Common themes in nutrient acquisition by plant symbiotic microbes, described by the Gene Ontology

A critical function for symbionts is the acquisition of nutrients from their host. Relationships between hosts and symbionts range from biotrophic mutualism to necrotrophic parasitism, with a corresponding range of structures to facilitate nutrient flow between host and symbiont. Here, we review common themes among the nutrient acquisition strategies of a range of plant symbiotic microorganisms, including mutualistic symbionts, biotrophic pathogens that feed from living tissue, necrotrophic pathogens that kill host tissue, and hemibiotrophic pathogens that switch from biotrophy to necrotrophy. We show how Gene Ontology (GO) terms developed by the Plant-Associated Microbe Gene Ontology (PAMGO) Consortium can be used for describing commonalities in nutrient acquisition among diverse plant symbionts. Where appropriate, parallels found among animal symbionts are also highlighted

Tethering of Multi-Vesicular Bodies and the Tonoplast to the Plasma Membrane in Plants

Tethering of the plasma membrane (PM) and many organelles to the endoplasmic reticulum (ER) for communication and lipid exchange has been widely reported. However, despite growing interest in multi-vesicular bodies (MVBs) as possible sources of exosomes, tethering of MVBs to the PM has not been reported. Here we show that MVBs and the vacuolar membrane (tonoplast) could be tethered to the PM (PM-MVB/TP tethering) by artificial protein fusions or bimolecular fluorescence complementation (BiFC) complexes that contain a peripheral membrane protein that binds the PM and also a protein that binds MVBs or the tonoplast. PM-binding proteins capable of participating in PM-MVB/TP tethering included StRem1.3, BIK1, PBS1, CPK21, and the PtdIns(4)-binding proteins FAPP1 and Osh2. MVB/TP-binding proteins capable of participating in tethering included ARA6, ARA7, RHA1, RABG3f, and the PtdIns(3)P-binding proteins Vam7p and Hrs-2xFYVE. BiFC complexes or protein fusions capable of producing PM-MVB/TP tethering were visualized as large well-defined patches of fluorescence on the PM that could displace PM proteins such as AtFlotillin1 and also could displace cytoplasmic proteins such as soluble GFP. Furthermore, we identified paralogous ubiquitin E3 ligase proteins, SAUL1 (AtPUB44), and AtPUB43 that could produce PM-MVB/TP tethering. SAUL1 and AtPUB43 could produce tethering in uninfected tissue when paired with MVB-binding proteins or when their E3 ligase domain was deleted. When Nicotiana benthamiana leaf tissue was infected with Phytophthora capsici, full length SAUL1 and AtPUB43 localized in membrane patches consistent with PM-MVB/TP tethering. Our findings define new tools for studying PM-MVB/TP tethering and its possible role in plant defense.Significance StatementAlthough not previously observed, the tethering of multi-vesicular bodies to the plasma membrane is of interest due to the potential role of this process in producing exosomes in plants. Here we describe tools for observing and manipulating the tethering of multi-vesicular bodies and the tonoplast to the plant plasma membrane, and describe two plant proteins that may naturally regulate this process during infection

Successful Models for Interdisciplinary Graduate Education in the U.S. : Report of the Taskforce on Interdisciplinary Graduate Education Programs

The Taskforce on Interdisciplinary Graduate Education Programs was given the mission to examine interdisciplinary graduate education programs at a range of institutions around the US, and to identify successful models that could inform the revitalization of interdisciplinary graduate education at Oregon State University. The taskforce examined programs at 29 universities of various sizes, including 11 land grant institutions, including site visits to 5 institutions

Protein secretion systems in bacterial-host associations, and their description in the Gene Ontology

