, eIF1, eIF2, eIF3, eIF5 and Met-tRNAiMet. It plays essential roles in stringent translation initiation at AUG codon. Human eIF5-mimic protein 1 (5MP1) binds eIF2 and eIF3 through eIF5-mimic C-terminal domain and inhibits translation. Thus, this is the first protein that was identified as the potential regulator of the MFC function. An important consequence of MFC regulation (eIF2 inhibition) is to up-regulate the translation of ATF4, the transcription factor governing amino acid synthesis and stress resistance. In this project, cultured insect cell lines, including neuronal cell lines, will be used to study 5MP interactions with translational machineries and 5MP regulation of ATF4 translation. In the fruit fly, genetic evidence suggests that 5MP is involved in axon guidance. Using the second insect model organism Tribolium castaneum (red flour beetle), it is aimed at examining whether 5MP or ATF4 knockdown causes defects in axon guidance and determining which protein partner of 5MP can regulate this biological process and whether this is done through ATF4.