Molecular variation of satellite DNA beta molecules associated with Malvastrum yellow vein virus and their role in pathogenicity

Previous studies have found that the diversity of begomovirus-associated DNA beta satellites is related to host and geographical origin. In this study, we have cloned and sequenced 20 different isolates of DNA beta molecules associated with Malvastrum yellow vein virus (MYVV) isolated from Malvastrum coromandelianum plants in different geographical locations of Yunnan Province, China. Analyses of their molecular variation indicate that the satellites are clustered together according to their geographical location but that they have only limited sequence diversity. Infectivity tests using infectious clones of MYVV and its associated DNA beta molecule indicate that MYVV DNA beta is indispensable for symptom induction in Nicotiana benthamiana, N. glutinosa, Petunia hybrida, and M. coromandelianum plants. Furthermore, we showed that MYVV interacts functionally with heterologous DNA beta molecules in N. benthamiana plants.     

Effects of the virus satellite gene βC1 on host plant defense signaling and volatile emission

Tomato Yellow Leaf Curl China virus spreads together with its invasive vector, the silverleaf whitefly B biotype, which exhibits higher growth rates on infected plants. Previous studies indicate that the virus satellite gene βC1 accounts for the visible symptoms of infection and inhibits the constitutive expression of jasmonic acid (JA)—a phytohormone involved in plant defense against whiteflies—and of some JA-regulated genes. Here we present new details of the effects of on plant signaling and defense, obtained with (non-host) transgenic Arabidopsis thaliana and Nicotiana benthamiana plants. We found that JA induction in response to wounding was reduced in plants expressing βC1. This result implies that βC1 acts on conserved plant regulation mechanisms and might impair the entire JA defense pathway. Furthermore, transformed N. benthamiana plants exhibited elevated emissions of the volatile compound linalool, suggesting that βC1 also influences plant-derived olfactory cues available to vector and non-vector insects.     

Fungal effector SIB1 of Colletotrichum orbiculare has unique structural features and can suppress plant immunity in Nicotiana benthamiana

Fungal plant pathogens secrete virulence-related proteins, called effectors, to establish host infection; however, the details are not fully understood yet. Functional screening of effector candidates using Agrobacterium-mediated transient expression assay in Nicotiana benthamiana identified two virulence-related effectors, named SIB1 and SIB2 (Suppression of Immunity in N. benthamiana), of an anthracnose fungus Colletotrichum orbiculare, which infects both cucurbits and N. benthamiana. The Agrobacterium-mediated transient expression of SIB1 or SIB2 increased the susceptibility of N. benthamiana to C. orbiculare, which suggested these effectors can suppress immune responses in N. benthamiana. The presence of SIB1 and SIB2 homologs was found to be limited to the genus Colletotrichum. SIB1 suppressed both (i) the generation of reactive oxygen species triggered by two different pathogen-associated molecular patterns, chitin and flg22, and (ii) the cell death response triggered by the Phytophthora infestans INF1 elicitin in N. benthamiana. We determined the NMR-based structure of SIB1 to obtain its structural insights. The three-dimensional structure of SIB1 comprises five β-strands, each containing three disulfide bonds. The overall conformation was found to be a cylindrical shape, such as the well-known antiparallel β-barrel structure. However, the β-strands were found to display a unique topology, one pair of these β-strands formed a parallel β-sheet. These results suggest that the effector SIB1 present in Colletotrichum fungi has unique structural features and can suppress pathogen-associated molecular pattern–triggered immunity in N. benthamiana.     

Selective autophagic receptor NbNBR1 prevents NbRFP1-mediated UPS-dependent degradation of βC1 to promote geminivirus infection

