All plant‐interacting microbes must acquire metabolites from their hosts to satisfy nutritional demands for growth. With carbon being crucial for all organisms, sufficient acquisition of sugars from plants is a cardinal task of plant pathogens for successful invasion. Blocking access to host sugars seems to be a promising strategy to control plant diseases. Plant sugar retrieval strengthens plant resistance to pathogens (Yamada et al., 2016). However, it is difficult to discriminate if this is a result of blocking the pathogen’s access to sugar, or a disturbance in sugar‐mediated signalling in plants (Milne et al., 2019; Moore et al., 2015). Since the identification of UfHXT1 provided the first evidence of sugar uptake in rust fungi (Voegele et al., 2001), many sugar transporters have been identified from different pathogenic fungi (Saitoh et al., 2014; Schuler et al., 2015). However, the effects of sugar starvation on pathogen growth, development and pathogenicity are still unclear.

Puccinia striiformis f.sp. tritici (Pst) is the causal agent of stripe (yellow) rust, which threatens global wheat production severely. So far, control of Pst mostly relies on the deployment of resistant cultivars carrying specific resistance (R) genes, and the use of chemical fungicides. However, novel, sustainable ways to control Pst are desperately needed. Recently, hexoses were shown to be the major form of sugars utilized by this obligate biotrophic fungus (Chang et al., 2017). In this study, we cloned the hexose transporter gene PsHXT1, which is the only one highly induced during Pst infection (Zheng et al., 2013). Further analysis of PsHXT1/PsHXT1 revealed typical characteristics of a major facilitator superfamily (MFS) symporter with 12 membrane‐spanning segments (Figure 1a). Intraspecies polymorphism of PsHXT1 seems to be fairly low, as all eleven compared Pst genomes show a similarity of greater 99% at the nucleotide level (Figure 1b). While the interspecies variation ranges between 83% and 91% among closely related species, PsHXT1 is clearly different from other rust fungal glucose transporters characterized so far. It only shares 26% similarity with UfHXT1 (Figure 1c). As genes involved in sugar acquisition are much more conserved compared with effectors (Oliva and Quibod, 2017), these genes/proteins might represent promising targets for novel ways to control plant diseases.

Transcript levels of PsHXT1 during Pst infection were analysed by qRT‐PCR for the complete invasion process (Figure 1d). Transcript levels of PsHXT1 increased from 12 h post‐inoculation (hpi), when primary infection starts with substomatal vesicle formation, and increased continuously to reach a maximum at 168 hpi, when branched hyphae develop and more haustoria are formed. Thereafter, transcript levels sharply decrease to a very low level. This result indicates that PsHXT1 is indispensable for establishing the Pst–wheat interaction.

In order to determine the subcellular localization of PsHXT1, a PsHXT1‐GFP fusion protein was generated and expressed in yeast. PsHXT1 was shown to localize to the plasma membrane (Figure 1e). The subcellular localization of PsHXT1 was further analysed by expression in Nicotiana benthamiana (Figure 1f). Both plasmolysis and staining with the membrane marker SynaptoRed™ C2 (FM4‐64) confirmed a plasma membrane localization of PsHXT1. Based on a similar subcellular localization, PsHXT1 could function as a transporter as UfHXT1 (Voegele et al., 2001).

In order to identify the biochemical characteristics of PsHXT1, PsHXT1 was expressed in the Saccharomyces cerevisiae mutant strain EBY.VW4000, which lacks all 20 hexose transporters identified. PsHXT1 was shown to exhibit a substrate preference of glucose (Figure 1g). The Km of PsHXT1 was 59 ± 12 μm, and the Vmax was 7.75 ± 2.33 nm under optimal conditions (Figure 1h). The optimum pH of PsHXT1 is about 5.5, but transport activity retained a high level within the pH range from 4 to 7. Two different protonophores, carbonyl cyanide m‐chlorophenylhydrazone (CCCP) and 2,4‐dinitrophenol (DNP), were both able to inhibit the activity of PsHXT1. The SH group inhibitor, p‐chloromercuribenzene sulphonate (pCMBS), had no effect on PsHXT1 activity. Competition experiments confirmed that PsHXT1 has a high affinity for glucose only. All these results indicate that PsHXT1 is a glucose–proton symporter.

