James H. Westwood

The evolution of parasitism in plants

The multiple independent origins of plant parasitism suggest that numerous ancestral plant lineages possessed the developmental flexibility to meet the requirements of a parasitic life style, including such adaptations as the ability to recognize host plants, form an invasive haustorium, and regulate the transfer of nutrients and other molecules between two different plants. In this review, we focus on the Orobanchaceae, which are unique among the parasitic plants in that extant member species include the full range of host dependence from facultative to obligate parasites. The recent emergence of genomic resources for these plants should provide new insights into parasitic plant evolution and enable the development of novel genetic strategies for controlling parasitic weeds.

Functional Analysis of a Predicted Flavonol Synthase Gene Family in Arabidopsis

The genome of Arabidopsis (Arabidopsis thaliana) contains five sequences with high similarity to FLAVONOL SYNTHASE1 (AtFLS1), a previously characterized flavonol synthase gene that plays a central role in flavonoid metabolism. This apparent redundancy suggests the possibility that Arabidopsis uses multiple isoforms of FLS with different substrate specificities to mediate the production of the flavonols, quercetin and kaempferol, in a tissue-specific and inducible manner. However, biochemical and genetic analysis of the six AtFLS sequences indicates that, although several of the members are expressed, only AtFLS1 encodes a catalytically competent protein. AtFLS1 also appears to be the only member of this group that influences flavonoid levels and the root gravitropic response in seedlings under nonstressed conditions. This study showed that the other expressed AtFLS sequences have tissue- and cell type-specific promoter activities that overlap with those of AtFLS1 and encode proteins that interact with other flavonoid enzymes in yeast two-hybrid assays. Thus, it is possible that these “pseudogenes” have alternative, noncatalytic functions that have not yet been uncovered.

Cross-Species Translocation of mRNA from Host Plants into the Parasitic Plant Dodder

An intriguing new paradigm in plant biology is that systemically mobile mRNAs play a role in coordinating development. In this process, specific mRNAs are loaded into the phloem transport stream for translocation to distant tissues, where they may impact on developmental processes. However, despite its potential significance for plant growth regulation, mRNA trafficking remains poorly understood and challenging to study. Here, we show that phloem-mobile mRNAs can also traffic between widely divergent species from a host to the plant parasite lespedeza dodder (Cuscuta pentagona Engelm.). Reverse transcription-polymerase chain reaction and microarray analysis were used to detect specific tomato (Lycopersicon esculentum Mill.) transcripts in dodder grown on tomato that were not present in control dodder grown on other host species. Foreign transcripts included LeGAI, which has previously been shown to be translocated in the phloem, as well as nine other transcripts not reported to be mobile. Dodders are parasitic plants that obtain resources by drawing from the phloem of a host plant and have joint plasmodesmata with host cortical cells. Although viruses are known to move between dodder and its hosts, translocation of endogenous plant mRNA has not been reported. These results point to a potentially new level of interspecies communication, and raise questions about the ability of parasites to recognize, use, and respond to transcripts acquired from their hosts.

A New Mutation in Plant Als Confers Resistance to Five Classes of Als-inhibiting Herbicides

Abstract Experiments were conducted to evaluate a biotype of smooth pigweed that had survived applications of sulfonylurea (SU) and imidazolinone (IMI) herbicides in a single season. The source field had a history of repeated acetolactate synthase (ALS)-inhibiting herbicide use over several years. Whole-plant response experiments evaluated the resistant (R11) biotype and an ALS-inhibitor susceptible (S) smooth pigweed biotype to herbicides from the SU, IMI, pyrimidinylthiobenzoate (PTB), and triazolopyrimidine sulfonanilide (TP) chemical families. The R11 biotype exhibited 60- to 3,200-fold resistance to all four ALS-inhibiting herbicide chemistries compared with the S biotype. Nucleotide sequence comparison of Als genes from R11 and S biotypes revealed a single nucleotide difference that resulted in R11 having an amino acid substitution of aspartate to glutamate at position 376, as numbered relative to the protein sequence of mouseearcress. This is the first report of an amino acid substitution at this position of an Als gene isolated from a field-selected weed biotype. To verify the role of this mutation in herbicide resistance, the Als gene was cloned and expressed in Arabidopsis. Transgenic Arabidopsis expressing this Als gene exhibited resistance to SU, IMI, PTB, TP, and sulfonylaminocarbonyltriazolinone ALS-inhibiting herbicide classes. Nomenclature: Chlorimuron cloransulam imazethapyr propoxycarbazone pyrithiobac thifensulfuron smooth pigweed, Amaranthus hybridus L. AMACH mouseearcress, Arabidopsis thaliana (L.) Heynh. ARBTH.