Autophagy is an evolutionarily conserved, lysosomal/vacuolar degradation mechanism that targets cell organelles and macromolecules. Autophagy and autophagy-related genes have been studied for their antiviral and pro-viral roles in virus-infected plants. Here, we demonstrate the pro-viral role of a selective autophagic receptor NbNBR1 in geminivirus-infected Nicotiana benthamiana plants. The βC1 protein encoded by tomato yellow leaf curl China betasatellite (TYLCCNB) that is associated with tomato yellow leaf curl China virus (TYLCCNV) enhanced the expression level of NbNBR1. Then NbNBR1 interacted with βC1 to form cytoplasmic granules. Interaction of NbNBR1 with βC1 could prevent degradation of βC1 by the NbRFP1, an E3 ligase. Overexpression of NbNBR1 in N. benthamiana plants increased βC1 accumulation and promoted virus infection. In contrast, silencing or knocking out NbNBR1 expression in N. benthamiana suppressed βC1 accumulation and inhibited virus infection. A single amino acid substitution in βC1 (βC1^K4A) abolished its interaction with NbNBR1, leading to a reduced level of βC1^K4A. The TYLCCNV/TYLCCNB^K4A mutant virus caused milder disease symptoms and accumulated much less viral genomic DNAs in the infected plants. Collectively, the results presented here show how a viral satellite-encoded protein hijacks host autophagic receptor NbNBR1 to form cytoplasmic granules to protect itself from NbRFP1-mediated degradation and facilitate viral infection.     

Phytophthora sojae apoplastic effector AEP1 mediates sugar uptake by mutarotation of extracellular aldose and is recognized as a MAMP

Diseases caused by Phytophthora pathogens devastate many crops worldwide. During infection, Phytophthora pathogens secrete effectors, which are central molecules for understanding the complex plant–Phytophthora interactions. In this study, we profiled the effector repertoire secreted by Phytophthora sojae into the soybean (Glycine max) apoplast during infection using liquid chromatography–mass spectrometry. A secreted aldose 1-epimerase (AEP1) was shown to induce cell death in Nicotiana benthamiana, as did the other two AEP1s from different Phytophthora species. AEP1 could also trigger immune responses in N. benthamiana, other Solanaceae plants, and Arabidopsis (Arabidopsis thaliana). A glucose dehydrogenase assay revealed AEP1 encodes an active AEP1. The enzyme activity of AEP1 is dispensable for AEP1-triggered cell death and immune responses, while AEP-triggered immune signaling in N. benthamiana requires the central immune regulator BRASSINOSTEROID INSENSITIVE 1-associated receptor kinase 1. In addition, AEP1 acts as a virulence factor that mediates P. sojae extracellular sugar uptake by mutarotation of extracellular aldose from the α-anomer to the β-anomer. Taken together, these results revealed the function of a microbial apoplastic effector, highlighting the importance of extracellular sugar uptake for Phytophthora infection. To counteract, the key effector for sugar conversion can be recognized by the plant membrane receptor complex to activate plant immunity.

Phytophthora sojae apoplastic effector AEP1 triggers pattern-triggered immunity in nonhost plants and contributes to P. sojae virulence by promoting the uptake of extracellular sugar.     

Suppression of RNA Silencing by a Plant DNA Virus Satellite Requires a Host Calmodulin-Like Protein to Repress RDR6 Expression

In plants, RNA silencing plays a key role in antiviral defense. To counteract host defense, plant viruses encode viral suppressors of RNA silencing (VSRs) that target different effector molecules in the RNA silencing pathway. Evidence has shown that plants also encode endogenous suppressors of RNA silencing (ESRs) that function in proper regulation of RNA silencing. The possibility that these cellular proteins can be subverted by viruses to thwart host defense is intriguing but has not been fully explored. Here we report that the Nicotiana benthamiana calmodulin-like protein Nbrgs-CaM is required for the functions of the VSR βC1, the sole protein encoded by the DNA satellite associated with the geminivirus Tomato yellow leaf curl China virus (TYLCCNV). Nbrgs-CaM expression is up-regulated by the βC1. Transgenic plants over-expressing Nbrgs-CaM displayed developmental abnormities reminiscent of βC1-associated morphological alterations. Nbrgs-CaM suppressed RNA silencing in an Agrobacterium infiltration assay and, when over-expressed, blocked TYLCCNV-induced gene silencing. Genetic evidence showed that Nbrgs-CaM mediated the βC1 functions in silencing suppression and symptom modulation, and was required for efficient virus infection. Moreover, the tobacco and tomato orthologs of Nbrgs-CaM also possessed ESR activity, and were induced by betasatellite to promote virus infection in these Solanaceae hosts. We further demonstrated that βC1-induced Nbrgs-CaM suppressed the production of secondary siRNAs, likely through repressing RNA-DEPENDENT RNA POLYMERASE 6 (RDR6) expression. RDR6-deficient N. benthamiana plants were defective in antiviral response and were hypersensitive to TYLCCNV infection. More significantly, TYLCCNV could overcome host range restrictions to infect Arabidopsis thaliana when the plants carried a RDR6 mutation. These findings demonstrate a distinct mechanism of VSR for suppressing PTGS through usurpation of a host ESR, and highlight an essential role for RDR6 in RNA silencing defense response against geminivirus infection.     