In order to determine the biological function of PsHXT1 in a Pst–wheat interaction, PsHXT1 was silenced by barley stripe mosaic virus (BSMV)‐mediated host‐induced gene silencing (HIGS). Two independent fragments (PsHXT1‐1as and PsHXT1‐2as) were chosen to silence PsHXT1, and PsINVas served as a positive control (Chang et al., 2017). Disease phenotypes of Pst infection were observed for 14 days. Disease phenotypes decreased on plants treated with either BSMV:PsHXT1‐1as or BSMV:PsHXT1‐2as (Figure 1i). Statistical analysis of the quantity of uredia on infected leaves further supports the differences in disease phenotypes. In addition, the biomass ratio indicates that the biomass of Pst in leaves treated with either BSMV:PsHXT1‐1as or BSMV:PsHXT1‐2as decreased significantly compared with leaves treated with BSMV:00. Development and growth of Pst were examined by histological observation in PsHXT1‐silenced plants (Figure 1j). At 24 hpi, Pst formed more branches, and inflated substomatal vesicles could be observed in nearly 30% of the cases. This indicates that there might be problems with the establishment of the Pst–wheat interaction with PsHXT1‐silenced plants. At 48 hpi, hyphae showed abnormal development and exhibited high levels of malformation (in nearly 70% of the cases). At 120 hpi, the infection area of Pst was significantly decreased in PsHXT1‐silenced plants. Taken together, these results indicate that silencing PsHXT1 restricts normal growth and development of Pst during the infection of wheat significantly, leading to a decrease in fungal biomass and disease symptoms.

Combined with the former study on PsINV (Chang et al., 2017), it can be concluded that sugar starvation not only impairs growth and development of Pst, but also slows down pathogen proliferation. To our knowledge, this is the first in vivo evidence demonstrating that sugar starvation restricts both pathogen’s growth and virulence without a possible confusion with signalling effects. This opens new vistas for sugar starvation‐mediated control of wheat stripe rust and suggests that blocking a pathogen’s sugar absorption could be a novel strategy to control disease with restricting pathogen’s growth and proliferation. Although most attention has been paid into seeking effectors and R genes, generating transgenic plants able to silence key transporters in the pathogen might be a future, sustainable alternative to conventional breeding efforts constantly introducing novel R gene combinations, which might easily be overcome. In addition, spraying dsRNA to silence key nutrient uptake elements in pathogens might provide another effective method to control plant diseases (Wang et al., 2016).

所有与植物相互作用的微生物都必须从宿主体内获取代谢产物，以满足生长所需的营养。碳对于所有生物至关重要，因此从植物中充分获取糖分是植物病原体成功入侵的基本任务。阻止获取宿主糖似乎是控制植物病害的有前途的策略。提取植物糖可增强植物对病原体的抗性（Yamada等，2016）。然而，很难区分这是由于阻止病原体获取糖分的结果，还是植物中糖介导的信号传导受到干扰的结果（Milne等人，2019年; Moore等人，2015年）。自鉴定如果HXT1成为铁锈真菌中糖吸收的第一个证据（Voegele等，2001），则已经从不同的病原真菌中鉴定出许多糖转运蛋白（Saitoh等，2014； Schuler等，2015）。然而，糖饥饿对病原体生长，发育和致病性的影响仍不清楚。

条锈菌病菌 Tritici（Pst）是条纹（黄色）锈病的病原体，严重威胁全球小麦产量。到目前为止，对Pst的控制主要取决于带有特定抗性（R）基因的抗性品种的部署以及化学杀真菌剂的使用。但是，迫切需要新颖，可持续的方法来控制Pst。最近，己糖被证明是这种专性生物营养真菌利用的主要糖形式（Chang et al。，2017）。在这项研究中，我们克隆了己糖转运蛋白基因PsHXT1，这是在Pst感染过程中唯一被高度诱导的基因（郑等人，2013）。对PsHXT1 / Ps HXT1的进一步分析显示，一个主要的促进子超家族（MFS）共转运体具有12个跨膜节段的典型特征（图1a）。PsHXT1的种内多态性似乎相当低，因为所有11个比较的Pst基因组在核苷酸水平上都显示出99％以上的相似性（图1b）。尽管在密切相关的物种中，物种间的变异范围在83％至91％之间，但Ps HXT1明显不同于迄今表征的其他锈菌真菌葡萄糖转运蛋白。它与Uf的相似度仅为26％HXT1（图1c）。由于与效应子相比，参与糖获取的基因更加保守（Oliva和Quibod，2017年），这些基因/蛋白质可能代表了控制植物病害的新方法的有希望的目标。

Transcript levels of PsHXT1 during Pst infection were analysed by qRT‐PCR for the complete invasion process (Figure 1d). Transcript levels of PsHXT1 increased from 12 h post‐inoculation (hpi), when primary infection starts with substomatal vesicle formation, and increased continuously to reach a maximum at 168 hpi, when branched hyphae develop and more haustoria are formed. Thereafter, transcript levels sharply decrease to a very low level. This result indicates that PsHXT1 is indispensable for establishing the Pst–wheat interaction.

