David J. Studholme

Genome sequence and analysis of the Irish potato famine pathogen Phytophthora infestans

Phytophthora infestans is the most destructive pathogen of potato and a model organism for the oomycetes, a distinct lineage of fungus-like eukaryotes that are related to organisms such as brown algae and diatoms. As the agent of the Irish potato famine in the mid-nineteenth century, P. infestans has had a tremendous effect on human history, resulting in famine and population displacement. To this day, it affects world agriculture by causing the most destructive disease of potato, the fourth largest food crop and a critical alternative to the major cereal crops for feeding the world’s population. Current annual worldwide potato crop losses due to late blight are conservatively estimated at

InterPro, progress and status in 2005

6.7 billion. Management of this devastating pathogen is challenged by its remarkable speed of adaptation to control strategies such as genetically resistant cultivars. Here we report the sequence of the P. infestans genome, which at ∼240 megabases (Mb) is by far the largest and most complex genome sequenced so far in the chromalveolates. Its expansion results from a proliferation of repetitive DNA accounting for ∼74% of the genome. Comparison with two other Phytophthora genomes showed rapid turnover and extensive expansion of specific families of secreted disease effector proteins, including many genes that are induced during infection or are predicted to have activities that alter host physiology. These fast-evolving effector genes are localized to highly dynamic and expanded regions of the P. infestans genome. This probably plays a crucial part in the rapid adaptability of the pathogen to host plants and underpins its evolutionary potential.

miRNAs control gene expression in the single-cell alga Chlamydomonas reinhardtii

InterPro, an integrated documentation resource of protein families, domains and functional sites, was created to integrate the major protein signature databases. Currently, it includes PROSITE, Pfam, PRINTS, ProDom, SMART, TIGRFAMs, PIRSF and SUPERFAMILY. Signatures are manually integrated into InterPro entries that are curated to provide biological and functional information. Annotation is provided in an abstract, Gene Ontology mapping and links to specialized databases. New features of InterPro include extended protein match views, taxonomic range information and protein 3D structure data. One of the new match views is the InterPro Domain Architecture view, which shows the domain composition of protein matches. Two new entry types were introduced to better describe InterPro entries: these are active site and binding site. PIRSF and the structure-based SUPERFAMILY are the latest member databases to join InterPro, and CATH and PANTHER are soon to be integrated. InterPro release 8.0 contains 11 007 entries, representing 2573 domains, 8166 families, 201 repeats, 26 active sites, 21 binding sites and 20 post-translational modification sites. InterPro covers over 78% of all proteins in the Swiss-Prot and TrEMBL components of UniProt. The database is available for text- and sequence-based searches via a webserver (http://www.ebi.ac.uk/interpro), and for download by anonymous FTP (ftp://ftp.ebi.ac.uk/pub/databases/interpro).

The Bacterial Enhancer-Dependent ς54(ςN) Transcription Factor

MicroRNAs (miRNAs) in eukaryotes guide post-transcriptional regulation by means of targeted RNA degradation and translational arrest. They are released by a Dicer nuclease as a 21–24-nucleotide RNA duplex from a precursor in which an imperfectly matched inverted repeat forms a partly double-stranded region. One of the two strands is then recruited by an Argonaute nuclease that is the effector protein of the silencing mechanism. Short interfering RNAs (siRNAs), which are similar to miRNAs, are also produced by Dicer but the precursors are perfectly double-stranded RNA. These siRNAs guide post-transcriptional regulation, as with miRNAs, and epigenetic genome modification. Diverse eukaryotes including fungi, plants, protozoans and metazoans produce siRNAs but, until now, miRNAs have not been described in unicellular organisms and it has been suggested that they evolved together with multicellularity in separate plant and animal lineages. Here we show that the unicellular alga Chlamydomonas reinhardtii contains miRNAs, putative evolutionary precursors of miRNAs and species of siRNAs resembling those in higher plants. The common features of miRNAs and siRNAs in an alga and in higher plants indicate that complex RNA-silencing systems evolved before multicellularity and were a feature of primitive eukaryotic cells.

Signatures of adaptation to obligate biotrophy in the Hyaloperonospora arabidopsidis genome.

The initiation of transcription is a complex process involving many different steps. These steps are all potential control points for regulating gene expression, and many have been exploited by bacteria to give rise to sophisticated regulatory mechanisms that allow the cell to adapt to changing