Protein secretion plays a central role in modulating the interactions of bacteria with their environments. This is particularly the case when symbiotic bacteria (whether pathogenic, commensal or mutualistic) are interacting with larger host organisms. In the case of Gram-negative bacteria, secretion requires translocation across the outer as well as the inner membrane, and a diversity of molecular machines have been elaborated for this purpose. A number of secreted proteins are destined to enter the host cell (effectors and toxins), and thus several secretion systems include apparatus to translocate proteins across the plasma membrane of the host also. The Plant-Associated Microbe Gene Ontology (PAMGO) Consortium has been developing standardized terms for describing biological processes and cellular components that play important roles in the interactions of microbes with plant and animal hosts, including the processes of bacterial secretion. Here we survey bacterial secretion systems known to modulate interactions with host organisms and describe Gene Ontology terms useful for describing the components and functions of these systems, and for capturing the similarities among the diverse systems

Efficient disruption and replacement of an effector gene in the oomycete Phytophthora sojae using CRISPR/Cas9

Phytophthora sojae is an oomycete pathogen of soybean. As a result of its economic importance, P. sojae has become a model for the study of oomycete genetics, physiology and pathology. The lack of efficient techniques for targeted mutagenesis and gene replacement have long hampered genetic studies of pathogenicity in Phytophthora species. Here, we describe a CRISPR/Cas9 system enabling rapid and efficient genome editing in P. sojae. Using the RXLR effector gene Avr4/6 as a target, we observed that, in the absence of a homologous template, the repair of Cas9-induced DNA double-strand breaks (DSBs) in P. sojae was mediated by non-homologous end-joining (NHEJ), primarily resulting in short indels. Most mutants were homozygous, presumably as a result of gene conversion triggered by Cas9-mediated cleavage of non-mutant alleles. When donor DNA was present, homology-directed repair (HDR) was observed, which resulted in the replacement of Avr4/6 with the NPT II gene. By testing the specific virulence of several NHEJ mutants and HDR-mediated gene replacements in soybean, we have validated the contribution of Avr4/6 to recognition by soybean R gene loci, Rps4 and Rps6, but also uncovered additional contributions to resistance by these two loci. Our results establish a powerful tool for the study of functional genomics in Phytophthora, which provides new avenues for better control of this pathogen.This is the publisher’s final pdf. The published article is copyrighted by the author(s) and published by John Wiley & Sons, Ltd and British Society for Plant Pathology. The published article can be found at: http://onlinelibrary.wiley.com/journal/10.1111/%28ISSN%291364-3703. Supporting information available online at: http://onlinelibrary.wiley.com/doi/10.1111/mpp.12318/suppinfoKeywords: Avr4/6, RXLR effector, Phytophthora sojae, CRISPR/Cas9, gene replacement, oomycetes, genome editin

VMD: a community annotation database for oomycetes and microbial genomes

The VBI Microbial Database (VMD) is a database system designed to host a range of microbial genome sequences. At present, the database contains genome sequence and annotation data of two plant pathogens Phytophthora sojae and Phytophthora ramorum. With the completion of the draft genome sequences of these pathogens in collaboration with the DOE Joint Genome Institute (JGI), we have created this resource to make the sequences publicly available. The genome sequences (95 MB for P.sojae and 65 MB for P.ramorum) were annotated with ∼19 000 and ∼16 000 gene models, respectively. We used two different statistical methods to validate these gene models, Fickett's and a log-likelihood method. Functional annotation of the gene models is based on results from BlastX and InterProScan screens. From the InterProScan results, we could assign putative functions to 17 694 genes in P.sojae and 14 700 genes in P.ramorum. We created an easy-to-use genome browser to view the genome sequence data, which opens to detailed annotation pages for each gene model. A community annotation interface is available for registered community members to add or edit annotations. There are ∼ 1600 gene models for P.sojae and ∼700 models for P.ramorum that have already been manually curated. A toolkit is provided as an additional resource for users to perform a variety of sequence analysis jobs. The database is publicly available at

Common processes in pathogenesis by fungal and oomycete plant pathogens, described with Gene Ontology terms