Tomato 14-3-3 Protein 7 Positively Regulates Immunity-Associated Programmed Cell Death by Enhancing Protein Abundance and Signaling Ability of MAPKKK α

Programmed cell death (PCD) is triggered when Pto, a Ser-Thr protein kinase, recognizes either the AvrPto or AvrPtoB effector from Pseudomonas syringae pv tomato. This PCD requires mitogen-activated protein kinase kinase kinase (MAPKKK α ) as a positive regulator in tomato (Solanum lycopersicum) and Nicotiana benthamiana. To examine how PCD-eliciting activity of the tomato MAPKKK α protein is regulated, we screened for MAPKKK α -interacting proteins in tomato and identified a 14-3-3 protein, TFT7. Virus-induced gene silencing using the TFT7 gene in N. benthamiana compromised both Pto- and MAPKKK α -mediated PCD, and coexpression of TFT7 with tomato MAPKKK α enhanced MAPKKK α -mediated PCD. TFT7 was also required for PCD associated with several other disease resistance proteins and contributed to resistance against P. syringae pv tomato. Coexpression of TFT7 with MAPKKK α in vivo caused increased accumulation of the kinase and enhanced phosphorylation of two MAP kinases. TFT7 protein contains a phosphopeptide binding motif that is present in human 14-3-3 ϵ, and substitutions in this motif abolished interaction with MAPKKK α in vivo and also the PCD-enhancing activity of TFT7. A 14-3-3 binding motif, including a putative phosphorylated Ser-535, is present in the C-terminal region of MAPKKK α. An S535A substitution in MAPKKK α reduced interaction with TFT7 and both PCD-eliciting ability and stability of MAPKKK α. Our results provide new insights into a role for 14-3-3 proteins in regulating immunity-associated PCD pathways in plants.     

Partitioning of beta‐diversity reveals distinct assembly mechanisms of plant and soil microbial communities in response to nitrogen enrichment

Nitrogen (N) deposition poses a serious threat to terrestrial biodiversity and alters plant and soil microbial community composition. Species turnover and nestedness reflect the underlying mechanisms of variations in community composition. However, it remains unclear how species turnover and nestedness contribute to different responses of taxonomic groups (plants and soil microbes) to N enrichment. Here, based on a 13‐year consecutive multi‐level N addition experiment in a semiarid steppe, we partitioned community β‐diversity into species turnover and nestedness components and explored how and why plant and microbial communities reorganize via these two processes following N enrichment. We found that plant, soil bacterial, and fungal β‐diversity increased, but their two components showed different patterns with increasing N input. Plant β‐diversity was mainly driven by species turnover under lower N input but by nestedness under higher N input, which may be due to a reduction in forb species, with low tolerance to soil Mn²⁺, with increasing N input. However, turnover was the main contributor to differences in soil bacterial and fungal communities with increasing N input, indicating the phenomenon of microbial taxa replacement. The turnover of bacteria increased greatly whereas that of fungi remained within a narrow range with increasing N input. We further found that the increased soil Mn²⁺ concentration was the best predictor for increasing nestedness of plant communities under higher N input, whereas increasing N availability and acidification together contributed to the turnover of bacterial communities. However, environmental factors could explain neither fungal turnover nor nestedness. Our findings reflect two different pathways of community changes in plants, soil bacteria, and fungi, as well as their distinct community assembly in response to N enrichment. Disentangling the turnover and nestedness of plant and microbial β‐diversity would have important implications for understanding plant–soil microbe interactions and seeking conservation strategies for maintaining regional diversity.     