Application of 'next-generation' sequencing technologies to microbial genetics

From Blight to Powdery Mildew Pathogenic effects of microbes on plants have widespread consequences. Witness, for example, the cultural upheavals driven by potato blight in the 1800s. A variety of microbial pathogens continue to afflict crop plants today, driving both loss of yield and incurring the increased costs of control mechanisms. Now, four reports analyze microbial genomes in order to understand better how plant pathogens function (see the Perspective by Dodds). Raffaele et al. (p. 1540) describe how the genome of the potato blight pathogen accommodates transfer to different hosts. Spanu et al. (p. 1543) analyze what it takes to be an obligate biotroph in barley powdery mildew, and Baxter et al. (p. 1549) ask a similar question for a natural pathogen of Arabidopsis. Schirawski et al. (p. 1546) compared genomes of maize pathogens to identify virulence determinants. Better knowledge of what in a genome makes a pathogen efficient and deadly is likely to be useful for improving agricultural crop management and breeding. A group of papers analyzes pathogen genomes to find the roots of virulence, opportunism, and life-style determinants. Many oomycete and fungal plant pathogens are obligate biotrophs, which extract nutrients only from living plant tissue and cannot grow apart from their hosts. Although these pathogens cause substantial crop losses, little is known about the molecular basis or evolution of obligate biotrophy. Here, we report the genome sequence of the oomycete Hyaloperonospora arabidopsidis (Hpa), an obligate biotroph and natural pathogen of Arabidopsis thaliana. In comparison with genomes of related, hemibiotrophic Phytophthora species, the Hpa genome exhibits dramatic reductions in genes encoding (i) RXLR effectors and other secreted pathogenicity proteins, (ii) enzymes for assimilation of inorganic nitrogen and sulfur, and (iii) proteins associated with zoospore formation and motility. These attributes comprise a genomic signature of evolution toward obligate biotrophy.

Uniparental expression of PolIV-dependent siRNAs in developing endosperm of Arabidopsis

New sequencing methods generate data that can allow the assembly of microbial genome sequences in days. With such revolutionary advances in technology come new challenges in methodologies and informatics. In this article, we review the capabilities of high-throughput sequencing technologies and discuss the many options for getting useful information from the data.

Genome Evolution Following Host Jumps in the Irish Potato Famine Pathogen Lineage

Most eukaryotes produce small RNA (sRNA) mediators of gene silencing that bind to Argonaute proteins and guide them, by base pairing, to an RNA target. MicroRNAs (miRNAs) that normally target messenger RNAs for degradation or translational arrest are the best-understood class of sRNAs. However, in Arabidopsis thaliana flowers, miRNAs account for only 5% of the sRNA mass and less than 0.1% of the sequence complexity. The remaining sRNAs form a complex population of more than 100,000 different small interfering RNAs (siRNAs) transcribed from thousands of loci. The biogenesis of most of the siRNAs in Arabidopsis are dependent on RNA polymerase IV (PolIV), a homologue of DNA-dependent RNA polymerase II. A subset of these PolIV-dependent (p4)-siRNAs are involved in stress responses, and others are associated with epigenetic modifications to DNA or chromatin; however, the biological role is not known for most of them. Here we show that the predominant phase of p4-siRNA accumulation is initiated in the maternal gametophyte and continues during seed development. Expression of p4-siRNAs in developing endosperm is specifically from maternal chromosomes. Our results provide the first evidence for a link between genomic imprinting and RNA silencing in plants.

A toolkit for analysing large-scale plant small RNA datasets

From Blight to Powdery Mildew Pathogenic effects of microbes on plants have widespread consequences. Witness, for example, the cultural upheavals driven by potato blight in the 1800s. A variety of microbial pathogens continue to afflict crop plants today, driving both loss of yield and incurring the increased costs of control mechanisms. Now, four reports analyze microbial genomes in order to understand better how plant pathogens function (see the Perspective by Dodds). Raffaele et al. (p. 1540) describe how the genome of the potato blight pathogen accommodates transfer to different hosts. Spanu et al. (p. 1543) analyze what it takes to be an obligate biotroph in barley powdery mildew, and Baxter et al. (p. 1549) ask a similar question for a natural pathogen of Arabidopsis. Schirawski et al. (p. 1546) compared genomes of maize pathogens to identify virulence determinants. Better knowledge of what in a genome makes a pathogen efficient and deadly is likely to be useful for improving agricultural crop management and breeding. A group of papers analyzes pathogen genomes to find the roots of virulence, opportunism, and life-style determinants. Many plant pathogens, including those in the lineage of the Irish potato famine organism Phytophthora infestans, evolve by host jumps followed by specialization. However, how host jumps affect genome evolution remains largely unknown. To determine the patterns of sequence variation in the P. infestans lineage, we resequenced six genomes of four sister species. This revealed uneven evolutionary rates across genomes with genes in repeat-rich regions showing higher rates of structural polymorphisms and positive selection. These loci are enriched in genes induced in planta, implicating host adaptation in genome evolution. Unexpectedly, genes involved in epigenetic processes formed another class of rapidly evolving residents of the gene-sparse regions. These results demonstrate that dynamic repeat-rich genome compartments underpin accelerated gene evolution following host jumps in this pathogen lineage.

PolIVb influences RNA-directed DNA methylation independently of its role in siRNA biogenesis

UNLABELLED Recent developments in high-throughput sequencing technologies have generated considerable demand for tools to analyse large datasets of small RNA sequences. Here, we describe a suite of web-based tools for processing plant small RNA datasets. Our tools can be used to identify micro RNAs and their targets, compare expression levels in sRNA loci, and find putative trans-acting siRNA loci. AVAILABILITY The tools are freely available for use at http://srna-tools.cmp.uea.ac.uk.