Using hierarchical clustering of secreted protein families to classify and rank candidate effectors of rust fungi

Rust fungi are obligate biotrophic pathogens that cause considerable damage on crop plants. Puccinia graminis f. sp. tritici, the causal agent of wheat stem rust, and Melampsora larici-populina, the poplar leaf rust pathogen, have strong deleterious impacts on wheat and poplar wood production, respectively. Filamentous pathogens such as rust fungi secrete molecules called disease effectors that act as modulators of host cell physiology and can suppress or trigger host immunity. Current knowledge on effectors from other filamentous plant pathogens can be exploited for the characterisation of effectors in the genome of recently sequenced rust fungi. We designed a comprehensive in silico analysis pipeline to identify the putative effector repertoire from the genome of two plant pathogenic rust fungi. The pipeline is based on the observation that known effector proteins from filamentous pathogens have at least one of the following properties: (i) contain a secretion signal, (ii) are encoded by in planta induced genes, (iii) have similarity to haustorial proteins, (iv) are small and cysteine rich, (v) contain a known effector motif or a nuclear localization signal, (vi) are encoded by genes with long intergenic regions, (vii) contain internal repeats, and (viii) do not contain PFAM domains, except those associated with pathogenicity. We used Markov clustering and hierarchical clustering to classify protein families of rust pathogens and rank them according to their likelihood of being effectors. Using this approach, we identified eight families of candidate effectors that we consider of high value for functional characterization. This study revealed a diverse set of candidate effectors, including families of haustorial expressed secreted proteins and small cysteine-rich proteins. This comprehensive classification of candidate effectors from these devastating rust pathogens is an initial step towards probing plant germplasm for novel resistance components.     

Infection assays in Arabidopsis reveal candidate effectors from the poplar rust fungus that promote susceptibility to bacteria and oomycete pathogens

Fungi of the Pucciniales order cause rust diseases which, altogether, affect thousands of plant species worldwide and pose a major threat to several crops. How rust effectors—virulence proteins delivered into infected tissues to modulate host functions—contribute to pathogen virulence remains poorly understood. Melampsora larici‐populina is a devastating and widespread rust pathogen of poplar, and its genome encodes 1184 identified small secreted proteins that could potentially act as effectors. Here, following specific criteria, we selected 16 candidate effector proteins and characterized their virulence activities and subcellular localizations in the leaf cells of Arabidopsis thaliana. Infection assays using bacterial (Pseudomonas syringae) and oomycete (Hyaloperonospora arabidopsidis) pathogens revealed subsets of candidate effectors that enhanced or decreased pathogen leaf colonization. Confocal imaging of green fluorescent protein‐tagged candidate effectors constitutively expressed in stable transgenic plants revealed that some protein fusions specifically accumulate in nuclei, chloroplasts, plasmodesmata and punctate cytosolic structures. Altogether, our analysis suggests that rust fungal candidate effectors target distinct cellular components in host cells to promote parasitic growth.     

玉米黑粉菌致病的分子机制研究进展

玉米黑粉菌（Ustilago maydis）可在其宿主植物玉米（Zea mays L.）地上部的所有器官诱导肿瘤发生。玉米黑粉菌成功定殖宿主并诱导形成肿瘤取决于与宿主植物多方位、多层次的相互作用以及该过程中发生的复杂的细胞和分子事件。本文综述了玉米黑粉菌与玉米互作研究的最新进展,介绍了玉米黑粉菌通过分泌效应子入侵、定殖玉米植株以及植株在分子水平上对入侵的响应;阐述了活体营养建立过程中,玉米黑粉菌与玉米通过效应子、激素、糖代谢酶和转运蛋白的差异调节,协调受感染宿主组织重新编程发育成膨大的植物肿瘤的关键因素,并对今后的研究方向进行了展望。     

Versatile effectors of phytopathogenic fungi target host immunity

Phytopathogenic fungi secrete a large arsenal of effector molecules, including proteinaceous effectors, small RNAs, phytohormones and derivatives thereof. The pathogenicity of fungal pathogens is primarily determined by these effectors that are secreted into host cells to undermine innate immunity, as well as to facilitate the acquisition of nutrients for their in planta growth and proliferation. After conventional and non-conventional secretion, fungal effectors are translocated into different subcellular compartments of the host cells to interfere with various biological processes. In extracellular spaces, apoplastic effectors cope with physical and chemical barriers to break the first line of plant defenses. Intracellular effectors target essential immune components on the plasma membrane, in the cytosol, including cytosolic organelles, and in the nucleus to suppress host immunity and reprogram host physiology, favoring pathogen colonization. In this review, we comprehensively summarize the recent advances in fungal effector biology, with a focus on the versatile virulence functions of fungal effectors in promoting pathogen infection and colonization. A perspective of future research on fungal effector biology is also discussed.     