Genomic-scale exchange of mRNA between a parasitic plant and its hosts

Strangleweed shares too much information Because RNA normally functions within an individual cell, we generally think that we keep our RNAs to ourselves. Kim et al. now show that the parasitic dodder plant breaks that rule. When dodder attacks a host plant, it opens up a conduit through which messenger and perhaps other regulatory RNAs are exchanged between parasite and host. Because a single dodder plant can attack multiple hosts, such exchanges may underlie instances of genes transferring between species. Science, this issue p. 808 Strangleweed (Cuscuta pentagona) can exchange large numbers of messenger RNAs with different host plants. Movement of RNAs between cells of a single plant is well documented, but cross-species RNA transfer is largely unexplored. Cuscuta pentagona (dodder) is a parasitic plant that forms symplastic connections with its hosts and takes up host messenger RNAs (mRNAs). We sequenced transcriptomes of Cuscuta growing on Arabidopsis and tomato hosts to characterize mRNA transfer between species and found that mRNAs move in high numbers and in a bidirectional manner. The mobile transcripts represented thousands of different genes, and nearly half the expressed transcriptome of Arabidopsis was identified in Cuscuta. These findings demonstrate that parasitic plants can exchange large proportions of their transcriptomes with hosts, providing potential mechanisms for RNA-based interactions between species and horizontal gene transfer.

Convergent evolution of strigolactone perception enabled host detection in parasitic plants

How plant parasites evolved to find hosts The seeds of parasitic plants need to be able to sense their hosts presence to germinate at the correct time and in the correct place. This is done through the detection of plant hormones, strigolactones. However, the origin of this sensory system is unknown. Conn et al. investigated the diversity of strigolactone receptors in multiple lineages of parasitic plants and their close relatives. They found a greater copy number and accelerated evolution in parasitic plants as compared with nonparasitic relatives. Functional analyses of parasitic plant strigolactone receptors in transgenic Arabidopsis suggested that convergent evolution has occurred to allow the parasitic plants to detect their hosts. Science, this issue p. 540 Obligate parasitic Orobanchaceae plants germinate after sensing strigolactones exuded from host roots. Obligate parasitic plants in the Orobanchaceae germinate after sensing plant hormones, strigolactones, exuded from host roots. In Arabidopsis thaliana, the α/β-hydrolase D14 acts as a strigolactone receptor that controls shoot branching, whereas its ancestral paralog, KAI2, mediates karrikin-specific germination responses. We observed that KAI2, but not D14, is present at higher copy numbers in parasitic species than in nonparasitic relatives. KAI2 paralogs in parasites are distributed into three phylogenetic clades. The fastest-evolving clade, KAI2d, contains the majority of KAI2 paralogs. Homology models predict that the ligand-binding pockets of KAI2d resemble D14. KAI2d transgenes confer strigolactone-specific germination responses to Arabidopsis thaliana. Thus, the KAI2 paralogs D14 and KAI2d underwent convergent evolution of strigolactone recognition, respectively enabling developmental responses to strigolactones in angiosperms and host detection in parasites.

Characterization of the Orobanche–Arabidopsis system for studying parasite–host interactions

Abstract Parasitization by Orobanche is a complex process, one that is mediated by host-derived chemical signals that control parasite seed germination and haustorium initiation and one that ultimately results in the union of two plant species. Experiments were conducted to characterize Orobanche parasitization of the model plant Arabidopsis thaliana and to begin to explore the role of host flavonoid metabolism in the interaction. Arabidopsis thaliana stimulated seed germination and allowed tubercle development of O. aegyptiaca and O. ramosa but did not significantly stimulate seeds of O. crenata, O. minor, or O. cernua. However, if Orobanche seeds were artificially stimulated, O. crenata and O. minor successfully established tubercles on A. thaliana. When compared to the recognized crop hosts, Daucus carota and Nicotiana tabacum, A. thaliana stimulated less O. aegyptiaca germination but allowed for formation of equivalent numbers of tubercles. These findings indicate that A. thaliana is not a large-scale producer of germination stimulant but is highly susceptible to the parasite once Orobanche seeds have germinated. Experiments comparing wild-type A. thaliana plants to mutant lines deficient in flavonoid biosynthesis revealed no differences in the ability to stimulate germination or to allow tubercle formation, indicating that host flavonoid production is not essential for Orobanche parasitization. The results of this work support the use of A. thaliana as a valuable host in understanding Orobanche parasitization. Nomenclature: Arabidopsis thaliana L. Heynh., ARBTH, mouseearcress; Daucus carota L. ‘Danver half long’, DAUCS, carrot; Nicotiana tabacum L. ‘Coker 319’, NIOTA, tobacco; Orobanche aegyptiaca Pers., ORAAE, Egyptian broomrape; Orobanche cernua Loefl., ORACE, nodding broomrape; Orobanche crenata Forsk., ORACR, crenate broomrape; Orobanche minor Smith, ORAMI, small broomrape; Orobanche ramosa L., ORARA, branched broomrape.