Phospholipase D δ knock-out mutants are tolerant to severe drought stress

Phospholipase D (PLD) is involved in different plant processes, ranging from responses to abiotic and biotic stress to plant development. Phospholipase Dδ (PLDδ) is activated in dehydration and salt stress, producing the lipid second messenger phosphatidic acid. In this work we show that pldδ Arabidopsis mutants were more tolerant to severe drought than wild-type plants. PLDδ has been shown to be required for ABA regulation of stomatal closure of isolated epidermal peels. However, there was no significant difference in stomatal conductance at the whole plant level between wild-type and pldδ mutants. Since PLD hydrolyses structural phospholipids, then we looked at membrane integrity. Ion leakage measurements showed that during dehydration of leaf discs pldδ mutant has less membrane degradation compared to the wild-type. We further analyzed the mutants and showed that pldδ have higher mRNA levels of RAB18 and RD29A compared to wild-type plants under normal growth conditions. Transient expression of AtPLDδ in Nicotiana benthamiana plants induced a wilting phenotype. These findings suggest that, in wt plants PLDδ disrupt membranes in severe drought stress and, in the absence of the protein (PLDδ knock-out) might drought-prime the plants, making them more tolerant to severe drought stress. The results are discussed in relation to PLDδ role in guard cell signaling and drought tolerance.     

Genetic Determinants of Symptoms on Viral DNA Satellites

Begomovirus-DNA-β disease complexes induce different symptom phenotypes in their hosts. To investigate the genetic determinants of the phenotypic differences, Nicotiana spp. and tomato plants were inoculated with infectious clones of Tobacco curly shoot virus (TbCSV)/TbCSV DNA-β (TbCSB) and Tomato yellow leaf curl China virus (TYLCCNV)/TYLCCNV DNA-β (TYLCCNB) pseudorecombinants and showed that TYLCCNB induced characteristic vein-thickening and enation symptoms, while TbCSB only slightly exacerbated the leaf-curling symptoms, regardless of the helper virus being used. The roles of DNA-β-encoded βC1 and a 430-nucleotide fragment containing the A-rich region and the putative βC1 promoter region of the βC1 gene (referred to as AP) in symptom development were further investigated by constructing hybrid satellites in which the βC1 coding region or AP was exchanged between the two satellite molecules. A TYLCCNB hybrid with TbCSB βC1 lost the ability to elicit the vein-thickening and enation phenotypes. TbCSB hybrids containing the TYLCCNB βC1 or AP fragment failed to induce the characteristic vein thickening and enations. A TYLCCNB hybrid having the TbCSB AP fragment produced the enations, but the number of enations was less and their sizes were reduced. Differently from the phloem-specific pattern of the TYLCCNB promoter, a full-length fragment upstream of the TbCSB βC1 gene confers a constitutive β-glucuronidase expression pattern in transgenic tobacco plants. The above results indicate that the DNA-β-encoded βC1 protein is the symptom determinant, but the promoter of the βC1 gene has influence on symptom production.Geminiviruses are small plant viruses with circular single-stranded DNA (ssDNA) genomes that are encapsidated in unique twinned (geminate) particles. Members of the genus Begomovirus are transmitted by whiteflies (Bemisia tabaci) and infect dicotyledonous plants (42). Begomoviruses have either one or two circular ssDNA genomic components (DNA-A and DNA-B). The DNA-A component is capable of autonomous replication and encapsidation, whereas the DNA-B component encodes two proteins (BC1 and BV1) involved in movement (14). Recently, some monopartite begomoviruses have been found in association with a novel satellite DNA molecule, referred to as DNA-β and now known as a betasatellite (2, 5, 20, 22, 38, 45). DNA-β is approximately half the size of the viral genomic DNA, and apart from a nonanucleotide sequence (TAATATTAC), it has little sequence identity with viral genomic DNA. DNA-β depends on the helper virus for replication and encapsidation and, in turn, is required for the induction of bona fide disease symptoms. DNA-β bears a βC1 open reading frame (ORF) on the complementary-sense strand, which is conserved among distinct betasatellites in terms of position and size. Mutational analyses and constitutive expression have shown that βC1 is a strong pathogenicity/symptom determinant (7, 34, 39).Begomovirus-DNA-β disease complexes are associated with a wide range of plant species and induce different sets of symptom phenotypes in their natural hosts (25). However, the contributions of the helper virus and the satellite molecule to symptom development are not clear. Tomato yellow leaf curl China virus (TYLCCNV) and Tobacco curly shoot virus (TbCSV) are monopartite begomoviruses associated with DNA-β, but they differ in the symptom phenotypes induced in Nicotiana spp. and Solanum lycopersicum (7, 22). In the present work, we report that the symptom differences between TYLCCNV/TYLCCNV DNA-β (TYLCCNB) and TbCSV/TbCSV DNA-β (TbCSB) are determined by DNA-β and the DNA-β-encoded βC1 protein is the symptom determinant, but the promoter of the βC1 gene has influence on symptom production.     