杨树叶锈病研究

松杨栅锈菌（Melampsora larici-populina）导致的杨树叶锈病是杨树病害中分布最广、危害最大的一种病害,它造成杨树林的材积损失和材质下降,严重制约着杨树商品林和生态林的发展。该文从病原菌的分布和寄主种类、病原菌生理分化、寄主抗锈性及分子标记、防治措施等几方面介绍了杨树叶锈病的研究。     

杨树抗锈性研究现状

综述了落叶松一杨栅锈菌生理小种分化、寄主杨树抗锈性、抗锈病基因分子标记和抗病基因连锁遗传图谱构建技术;分析了我国杨树抗锈病育种研究的现状和亟待解决的问题。     

Disentangling the genetic origins of a plant pathogen during disease spread using an original molecular epidemiology approach

The advent of molecular epidemiology has greatly improved our ability to identify the population sources and track the pathogen movement. Yet the wide spatial and temporal scales usually considered are useful only to infer historical migration pathways. In this study, Bayesian genetic assignments and a landscape epidemiology approach were combined to unravel genetic origin and annual spread during a single epidemic of a plant pathogen: the poplar rust fungus Melampsora larici-populina. The study focused on a particular area-the Durance River valley-which enabled inoculum sources to be identified and channelled spread of the epidemic along a one-dimensional corridor. Spatio-temporal monitoring of disease showed that the epidemic began in the upstream part of the valley and spread out downstream. Using genetic assignment tests, individuals collected at the end of the epidemic were sorted into two genetic groups; very few hybrids were detected, although individuals from both groups coexisted locally downstream in the valley. The epidemic originated from two genetically distinct inoculum sources. Individuals of each group then dispersed southwards along the Durance River and became mixed in poplar riparian stands. These two genetic groups were found previously at a wider spatial scale and proved to result from distinct evolutionary histories on either wild or cultivated poplars. This study showed that the two groups can mix during an epidemic but do not hybridize because they then reproduce asexually. In general, the methods employed here could be useful for elucidating the genetic origin and retracing the colonization history and migration pathways of recent epidemics.     

Quantitative mass spectrometry catalogues Salmonella pathogenicity island-2 effectors and identifies their cognate host binding partners

Gram-negative bacterial pathogens have developed specialized secretion systems to transfer bacterial proteins directly into host cells. These bacterial effectors are central to virulence and reprogram host cell processes to favor bacterial survival, colonization, and proliferation. Knowing the complete set of effectors encoded by a particular pathogen is the key to understanding bacterial disease. In addition, the identification of the molecular assemblies that these effectors engage once inside the host cell is critical to determining the mechanism of action of each effector. In this work we used stable isotope labeling of amino acids in cell culture (SILAC), a powerful quantitative proteomics technique, to identify the proteins secreted by the Salmonella pathogenicity island-2 type three secretion system (SPI-2 T3SS) and to characterize the host interaction partners of SPI-2 effectors. We confirmed many of the known SPI-2 effectors and were able to identify several novel substrate candidates of this secretion system. We verified previously published host protein-effector binding pairs and obtained 11 novel interactions, three of which were investigated further and confirmed by reciprocal co-immunoprecipitation. The host cell interaction partners identified here suggest that Salmonella SPI-2 effectors target, in a concerted fashion, cellular processes such as cell attachment and cell cycle control that are underappreciated in the context of infection. The technology outlined in this study is specific and sensitive and serves as a robust tool for the identification of effectors and their host targets that is readily amenable to the study of other bacterial pathogens.     

Sequencing the genome of Marssonina brunnea reveals fungus-poplar co-evolution

ABSTRACT: BACKGROUND: The fungus Marssonina brunnea is a causal pathogen of Marssonina leaf spot that devastates poplar plantations by defoliating susceptible trees before normal fall leaf drop. RESULTS: \We sequence the genome of M. brunnea with a size of 52 Mb assembled into 89 scaffolds, representing the first sequenced Dermateaceae genome. By inoculating this fungus onto a poplar hybrid clone, we investigate how M. brunnea interacts and co-evolves with its host to colonize poplar leaves. While a handful of virulence genes in M. brunnea, mostly from the LysM family, are detected to up-regulate during infection, the poplar down-regulates its resistance genes, such as nucleotide binding site domains and leucine rich repeats, in response to infection. From 10,027 predicted proteins of M. brunnea in a comparison with those from poplar, we identify four poplar transferases that stimulate the host to resist M. brunnea. These transferas-encoding genes may have driven the co-evolution of M. brunnea and Populus during the process of infection and anti-infection. CONCLUSIONS: Our results from the draft sequence of the M. brunnea genome provide evidence for genomegenome interactions that play an important role in poplar-pathogen co-evolution. This knowledge could help to design effective strategies for controlling Marssonina leaf spot in poplar.