本文目录一览：
求翻译！！
SIGNIFICANCE
Drought is a major cause of lost agricultural productivity. Even moderate water limitation can lead to down-regulation of plant growth; however, the underlying mechanisms of stress sensing and growth regulation are little understood. We identified At14a-Like1 (AFL1) and its interacting proteins protein disulfide isomerase 5 (PDI5) and NAI2 as positive and negative regulators, respectively, of growth and proline accumulation. Despite numerous ideas that membrane-based mechanisms are important for drought sensing and initial signaling, AFL1 is one of only a few membrane proteins with a demonstrated effect on drought resistance. AFL1 structure, localization, and interaction with endomembrane proteins indicate novel functions in drought signaling. Increased growth of AFL1 overexpression in plants under stress without negative effects on unstressed plants make AFL1 an attractive target for biotechnology.
Keywords: drought, At14a, vesicle endocytosis, protein disulfide isomerase, clathrin adaptor AP2-2a
意义
干旱是农业生产力下降的主要原因。即使是适度的水分限制也会导致植物生长下调;然而，压力感知和生长调节的潜在机制很少被理解。我们鉴定了At14a-Like1（AFL1）及其相互作用蛋白质蛋白质二硫键异构酶5（PDI5）和NAI2分别作为生长和脯氨酸积累的正调节剂和负调节剂。尽管有许多观点认为基于膜的机制对干旱传感和初始信号传导很重要，但AFL1是少数几种对抗旱性有显着作用的膜蛋白之一。 AFL1结构，定位和与内膜蛋白的相互作用表明干旱信号传导中的新功能。在压力下植物中AFL1过表达的增加，对无应激植物没有负面影响，使AFL1成为生物技术的一个有吸引力的目标。
关键词：干旱，At14a，囊泡内吞作用，蛋白质二硫键异构酶，网格蛋白衔接子AP2-2a
ABSTRACT
Limited knowledge of how plants regulate their growth and metabolism in response to drought and reduced soil water potential has impeded efforts to improve stress tolerance. Increased expression of the membrane-associated protein At14a-like1 (AFL1) led to increased growth and accumulation of the osmoprotective solute proline without negative effects on unstressed plants. Conversely, inducible RNA-interference suppression of AFL1 decreased growth and proline accumulation during low water potential while having no effect on unstressed plants. AFL1 overexpression lines had reduced expression of many stress-responsive genes, suggesting AFL1 may promote growth in part by suppression of negative regulatory genes. AFL1 interacted with the endomembrane proteins protein disulfide isomerase 5 (PDI5) and NAI2, with the PDI5 interaction being particularly increased by stress. PDI5 and NAI2 are negative regulatory factors, as pdi5, nai2, and pdi5-2nai2-3 mutants had increased growth and proline accumulation at low water potential. AFL1 also interacted with Adaptor protein2-2A (AP2-2A), which is part of a complex that recruits cargo proteins and promotes assembly of clathrin-coated vesicles. AFL1 colocalization with clathrin light chain along the plasma membrane, together with predictions of AFL1 structure, were consistent with a role in vesicle formation or trafficking. Fractionation experiments indicated that AFL1 is a peripheral membrane protein associated with both plasma membrane and endomembranes. These data identify classes of proteins (AFL1, PDI5, and NAI2) not previously known to be involved in drought signaling. AFL1-predicted structure, protein interactions, and localization all indicate its involvement in previously uncharacterized membrane-associated drought sensing or signaling mechanisms.
Even relatively mild drought that causes reduced soil water potential (ψw) can result in dramatically reduced plant growth and agricultural productivity. Physiological analyses have shown that plant growth is actively down-regulated during drought and is not limited by carbon supply (1–3). Reductions of growth help ensure survival by conserving water but can be undesirable for agriculture, as plant productivity is reduced more than need be if growth were less sensitive to changes in water status (3). Also, specific metabolic pathways, such as proline metabolism, are stress regulated and contribute to drought tolerance.
The sensing and signaling mechanisms controlling growth and metabolic responses to drought remain unclear. Many hypotheses of how plants sense water loss center on detection of mechanical stimuli generated by loss of turgor and cell shrinkage. This includes changes in membrane shape or disruption of cell wall–cell membrane connections possibly detected by proteins, such as mechanosensitive channels or receptor-like kinases that bind cell wall components (4–9). Proteins that induce or detect membrane curvature are known in mammalian cells (10) but have been little considered in plants. Also, in analogy to mammalian cells, integrin-related proteins have been hypothesized to play stress-sensing roles in plant cells. Plants lack clear orthologs to integrins. Nonetheless, modeling has identified at least one Arabidopsis protein with integrin-like structure and possible stress-related function (11), and other proteins with small integrin similarity domains also have been identified (12). Endomembrane compartments, particularly the endoplasmic reticulum (ER), are involved in responding to cytotoxic stresses such as the accumulation of unfolded proteins (13). How endomembrane proteins may be involved in responding to water limitation, and whether this may occur via mechanisms other than the unfolded protein response, is less understood. Trafficking of membrane proteins between cellular compartments is also emerging as an important aspect of plant signaling, with plasma membrane (PM) aquaporins being one example of intracellular trafficking affecting drought resistance (14). Sites of ER–PM contact have also been proposed to be critical for mechanosensing and stress tolerance (15).