Evolution,Expression Differentiation and Interaction Specificity of Heterotrimeric G-Protein Subunit Gene Family in the Mesohexaploid Brassica rapa

Heterotrimeric G-proteins, comprising of Gα, Gβ, and Gγ subunits, are important signal transducers which regulate many aspects of fundamental growth and developmental processes in all eukaryotes. Initial studies in model plants Arabidopsis and rice suggest that the repertoire of plant G-protein is much simpler than that observed in metazoans. In order to assess the consequence of whole genome triplication events within Brassicaceae family, we investigated the multiplicity of G-protein subunit genes in mesohexaploid Brassica rapa, a globally important vegetable and oilseed crop. We identified one Gα (BraA.Gα1), three Gβ (BraA.Gβ1, BraA.Gβ2, and BraA.Gβ3), and five Gγ (BraA.Gγ1, BraA.Gγ2, BraA.Gγ3, BraA.Gγ4, and BraA.Gγ5) genes from B. rapa, with a possibility of 15 Gαβγ heterotrimer combinations. Our analysis suggested that the process of genome triplication coupled with gene-loss (gene-fractionation) phenomenon have shaped the quantitative and sequence diversity of G-protein subunit genes in the extant B. rapa genome. Detailed expression analysis using qRT-PCR assays revealed that the G-protein genes have retained ubiquitous but distinct expression profiles across plant development. The expression of multiple G-protein genes was differentially regulated during seed-maturation and germination stages, and in response to various phytohormone treatments and stress conditions. Yeast-based interaction analysis showed that G-protein subunits interacted in most of the possible combinations, with some degree of subunit-specific interaction specificity, to control the functional selectivity of G-protein heterotrimer in different cell and tissue-types or in response to different environmental conditions. Taken together, this research identifies a highly diverse G-protein signaling network known to date from B. rapa, and provides a clue about the possible complexity of G-protein signaling networks present across globally important Brassica species.     

Physiological Functions of the COPI Complex in Higher Plants

COPI vesicles are essential to the retrograde transport of proteins in the early secretory pathway. The COPI coatomer complex consists of seven subunits, termed α-, β-, β′-, γ-, δ-, ε-, and ζ-COP, in yeast and mammals. Plant genomes have homologs of these subunits, but the essentiality of their cellular functions has hampered the functional characterization of the subunit genes in plants. Here we have employed virus-induced gene silencing (VIGS) and dexamethasone (DEX)-inducible RNAi of the COPI subunit genes to study the in vivo functions of the COPI coatomer complex in plants. The β′-, γ-, and δ-COP subunits localized to the Golgi as GFP-fusion proteins and interacted with each other in the Golgi. Silencing of β′-, γ-, and δ-COP by VIGS resulted in growth arrest and acute plant death in Nicotiana benthamiana, with the affected leaf cells exhibiting morphological markers of programmed cell death. Depletion of the COPI subunits resulted in disruption of the Golgi structure and accumulation of autolysosome-like structures in earlier stages of gene silencing. In tobacco BY-2 cells, DEX-inducible RNAi of β′-COP caused aberrant cell plate formation during cytokinesis. Collectively, these results suggest that COPI vesicles are essential to plant growth and survival by maintaining the Golgi apparatus and modulating cell plate formation.     

Response of Endophytic Bacterial Communities in Potato Plants to Infection with Erwinia carotovora subsp. atroseptica

The term endophyte refers to interior colonization of plants by microorganisms that do not have pathogenic effects on their hosts, and various endophytes have been found to play important roles in plant vitality. In this study, cultivation-independent terminal restriction fragment length polymorphism analysis of 16S ribosomal DNA directly amplified from plant tissue DNA was used in combination with molecular characterization of isolates to examine the influence of plant stress, achieved by infection with the blackleg pathogen Erwinia carotovora subsp. atroseptica, on the endophytic population in two different potato varieties. Community analysis clearly demonstrated increased bacterial diversity in infected plants compared to that in control plants. The results also indicated that the pathogen stress had a greater impact on the bacteria population than the plant genotype had. Partial sequencing of the 16S rRNA genes of isolated endophytes revealed a broad phylogenetic spectrum of bacteria, including members of the α, β, and γ subgroups of the Proteobacteria, high- and low-G+C-content gram-positive organisms, and microbes belonging to the Flexibacter-Cytophaga-Bacteroides group. Screening of the isolates for antagonistic activity against E. carotovora subsp. atroseptica revealed that 38% of the endophytes protected tissue culture plants from blackleg disease.     

