Expression profiling across wild and cultivated tomatoes supports the relevance of early miR482/2118 suppression for Phytophthora resistance.

Plants possess a battery of specific pathogen resistance (R-)genes. Precise R-gene regulation is important in the presence and absence of a pathogen. Recently, a microRNA family, miR482/2118, was shown to regulate the expression of a major class of R-genes, nucleotide-binding site leucine-rich repeats (NBS-LRRs). Furthermore, RNA silencing suppressor proteins, secreted by pathogens, prevent the accumulation of miR482/2118, leading to an upregulation of R-genes. Despite this transcriptional release of R-genes, RNA silencing suppressors positively contribute to the virulence of some pathogens. To investigate this paradox, we analysed how the regulation of NBS-LRRs by miR482/2118 has been shaped by the coevolution between Phytophthora infestans and cultivated and wild tomatoes. We used degradome analyses and qRT-PCR to evaluate and quantify the co-expression of miR482/2118 and their NBS-LRR targets. Our data show that miR482/2118-mediated targeting contributes to the regulation of NBS-LRRs in Solanum lycopersicum. Based on miR482/2118 expression profiling in two additional tomato species-with different coevolutionary histories with P. infestans-we hypothesize that pathogen-mediated RNA silencing suppression is most effective in the interaction between S. lycopersicum and P. infestans Furthermore, an upregulation of miR482/2118 early in the infection may increase susceptibility to P. infestans.

Population Genomics in Wild Tomatoes-The Interplay of Divergence and Admixture.

Hybridization between closely related plant species is widespread, but the outcomes of hybridization are not fully understood. This study investigates phylogenetic relationships and the history of hybridization in the wild tomato clade (Solanum sect. Lycopersicon). We sequenced RNA from individuals of 38 different populations and, by combining this with published data, build a comprehensive genomic data set for the entire clade. The data indicate that many taxa are not monophyletic and many individuals are admixed due to repeated hybridization. The most polymorphic species, Solanum peruvianum, has two genetic and geographical subpopulations, while its sister species, Solanum chilense, has distinct coastal populations and reduced heterozygosity indicating a recent expansion south following speciation from S. peruvianum circa 1.25 Ma. Discontinuous populations west of 72° are currently described as S. chilense, but are genetically intermediate between S. chilense and S. peruvianum. Based upon molecular, morphological, and crossing data, we test the hypothesis that these discontinuous "S. chilense" populations are an example of recombinational speciation. Recombinational speciation is rarely reported, and we discuss the difficulties in identifying it and differentiating between alternative demographic scenarios. This discovery presents a new opportunity to understand the genomic outcomes of hybridization in plants.

Signatures of selection and host-adapted gene expression of the Phytophthora infestans RNA silencing suppressor PSR2.

Phytophthora infestans is a devastating pathogen in agricultural systems. Recently, an RNA silencing suppressor (PSR2, 'Phytophthora suppressor of RNA silencing 2') has been described in P. infestans. PSR2 has been shown to increase the virulence of Phytophthora pathogens on their hosts. This gene is one of the few effectors present in many economically important Phytophthora species. In this study, we investigated: (i) the evolutionary history of PSR2 within and between species of Phytophthora; and (ii) the interaction between sequence variation, gene expression and virulence. In P. infestans, the highest PiPSR2 expression was correlated with decreased symptom expression. The highest gene expression was observed in the biotrophic phase of the pathogen, suggesting that PSR2 is important during early infection. Protein sequence conservation was negatively correlated with host range, suggesting host range as a driver of PSR2 evolution. Within species, we detected elevated amino acid variation, as observed for other effectors; however, the frequency spectrum of the mutations was inconsistent with strong balancing selection. This evolutionary pattern may be related to the conservation of the host target(s) of PSR2 and the absence of known corresponding R genes. In summary, our study indicates that PSR2 is a conserved effector that acts as a master switch to modify plant gene regulation early during infection for the pathogen's benefit. The conservation of PSR2 and its important role in virulence make it a promising target for pathogen management.

Evolutionarily Dynamic, but Robust, Targeting of Resistance Genes by the miR482/2118 Gene Family in the Solanaceae.

