玉米AKINβγ蛋白在水稻黑条矮缩病毒侵染过程中的作用研究

Maize rough dwarf caused by Rice black streaked dwarf virus (RBSDV) is a widespread and destructive viral disease in China. It is essential to elucidate the molecular mechanism underlying the interactions between RBSDV and maize, for revealing the mechanisms of symptom development or host resistance to viral infection. We have confirmed the interaction between ZmAKINβr and P8, the minor core protein of RBSDV, in vitro and in vivo. Furthermore, our preliminary results suggested that the subcellular localization of ZmAKINβγ and the accumulation of ZmAKINβγ mRNAs in maize could be affected by RBSDV infection. AKINβγ is a subunit of SnRK1 complex which regulates the carbohydrate metabolism, sugar signaling and the responses to biotic or abiotic stresses. We will analyze the effects of RBSDV infection, on the subcellular localizations and functions of P8 and ZmAKINβγ, respectively. Moreover, the transcription level of ZmAKINβγ will be manipulated through virus-induced gene silencing (VIGS) and stable overexpression assays, with the purpose of revealing the roles of ZmAKINβγ during maize developmental processes or upon RBSDV infection. A series of experiments will be performed to elucidate the mechanisms underlying the influences of ZmAKINβγ on RBSDV infection or symptoms development. Eventually these efforts could contribute to the effective control of maize rough dwarf disease by precisely regulating specific infection-responsive host genes.

水稻黑条矮缩病毒（Rice black streaked dwarf virus, RBSDV）引起的玉米粗缩病是我国玉米上的主要病害。探索RBSDV与玉米互作在病害发生过程中的分子机制具有重要的科学意义。我们在前期鉴定出了与RBSDV结构蛋白P8互作的玉米蛋白ZmAKINβγ，初步证明RBSDV侵染可影响ZmAKINβγ的亚细胞定位及ZmAKINβγ基因的表达。AKINβγ是激酶复合体SnRK1的组分，参与碳代谢和糖信号途径，可调控植物生长发育并响应非生物和生物胁迫。我们拟研究P8与ZmAKINβγ蛋白直接互作对二者的亚细胞定位和生物学功能的影响；利用病毒诱导的基因沉默（VIGS）和过量表达等方法明确ZmAKINβγ在玉米生长发育和RBSDV侵染过程中的作用，解析ZmAKINβγ影响病毒侵染或病害症状形成的分子机制和生理途径，为调控玉米相关基因的表达、培育抗玉米粗缩病新种质提供理论依据。

水稻黑条矮缩病毒 (RBSDV)侵染玉米导致粗缩病。RBSDV的副核心蛋白P8与玉米AKINβΥ蛋白发生互作。AKINβΥ是SnRK1复合体的Υ亚基，参与糖代谢。本研究证实P8与ZmAKINβΥ-2及ZmAKINβΥ-1在玉米细胞中发生互作。P8与ZmAKINβΥ的互作依赖于后者N端的KIS域和首个 CBS结构域。P8 与 ZmAKINβΥ-1/-2 均定位于玉米细胞质与细胞核中, 并且P8 可在核仁中聚集。ZmAKINβΥ-1/-2可互补酵母突变体snf4∆的功能, 具有Υ亚基活性；P8能够抑制ZmAKINβΥ-1/-2作为Υ亚基的功能。RBSDV 侵染后能够差异化地调控 ZmAKINβΥ-1/-2 基因的表达，并改变ZmAKINβΥ-1/-2在玉米叶片中的亚细胞定位。利用BMV-VIGS系统同时沉默ZmAKINβΥ-1和 ZmAKINβΥ-2基因对玉米的生长发育没有明显影响，但却调控了玉米的蔗糖和淀粉代谢途径相关基因的表达,导致葡萄糖和蔗糖的积累量增加, 但对淀粉的积累量却没有显著影响。用BMV-VIGS同时沉默ZmAKINβΥ-1/-2后，立即用RBSDV挑战接种玉米植株，促进了RBSDV的积累。RBSDV侵染可调控糖代谢途径相关基因的表达, 导致葡萄糖含量增加而蔗糖含量下降；外施葡萄糖能够促进RBSDV的积累。因此，ZmAKINβΥ-1和ZmAKINβΥ/-2的表达影响了葡萄糖的含量，进而影响了RBSDV的积累。此外，我们用另外两种玉米病毒探索ZmAKINβΥ对不同病毒侵染的反应。用玉米褪绿斑驳病毒（MCMV，番茄丛矮病毒科)接种玉米植株后第1片系统侵染叶中ZmAKINβΥ的表达水平均先显著上调，然后显著下调。用甘蔗花叶病毒（SCMV)接种玉米植株后的第1片系统侵染叶中两种ZmAKINβΥ的表达均显著下调。因此，ZmAKINβΥ可能是抗病途径中的关键调节因子。本研究表明 ZmAKINβΥ-1/-2参与玉米的糖代谢, 是普遍响应病毒侵染的寄主蛋白, 在 RBSDV 侵染玉米过程中与P8蛋白互作并作为抵抗RBSDV侵染的正调节因子发挥作用。在研究与P8互作的玉米因子的过程中，我们发现了对RBSDV侵染有重要作用的两类蛋白，即脂滴蛋白与细胞数目调控因子。我们将研究ZmAKINβΥ在玉米抗病毒侵染中的作用机制，期望通过基因编辑调控相关基因的表达并控制病毒的侵染。