Plant Translation Elongation Factor 1Bβ Facilitates Potato Virus X (PVX) Infection and Interacts with PVX Triple Gene Block Protein 1

The eukaryotic translation elongation factor 1 (eEF1) has two components: the G-protein eEF1A and the nucleotide exchange factor eEF1B. In plants, eEF1B is itself composed of a structural protein (eEF1Bγ) and two nucleotide exchange subunits (eEF1Bα and eEF1Bβ). To test the effects of elongation factors on virus infection, we isolated eEF1A and eEF1B genes from pepper (Capsicum annuum) and suppressed their homologs in Nicotiana benthamiana using virus-induced gene silencing (VIGS). The accumulation of a green fluorescent protein (GFP)-tagged Potato virus X (PVX) was significantly reduced in the eEF1Bβ- or eEF1Bɣ-silenced plants as well as in eEF1A-silenced plants. Yeast two-hybrid and co-immunoprecipitation analyses revealed that eEF1Bα and eEF1Bβ interacted with eEF1A and that eEF1A and eEF1Bβ interacted with triple gene block protein 1 (TGBp1) of PVX. These results suggest that both eEF1A and eEF1Bβ play essential roles in the multiplication of PVX by physically interacting with TGBp1. Furthermore, using eEF1Bβ deletion constructs, we found that both N- (1-64 amino acids) and C-terminal (150-195 amino acids) domains of eEF1Bβ are important for the interaction with PVX TGBp1 and that the C-terminal domain of eEF1Bβ is involved in the interaction with eEF1A. These results suggest that eEF1Bβ could be a potential target for engineering virus-resistant plants.     

Oxygen and Hydrogen Stable Isotope Ratios of Bulk Needles Reveal the Geographic Origin of Norway Spruce in the European Alps

Background

Tracking timber is necessary in order to prevent illegal logging and protect local timber production, but there is as yet no suitable analytical traceability method. Stable isotope ratios in plants are known to reflect geographical variations. In this study we analysed four stable isotope ratios in order to develop a model able to identify the geographic origin of Norway spruce in the European Alps.

Methodology and Principal Findings

δ¹⁸O, δ²H, δ¹³C and δ¹⁵N were measured in bulk needles of Picea abies sampled in 20 sites in and around the European Alps. Environmental and spatial variables were found to be related to the measured isotope ratios. An ordinary least squares regression was used to identify the most important factor in stable isotope variability in bulk needles. Spatial autocorrelation was tested for all isotope ratios by means of Moran’s I. δ¹⁸O, δ²H and δ¹⁵N values differed significantly between sites. Distance from the coast had the greatest influence on δ²H, while latitude and longitude were strongly related to δ¹⁸O. δ¹³C values did not appear to have any relationship with geographical position, while δ¹⁵N values were influenced by distance from the motorway. The regression model improved the explanatory power of the spatial and environmental variables. Positive spatial autocorrelations were found for δ¹⁸O and δ²H values.

Conclusions

The δ ¹⁸O, δ²H and δ¹⁵N values in P. abies bulk needles are a suitable proxy to identify geographic origin as they vary according to geographical position. Although the regression model showed the explanatory variables to have significant power and stability, we conclude that our model might be improved by multivariate spatial interpolation of the δ ¹⁸O and δ²H values.     