With the motivation of testing a different class of protein that could have roles in sensing or signaling abiotic stress, we investigated the function of At14a-Like1 (AFL1, At3g28270). At14a (At3g28300) was first identified by immunoscreening an Arabidopsis expression library with antisera recognizing mammalian β1-integrin and was reported to be a PM-associated protein (12). A cluster of At14a-related genes, including AFL1, is present in Arabidopsis. AFL1 contains a small domain with similarity to integrins (domain of unknown function 677), but there is little other information that could reveal its cellular function. Our investigation found that AFL1 has a dramatic effect on plant growth during drought and identified AFL1 association with endomembrane proteins and clathrin-coated vesicle formation at the PM as key aspects of AFL1 cellular function.
抽象
关于植物如何响应干旱和降低土壤水势来调节其生长和代谢的知识有限，阻碍了改善胁迫耐受性的努力。膜相关蛋白At14a-like1（AFL1）的表达增加导致渗透保护性溶质脯氨酸的生长和积累增加而对无应激植物没有负面影响。相反，AFL1的诱导型RNA干扰抑制在低水势期间降低了生长和脯氨酸积累，而对无应激植物没有影响。 AFL1过表达株系降低了许多应激反应基因的表达，表明AFL1可能通过抑制负调控基因促进生长。 AFL1与内膜蛋白蛋白质二硫键异构酶5（PDI5）和NAI2相互作用，PDI5相互作用通过应激特别增加。 PDI5和NAI2是负调节因子，因为pdi5，nai2和pdi5-2nai2-3突变体在低水势下具有增加的生长和脯氨酸积累。 AFL1还与衔接蛋白2-2A（AP2-2A）相互作用，后者是招募货物蛋白并促进网格蛋白包被囊泡组装的复合物的一部分。 AFL1与质膜上的网格蛋白轻链共定位，以及AFL1结构的预测，与囊泡形成或运输中的作用一致。分级实验表明AFL1是与质膜和内膜相关的外周膜蛋白。这些数据识别以前未知参与干旱信号传导的蛋白质类别（AFL1，PDI5和NAI2）。 AFL1预测的结构，蛋白质相互作用和定位均表明其参与了之前未表征的膜相关干旱传感或信号传导机制。
即使相对温和的干旱导致土壤水势降低（ψw）也会导致植物生长和农业生产力大幅下降。生理分析表明，在干旱期间植物生长受到积极的下调，并且不受碳供应的限制（1-3）。减少生长有助于通过节约水来确保生存，但对农业来说可能是不合需要的，因为如果生长对水的状况变化不太敏感，植物生产力会降低（3）。此外，特定的代谢途径，例如脯氨酸代谢，受到压力调节并有助于耐旱性。
控制生长和对干旱的代谢反应的传感和信号传导机制仍不清楚。关于植物如何感知水分流失的许多假设都集中在检测由于膨胀和细胞萎缩造成的机械刺激。这包括膜形状的变化或可能由蛋白质检测到的细胞壁 - 细胞膜连接的破坏，例如机械敏感性通道或结合细胞壁组分的受体样激酶（4-9）。诱导或检测膜弯曲的蛋白质在哺乳动物细胞中是已知的（10），但在植物中很少考虑。此外，与哺乳动物细胞类似，假设整联蛋白相关蛋白在植物细胞中发挥应激感应作用。植物缺乏整合素的明确直向同源物。尽管如此，建模已经鉴定出至少一种具有整合素样结构和可能的应激相关功能的拟南芥蛋白（11），并且已经鉴定了具有小整联蛋白相似结构域的其他蛋白（12）。内膜隔室，特别是内质网（ER），参与细胞毒性应激的响应，例如未折叠蛋白的积累（13）。内膜蛋白如何参与响应水限制，以及这是否可能通过除了未折叠的蛋白质反应之外的机制发生，还不太了解。在细胞区室之间运输膜蛋白也是植物信号传导的一个重要方面，质膜（PM）水通道蛋白是影响抗旱性的细胞内运输的一个例子（14）。 ER-PM接触点也被认为对机械传感和应力耐受至关重要（15）。
为了测试可能在感知或发出非生物应激信号中起作用的不同类别蛋白质的动机，我们研究了At14a-Like1（AFL1，At3g28270）的功能。 At14a（At3g28300）首先通过用识别哺乳动物β1整联蛋白的抗血清免疫筛选拟南芥表达文库来鉴定，并且据报道是PM相关蛋白（12）。拟南芥中存在At14a相关基因簇，包括AFL1。 AFL1包含一个与整合素相似的小域（未知功能域677），但几乎没有其他信息可以揭示其细胞功能。我们的研究发现，AFL1对干旱期间的植物生长具有显着影响，并确定AFL1与内膜蛋白的结合以及PM处的网格蛋白包被的囊泡形成是A的关键方面。
AFL Fuzzer如何在LAVA-M数据集上进行测试？
不要因追求完美而为自己设置可望而不可及的目标，其实幸福很简单，就像有一所大房子，里面想要的东西一样都不缺，多余的东西一样也没有。敲响的是钟声，走过的是岁月，种下的是希望，留下的是故事，盼望的是美好，送来的...
AFL装置由三哪组成
A.快速动作的基本级
B.带延时的附加级
C.延时级
三部分
AFL指按频率自动减负荷装置。为了提高电能质量，保证重要用户供电的可*性，当系统中出现有功功率缺额引起频率下降时，根据频率下降的程度，自动断开一部分不重要的用户，阻止频率下降，以便使频率迅速恢复到正常值，这种装置叫按频率自动减负荷装置，简称AFL装置。它不仅可以保证重要用户的供电，而且可以避免频率下降引起的系统瓦解事故。继电保护测试仪中AFL装置基本工作原理 自动按频率负荷装置的工作原理,可用下图说明,假定变电所馈电母线上有多条供配电线路,俺店里用户的重要性分为n个级别