为了确定Ps HXT1的亚细胞定位，生成了Ps HXT1-GFP融合蛋白并在酵母中表达。Ps HXT1被证明定位于质膜（图1e）。通过在本氏烟草中的表达进一步分析了PS HXT1的亚细胞定位（图1f）。质膜裂解和膜标记SynaptoRed™C2（FM4-64）染色均证实了Ps HXT1的质膜定位。基于相似的亚细胞定位，Ps HXT1可以作为Uf HXT1发挥转运蛋白的作用（Voegele等，2001）。

为了识别的生化特性诗HXT1，PsHXT1在表达酿酒酵母突变株EBY.VW4000，它缺乏识别的所有20个己糖转运。Ps HXT1显示出对葡萄糖的底物偏爱（图1g）。在最佳条件下，Ps HXT1的Km为59± 12μm，Vmax为7.75±2.33 n m（图1h）。Ps HXT1的最佳pH约为5.5，但在4至7的pH范围内，转运活性仍保持较高水平。两种不同的质子体，分别是羰基氰化物间氯苯CC（CCCP）和2,4-二硝基苯酚（DNP）。能够抑制PS HXT1。SH基团抑制剂对氯汞苯磺酸盐（pCMBS）对Ps HXT1活性没有影响。竞争实验证实，Ps HXT1仅对葡萄糖具有高亲和力。所有这些结果表明，Ps HXT1是葡萄糖-质子的同向转运体。

为了确定的生物学功能PsHXT1在Pst-小麦相互作用，PsHXT1用大麦条纹花叶病毒（BSMV）介导的宿主诱导的基因沉默（HIGS）沉默。选择两个独立的片段（PsHXT1-1as和PsHXT1-2as）使PsHXT1沉默，而PsINVas用作阳性对照（Chang等，2017）。Pst的疾病表型观察感染14天。用BSMV：PsHXT1-1as或BSMV：PsHXT1-2as处理的植物的病害表型降低（图1i）。对被感染叶片上尿素含量的统计分析进一步支持了疾病表型的差异。此外，生物量比表明，用BSMV：PsHXT1-1as或BSMV：PsHXT1-2as处理的叶片中Pst的生物量与用BSMV：00处理的叶片显着降低。通过组织学观察在PsHXT1沉默的植物中检查了Pst的发育和生长（图1j）。在24 HPI，的Pst形成更多的分支，在近30％的病例中可以观察到气孔下囊泡膨胀。这表明建立与PsHXT1沉默的植物的Pst-小麦相互作用可能存在问题。在48 hpi时，菌丝显示异常发育并显示出高水平的畸形（在近70％的病例中）。在120 hpi，PsHXT1沉默的植物中Pst的感染面积显着减少。两者合计，这些结果表明沉默PsHXT1显着限制了小麦感染期间Pst的正常生长和发育，从而导致真菌生物量和疾病症状的减少。

结合以前关于Ps INV的研究（Chang等，2017），可以得出结论，糖饥饿不仅会损害Pst的生长和发育，而且会减慢病原体的增殖。据我们所知，这是第一个体内有证据表明，糖饥饿会限制病原体的生长和毒力，而不会与信号作用混淆。这为糖饥饿介导的小麦条锈病的控制开辟了新的前景，并表明阻断病原体的糖吸收可能是一种控制病原菌生长和繁殖的新策略。尽管已将大多数注意力集中在寻找效应子和R基因上，但是产生能够使病原体中关键转运蛋白沉默的转基因植物可能是常规育种努力的未来可持续发展选择，不断引入新的R基因组合，这很容易克服。此外，喷洒dsRNA沉默病原体中关键的养分吸收元素可能提供另一种有效的控制植物病害的方法（Wang等人，2016）。