Plant diseases caused by fungi and oomycetes result in significant economic losses every year. Although phylogenetically distant, the infection processes by these organisms share many common features. These include dispersal of an infectious particle, host adhesion, recognition, penetration, invasive growth, and lesion development. Previously, many of these common processes did not have corresponding Gene Ontology (GO) terms. For example, no GO terms existed to describe processes related to the appressorium, an important structure for infection by many fungi and oomycetes. In this mini-review, we identify common features of the pathogenic processes of fungi and oomycetes and create a pathogenesis model using 256 newly developed and 38 extant GO terms, with an emphasis on the appressorium and signal transduction. This set of standardized GO terms provides a solid base to further compare and contrast the molecular underpinnings of fungal and oomycete pathogenesis

Common and contrasting themes in host cell-targeted effectors from bacterial, fungal, oomycete and nematode plant symbionts described using the Gene Ontology

A wide diversity of plant-associated symbionts, including microbes, produce proteins that can enter host cells, or are injected into host cells in order to modify the physiology of the host to promote colonization. These molecules, termed effectors, commonly target the host defense signaling pathways in order to suppress the defense response. Others target the gene expression machinery or trigger specific modifications to host morphology or physiology that promote the nutrition and proliferation of the symbiont. When recognized by the host's surveillance machinery, which includes cognate resistance (R) gene products, defense responses are engaged to restrict pathogen proliferation. Effectors from diverse symbionts may be delivered into plant cells via varied mechanisms, including whole organism cellular entry (viruses, some bacteria and fungi), type III and IV secretion (in bacteria), physical injection (nematodes and insects) and protein translocation signal sequences (oomycetes and fungi). This mini-review will summarize both similarities and differences in effectors and effector delivery systems found in diverse plant-associated symbionts as well as how these are described with Plant-Associated Microbe Gene Ontology (PAMGO) terms

Zoospore interspecific signaling promotes plant infection by Phytophthora

<p>Abstract</p> <p>Background</p> <p>Oomycetes attack a huge variety of economically and ecologically important plants. These pathogens release, detect and respond to signal molecules to coordinate their communal behaviors including the infection process. When signal molecules are present at or above threshold level, single zoospores can infect plants. However, at the beginning of a growing season population densities of individual species are likely below those required to reach a quorum and produce threshold levels of signal molecules to trigger infection. It is unclear whether these molecules are shared among related species and what their chemistries are.</p> <p>Results</p> <p>Zoospore-free fluids (ZFF) from <it>Phytophthora capsici</it>, <it>P. hydropathica</it>, <it>P. nicotianae </it>(ZFFnic), <it>P. sojae </it>(ZFFsoj) and <it>Pythium aphanidermatum </it>were cross tested for stimulating plant infection in three pathosystems. All ZFFs tested significantly increased infection of <it>Catharanthus roseus </it>by <it>P. nicotianae</it>. Similar cross activities were observed in infection of <it>Lupinus polyphyllus </it>and <it>Glycine max </it>by <it>P. sojae</it>. Only ZFFnic and ZFFsoj cross induced zoospore aggregation at a density of 2 × 10³ml^-1. Pure autoinducer-2 (AI-2), a component in ZFF, caused zoospore lysis of <it>P. nicotianae </it>before encystment and did not stimulate plant infection at concentrations from 0.01 to 1000 μM. <it>P. capsici </it>transformants with a transiently silenced AI-2 synthase gene, ribose phosphate isomerase (<it>RPI</it>), infected <it>Capsicum annuum </it>seedlings at the same inoculum concentration as the wild type. Acyl-homoserine lactones (AHLs) were not detected in any ZFFs. After freeze-thaw treatments, ZFF remained active in promoting plant infection but not zoospore aggregation. Heat treatment by boiling for 5 min also did not affect the infection-stimulating property of ZFFnic.</p> <p>Conclusion</p> <p>Oomycetes produce and use different molecules to regulate zoospore aggregation and plant infection. We found that some of these signal molecules could act in an inter-specific manner, though signals for zoospore aggregation were somewhat restricted. This self-interested cooperation among related species gives individual pathogens of the same group a competitive advantage over pathogens and microbes from other groups for limited resources. These findings help to understand why these pathogens often are individually undetectable until severe disease epidemics have developed. The signal molecules for both zoospore aggregation and plant infection are distinct from AI-2 and AHL.</p