RNA translocation between parasitic plants and their hosts

Recent research indicates that RNA translocation occurs between certain parasitic plant species and their hosts. The movement of at least 27 mRNAs has been demonstrated between hosts and Cuscuta pentagona Engelm., with the largest proportion of these being regulatory genes. Movement of RNAi signals has been documented from hosts to the parasites Triphysaria versicolor (Frisch & CA Mey) and Orobanche aegyptiaca (Pers.), demonstrating that the regulation of genes in one species can be influenced by transfer of RNA signals through a parasitic association. This review considers the implications of these findings in light of present understanding of host-parasite connections and the growing body of evidence that RNAs are able to act as signal molecules that convey regulatory information in a cell- and tissue-specific manner. Together, this suggests that parasitic plants can exchange RNAs with their hosts, and that this may be part of the coordinated growth and development that occurs during the process of parasitism. This phenomenon offers promise for new insights into parasitic plants, and new opportunities for the control of parasitic weeds.

Expression of a defense-related 3-hydroxy-3-methylglutaryl CoA reductase gene in response to parasitization by Orobanche spp

Orobanche spp. are angiosperms that live parasitically on the roots of other plants, and are capable of significantly reducing the yield and quality of their crop hosts. We have demonstrated that parasitization by Orobanche induces expression of hmg2, a defense-related isogene of 3-hydroxy-3-methylglutaryl CoA reductase (HMGR) in tobacco. Transgenic tobacco plants expressing a construct containing 2.3 kb of the tomato hmg2 gene promoter fused to the beta-glucuronidase (GUS) reporter gene were parasitized by O. aegyptiaca. Expression of the hmg2:GUS construct was detected within 1 day following penetration of the host root by the O. aegyptiaca radicle and was localized to the region immediately around the site of parasite invasion. This expression continued and intensified over the course of O. aegyptiaca development. In addition, the hmg2:GUS expression was induced by secondary parasitization, where secondary roots of O. aegyptiaca contacted the host root at a distance from the primary attachment site. This GUS expression was specific to plants containing the hmg2:GUS construct, and was not observed in control plants transformed with a construct of the cauliflower mosaic virus 35S promoter fused to the GUS gene. These results indicate that Orobanche parasitization initiates rapid and sustained induction of a defense-related gene in the host root.

The Parasitic Plant Genome Project: New Tools for Understanding the Biology of Orobanche and Striga

Abstract The Parasitic Plant Genome Project has sequenced transcripts from three parasitic species and a nonparasitic relative in the Orobanchaceae with the goal of understanding genetic changes associated with parasitism. The species studied span the trophic spectrum from free-living nonparasite to obligate holoparasite. Parasitic species used were Triphysaria versicolor, a photosynthetically competent species that opportunistically parasitizes roots of neighboring plants; Striga hermonthica, a hemiparasite that has an obligate need for a host; and Orobanche aegyptiaca, a holoparasite with absolute nutritional dependence on a host. Lindenbergia philippensis represents the closest nonparasite sister group to the parasitic Orobanchaceae and was included for comparative purposes. Tissues for transcriptome sequencing from each plant were gathered to identify expressed genes for key life stages from seed conditioning through anthesis. Two of the species studied, S. hermonthica and O. aegyptiaca, are economically important weeds and the data generated by this project are expected to aid in research and control of these species and their relatives. The sequences generated through this project will provide an abundant resource of molecular markers for understanding population dynamics, as well as provide insight into the biology of parasitism and advance progress toward understanding parasite virulence and host resistance mechanisms. In addition, the sequences provide important information on target sites for herbicide action or other novel control strategies such as trans-specific gene silencing. Nomenclature: Egyptian broomrape, Orobanche aegyptiaca (Pers.) (Syn. Phelipanche aegyptiaca) ORAAE; Lindenbergia philippensis (Cham. & Schltdl.) Benth. LINPH; yellowbeak owls-clover, Triphysaria versicolor (Fisch. & C.A. Mey) TRVEV; purple witchweed, Striga hermonthica, (Del.) Benth. STRHE.