TaADF3, an actin-depolymerizing factor, negatively modulates wheat resistance against Puccinia striiformis

The actin cytoskeleton has been implicated in plant defense against pathogenic fungi, oomycetes and bacteria. Actin depolymerizing factors (ADF) are stimulus responsive actin cytoskeleton modulators. However, there is limited evidence linking ADFs with plant defense against pathogens. In this study, we have isolated and functionally characterized a stress-responsive ADF gene (TaADF3) from wheat, which was detectable in all examined wheat tissues. TaADF3 is a three-copy gene located on chromosomes 5AL, 5BL and 5DL. A particle bombardment assay in onion epidermal cells revealed the cytoplasmic and nuclear localization of TaADF3. The expression of TaADF3 was inducible by abscisic acid (ABA), as well as various abiotic stresses (drought and cold) and virulent Puccinia striiformis f. sp. tritici (Pst) but was down regulated in response to avirulent Pst. Virus-induced silencing of TaADF3 copies enhanced wheat resistance to avirulent Pst, with decreased reactive oxygen species (ROS) accumulation and hypersensitive response (HR). Upon treatment with virulent Pst, TaADF3-knockdown plants exhibited reduced susceptibility, which was accompanied by increased ROS production and HR. Interestingly, the silencing of TaADF3 resulted in hindered pathogen penetration and haustoria formation for both avirulent and virulent Pst. Moreover, the array and distribution of actin filaments was transformed in TaADF3-knockdown epidermal cells, which possibly facilitated attenuating the fungus penetration. Thus, our findings suggest that TaADF3 positively regulates wheat tolerance to abiotic stresses and negatively regulates wheat resistance to Pst in an ROS-dependent manner, possibly underlying the mechanism of impeding fungal penetration dependent on the actin architecture dynamics

Nitric oxide and reactive oxygen species coordinately regulate the germination of Puccinia striiformis f. sp. tritici urediniospores

Nitric oxide (NO) and reactive oxygen species (ROS) function as signaling molecules in a number of critical signal transduction pathways in plants, including plant biotic interactions. In addition to the role of plant-derived NO and ROS in plant resistance, which has been well documented, pathogen-produced NO and ROS have recently emerged as important players in fungal development and pathogenesis. However, the effects of pathogenic fungi-derived NO and ROS on signaling pathways during fungal pre-infection development remain unknown. Here, using a combination of pharmacological approaches and confocal microscopy, we investigated the roles of NO and ROS during the germination of Puccinia striiformis Westend f. sp. tritici (Pst) the wheat stripe rust pathogen. Both NO and ROS have a crucial role in uredinial germination. The scavengers of NO and ROS delayed spore germination and decreased the lengths of germ tubes. A similar phenotype was produced after treatment with the promoter. However, the spores germinated and grew normally when the levels of NO and ROS were simultaneously elevated by the application of a promoter of NO and a donor of ROS. Confocal laser microscopy indicated that both NO and ROS preferentially localized at the germ pores and apexes of growing germ tubes when the ROS/NO ratio in the spores was maintained in a specific range. We concluded that both NO and ROS are critical signaling molecules in the pre-infection development of Pst and that the polar growth of the germ tube is coordinately regulated by NO and ROS

Candidate effector proteins of the necrotrophic apple canker pathogen Valsa mali can suppress BAX-induced PCD.