Phytophthora infestans RXLR Effector PexRD2 Interacts with Host MAPKKKε to Suppress Plant Immune Signaling

Mitogen-activated protein kinase cascades are key players in plant immune signaling pathways, transducing the perception of invading pathogens into effective defense responses. Plant pathogenic oomycetes, such as the Irish potato famine pathogen Phytophthora infestans, deliver RXLR effector proteins to plant cells to modulate host immune signaling and promote colonization. Our understanding of the molecular mechanisms by which these effectors act in plant cells is limited. Here, we report that the P. infestans RXLR effector PexRD2 interacts with the kinase domain of MAPKKKε, a positive regulator of cell death associated with plant immunity. Expression of PexRD2 or silencing MAPKKKε in Nicotiana benthamiana enhances susceptibility to P. infestans. We show that PexRD2 perturbs signaling pathways triggered by or dependent on MAPKKKε. By contrast, homologs of PexRD2 from P. infestans had reduced or no interaction with MAPKKKε and did not promote disease susceptibility. Structure-led mutagenesis identified PexRD2 variants that do not interact with MAPKKKε and fail to support enhanced pathogen growth or perturb MAPKKKε signaling pathways. Our findings provide evidence that P. infestans RXLR effector PexRD2 has evolved to interact with a specific host MAPKKK to perturb plant immunity–related signaling.     

Biogeographic,climatic and spatial drivers differentially affect α-, β- and γ-diversities on oceanic archipelagos

Island biogeographic studies traditionally treat single islands as units of analysis. This ignores the fact that most islands are spatially nested within archipelagos. Here, we took a fundamentally different approach and focused on entire archipelagos using species richness of vascular plants on 23 archipelagos worldwide and their 174 constituent islands. We assessed differential effects of biogeographic factors (area, isolation, age, elevation), current and past climate (temperature, precipitation, seasonality, climate change velocity) and intra-archipelagic spatial structure (archipelago area, number of islands, area range, connectivity, environmental volume, inter-island distance) on plant diversity. Species diversity of each archipelago (γ) was additively partitioned into α, β, nestedness and replacement β-components to investigate the relative importance of environmental and spatial drivers. Multiple regressions revealed strong effects of biogeography and climate on α and γ, whereas spatial factors, particularly number of islands, inter-island distance and area range, were key to explain β. Structural equation models additionally suggested that γ is predominantly determined by indirect abiotic effects via its components, particularly β. This highlights that β and the spatial arrangement of islands are essential to understand insular ecology and evolution. Our methodological framework can be applied more widely to other taxa and archipelago-like systems, allowing new insights into biodiversity origin and maintenance.     

Global Analyses of Small Interfering RNAs Derived from Bamboo mosaic virus and Its Associated Satellite RNAs in Different Plants

Background

Satellite RNAs (satRNAs), virus parasites, are exclusively associated with plant virus infection and have attracted much interest over the last 3 decades. Upon virus infection, virus-specific small interfering RNAs (vsiRNAs) are produced by dicer-like (DCL) endoribonucleases for anti-viral defense. The composition of vsiRNAs has been studied extensively; however, studies of satRNA-derived siRNAs (satsiRNAs) or siRNA profiles after satRNA co-infection are limited. Here, we report on the small RNA profiles associated with infection with Bamboo mosaic virus (BaMV) and its two satellite RNAs (satBaMVs) in Nicotiana benthamiana and Arabidopsis thaliana.

Methodology/Principal Findings

Leaves of N. benthamiana or A. thaliana inoculated with water, BaMV alone or co-inoculated with interfering or noninterfering satBaMV were collected for RNA extraction, then large-scale Solexa sequencing. Up to about 20% of total siRNAs as BaMV-specific siRNAs were accumulated in highly susceptible N. benthamiana leaves inoculated with BaMV alone or co-inoculated with noninterfering satBaMV; however, only about 0.1% of vsiRNAs were produced in plants co-infected with interfering satBaMV. The abundant region of siRNA distribution along BaMV and satBaMV genomes differed by host but not by co-infection with satBaMV. Most of the BaMV and satBaMV siRNAs were 21 or 22 nt, of both (+) and (−) polarities; however, a higher proportion of 22-nt BaMV and satBaMV siRNAs were generated in N. benthamiana than in A. thaliana. Furthermore, the proportion of non-viral 24-nt siRNAs was greatly increased in N. benthamiana after virus infection.

Conclusions/Significance

The overall composition of vsiRNAs and satsiRNAs in the infected plants reflect the combined action of virus, satRNA and different DCLs in host plants. Our findings suggest that the structure and/or sequence demands of various DCLs in different hosts may result in differential susceptibility to the same virus. DCL2 producing 24-nt siRNAs under biotic stresses may play a vital role in the antiviral mechanism in N. benthamiana.