Push on through to the other side - molecular basis of viral cell-to-cell movement in plants

植物病毒是许多重要经济作物病害的罪魁祸首。在发展中国家，病毒造成的主要作物产量损失可能对当地人口造成直接和毁灭性的影响，但病毒性疾病也给工业化农业造成重大的经济损失。随着世界人口不断增长和气候变化给全球农业生产力带来越来越大的压力，保护和防御作物免受病毒侵害的能力成为实现可持续粮食安全的全球战略的重要组成部分。该项目将研究植物病毒生命周期中的一个关键步骤，其长期目标是开发具有更好病毒抗性的作物。由于植物细胞被纤维素细胞壁（造纸的原料）隔开，病毒将其感染传播到新的、未感染的宿主细胞的唯一途径是通过细胞壁上连接植物细胞的称为胞间连丝的极小的孔。细胞间的运动发生在感染的早期，因为病毒必须走在宿主的防御之前，只有少数病毒会在下一个细胞中发起感染。因此，这一步是中断感染周期以保护寄主植物的主要目标。植物病毒拥有专门的“运动蛋白”来介导这种细胞间运输。由于病毒比胞间连丝大，运动蛋白需要1)强行打开孔，2)推动或拉动它们的病毒“货物”通过孔。这两项任务中的前一项在一定程度上得到了理解，但对后一项却几乎一无所知。这个项目将测试一个新的假设，假设运动蛋白通过组装成细丝产生定向运输所需的机械力。单个蛋白质亚基聚合成细丝释放结合能，从而产生机械力并提供方向性。尽管总体上存在差异，但大多数植物病毒的运动蛋白形成某种聚合物或聚集体。这一间接证据表明，所提出的运动机制可能为大多数植物病毒所共有，因此由此产生的抗病毒策略将具有广泛的影响。该项目将通过在“试管”中重构运动蛋白聚合物，并详细分析细丝组装过程，对拟议的运动机制进行实验测试。这将涉及纯化运动蛋白，将它们与作为支架的脂质囊泡结合，并确定生化条件以及触发聚合所需的植物蛋白。复杂的光学和电子显微镜技术将被用来分析组装的动力学和聚合物的物理性质。如果基本假设得到证实，聚合所需的植物蛋白构成了使病毒感染的易感因子。对这些易感因子的了解将有助于通过筛选种子库和种质库中不支持病毒转运的变异形式，有针对性地培育更具抗病毒性的作物。

Plant viruses are responsible for many diseases of economically important crops. In the developing world, virally caused yield losses of staple crops can have direct and devastating effects for the local population, but viral diseases also cause substantial financial losses in industrialised agriculture. As the growing world population and climate change put increasing pressure on world-wide agricultural productivity, the ability to protect and defend crop plants from viruses forms an important part of global strategies towards sustainable food security. This project will investigate a crucial step in the life cycle of plant viruses, with the long-term aim of developing crops with improved virus resistance.Because plant cells are separated by a cellulose cell wall (the source for making paper), the only pathway for viruses to spread their infection into new, uninfected host cells is through extremely small pores called plasmodesmata that connect plant cells across the cell wall. Cell-to-cell movement happens early during the infection as the virus has to stay ahead of host defences, and only a handful of viruses initiate infection in the next cell. Therefore this step is a prime target for interupting the infection cycle to protect the host plant. Plant viruses possess specialised "movement proteins" that mediate this intercellular transport. Because the viruses are large compared to the size of plasmodesmata, movement poteins need to 1) force open the pores and 2) push or pull their viral 'cargo' through them. The former of these two tasks is understood to some degree, but virtually nothing is yet known about the latter.This project will test a new hypothesis which posits that movement proteins generate the mechanical force required for directional transport through plasmodesmata by assembling into filaments. Polymerisation of individual protein subunits into filaments releases binding energy and can thus generate mechanical force and also provide directionality. Despite overall differences, the movement proteins of most plant viruses form some sort of polymer or aggregates. This circumstancial evidence suggests that the proposed movement mechanism may be shared by the majority of plant viruses, so that resulting antiviral strategies will have a wide impact.The project will test the proposed movement mechanism experimentally by reconstituting movement protein polymers in a 'test tube', and analysing the filament assembly process in detail. This will involve purifying movement proteins, combining them with lipid vesicles that serve as scaffolds, and identifying biochemical conditions as well as plant proteins that are required to trigger polymerisation. Sophisticated light and electron microscopy techniques will be employed to analyse the kinetics of the assembly and the physical properties of the polymers. If the underlying hypothesis is verified, plant proteins required for polymerisation constitute susceptibility factors that enable viral infection. Knowledge of these susceptibility factors will enable targeted breeding of more virus-resistant crops by screening seed banks and germplasm libraries for variant forms that do not support virus transport.