Canker caused by the Ascomycete Valsa mali is the most destructive disease of apple in Eastern Asia, resulting in yield losses of up to 100 %. This necrotrophic fungus induces severe necrosis on apple, eventually leading to the death of the whole tree. Identification of necrosis inducing factors may help to unravel the molecular bases for colonization of apple trees by V. mali. As a first step towards this goal, we identified and characterized the V. mali repertoire of Candidate Effector Proteins (CEPs). In total, 193 secreted proteins with no known function were predicted from genomic data, of which 101 were V. mali-specific. Compared to non-CEPs predicted for the V. mali secretome, CEPs have shorter sequence length and a higher content of cysteine residues. Based on transient over-expression in Nicotiana benthamiana performed for 70 randomly selected CEPs, seven V. mali Effector Proteins (VmEPs) were shown to significantly suppress BAX-induced PCD. Furthermore, targeted deletion of VmEP1 resulted in a significant reduction of virulence. These results suggest that V. mali expresses secreted proteins that can suppress PCD usually associated with effector-triggered immunity (ETI). ETI in turn may play an important role in the V. mali-apple interaction. The ability of V. mali to suppress plant ETI sheds a new light onto the interaction of a necrotrophic fungus with its host plant

TaSYP71, a Qc-SNARE, Contributes to Wheat Resistance against Puccinia striiformis f. sp. tritici

N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are involved in plant resistance; however, the role of SYP71 in the regulation of plant–pathogen interactions is not well known. In this study, we characterized a plant-specific SNARE in wheat, TaSYP71, which contains a Qc-SNARE domain. Three homologues are localized on chromosome 1AL, 1BL and 1DL. Using Agrobacterium-mediated transient expression, TaSYP71 was localized to the plasma membrane in Nicotiana benthamiana. Quantitative real-time PCR assays revealed that TaSYP71 homologues was induced by NaCl, H2O2 stress and infection by virulent and avirulent Puccinia striiformis f. sp. tritici (Pst) isolates. Heterologous expression of TaSYP71 in Schizosaccharomyces pombe elevated tolerance to H2O2. Meanwhile, H2O2 scavenging gene (TaCAT) was downregulated in TaSYP71 silenced plants treated by H2O2 compared to that in control, which indicated that TaSYP71 enhanced tolerance to H2O2 stress possibly by influencing the expression of TaCAT to remove the excessive H2O2 accumulation. When TaSYP71 homologues were all silenced in wheat by the virus-induced gene silencing system, wheat plants were more susceptible to Pst, with larger infection area and more haustoria number, but the necrotic area of wheat mesophyll cells were larger, one possible explanation that minor contribution of resistance to Pst was insufficient to hinder pathogen extension when TaSYP71were silenced, and the necrotic area was enlarged accompanied with the pathogen growth. Of course, later cell death could not be excluded. In addition, the expression of pathogenesis-related genes were down-regulated in TaSYP71 silenced wheat plants. These results together suggest that TaSYP71 play a positive role in wheat defence against Pst

Exploration of microRNAs and their targets engaging in the resistance interaction between wheat and stripe rust

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat worldwide. miRNAs are important regulators, they play very central roles in plant organ development, vegetable phase change and defense responses. In this study, two miRNA libraries from wheat cultivar Xingzi 9104 (XZ) challenged with the avirulent Pst race CYR32 and sterile water were constructed, respectively. A total of 596 miRNA candidates were obtained. 420 wheat-specific candidate miRNAs were screened in adult plants challenged with Pst using microarray-based analyses. We analyzed the abundance of candidate miRNAs, and the levels of a subset of candidate miRNAs were determined by quantitative real time PCR (qRT-PCR). The qRT-PCR results indicated that some miRNAs were involved in the incompatible interaction between wheat and Pst. In addition, we identified some miRNAs differentially expressed in different leaves. Additionally, the target genes of wheat miRNAs were confirmed by using degradome sequencing technology. Most of the annotated target genes are related to signal transduction, energy metabolism, and other functions. We selected some target genes for relative expression analysis using qRT-PCR, and found that RabGAP/TBC domain-containing protein, zinc finger protein and Cysteine-rich receptor-like protein kinase 41 may play important role in the incompatible interaction between XZ and CYR32. Intriguingly, miRNAs and target gene seem to form a complicated regulation network that regulates the wheat-Pst interaction. Our data provide the foundation for evaluating the important regulatory roles of miRNAs in the wheat-Pst interaction