Plants are exposed to pathogens around the clock. A common resistance response in plants upon pathogen detection is localized cell death. Given the irreversible nature of this response, multiple layers of negative regulation are present to prevent the untimely or misexpression of resistance genes. One layer of negative regulation is provided by a recently discovered microRNA (miRNA) gene family, miR482/2118. This family targets the transcripts of resistance genes in plants. We investigated the evolutionary history and specificity of this miRNA gene family within the Solanaceae. This plant family includes many important crop species, providing a set of well-defined resistance gene repertoires. Across 14 species from the Solanaceae, we identified eight distinct miR482/2118 gene family members. Our studies show conservation of miRNA type and number in the group of wild tomatoes and, to a lesser extent, throughout the Solanaceae. The eight orthologous miRNA gene clusters evolved under different evolutionary constraints, allowing for individual subfunctionalization of the miRNAs. Despite differences in the predicted targeting behavior of each miRNA, the miRNA-R-gene network is robust due to its high degree of interconnectivity and redundant targeting. Our data suggest that the miR482/2118 gene family acts as an evolutionary buffer for R-gene sequence diversity.

A machine learning approach to identify hydrogenosomal proteins in Trichomonas vaginalis.

The protozoan parasite Trichomonas vaginalis is the causative agent of trichomoniasis, the most widespread nonviral sexually transmitted disease in humans. It possesses hydrogenosomes-anaerobic mitochondria that generate H(2), CO(2), and acetate from pyruvate while converting ADP to ATP via substrate-level phosphorylation. T. vaginalis hydrogenosomes lack a genome and translation machinery; hence, they import all their proteins from the cytosol. To date, however, only 30 imported proteins have been shown to localize to the organelle. A total of 226 nuclear-encoded proteins inferred from the genome sequence harbor a characteristic short N-terminal presequence, reminiscent of mitochondrial targeting peptides, which is thought to mediate hydrogenosomal targeting. Recent studies suggest, however, that the presequences might be less important than previously thought. We sought to identify new hydrogenosomal proteins within the 59,672 annotated open reading frames (ORFs) of T. vaginalis, independent of the N-terminal targeting signal, using a machine learning approach. Our training set included 57 gene and protein features determined for all 30 known hydrogenosomal proteins and 576 nonhydrogenosomal proteins. Several classifiers were trained on this set to yield an import score for all proteins encoded by T. vaginalis ORFs, predicting the likelihood of hydrogenosomal localization. The machine learning results were tested through immunofluorescence assay and immunodetection in isolated cell fractions of 14 protein predictions using hemagglutinin constructs expressed under the homologous SCSα promoter in transiently transformed T. vaginalis cells. Localization of 6 of the 10 top predicted hydrogenosome-localized proteins was confirmed, and two of these were found to lack an obvious N-terminal targeting signal.

Networks of gene sharing among 329 proteobacterial genomes reveal differences in lateral gene transfer frequency at different phylogenetic depths.

Lateral gene transfer (LGT) is an important mechanism of natural variation among prokaryotes. Over the full course of evolution, most or all of the genes resident in a given prokaryotic genome have been affected by LGT, yet the frequency of LGT can vary greatly across genes and across prokaryotic groups. The proteobacteria are among the most diverse of prokaryotic taxa. The prevalence of LGT in their genome evolution calls for the application of network-based methods instead of tree-based methods to investigate the relationships among these species. Here, we report networks that capture both vertical and horizontal components of evolutionary history among 1,207,272 proteins distributed across 329 sequenced proteobacterial genomes. The network of shared proteins reveals modularity structure that does not correspond to current classification schemes. On the basis of shared protein-coding genes, the five classes of proteobacteria fall into two main modules, one including the alpha-, delta-, and epsilonproteobacteria and the other including beta- and gammaproteobacteria. The first module is stable over different protein identity thresholds. The second shows more plasticity with regard to the sequence conservation of proteins sampled, with the gammaproteobacteria showing the most chameleon-like evolutionary characteristics within the present sample. Using a minimal lateral network approach, we compared LGT rates at different phylogenetic depths. In general, gene evolution by LGT within proteobacteria is very common. At least one LGT event was inferred to have occurred in at least 75% of the protein families. The average LGT rate at the species and class depth is about one LGT event per protein family, the rate doubling at the phylum level to an average of two LGT events per protein family. Hence, our results indicate that the rate of gene acquisition per protein family is similar at the level of species (by recombination) and at the level of classes (by LGT). The frequency of LGT per genome strongly depends on the species lifestyle, with endosymbionts showing far lower LGT frequencies than free-living species. Moreover, the nature of the transferred genes suggests that gene transfer in proteobacteria is frequently mediated by conjugation.