James V. Anderson

Transcriptome analysis identifies novel responses and potential regulatory genes involved in seasonal dormancy transitions of leafy spurge (Euphorbia esula L.).

BackgroundDormancy of buds is a critical developmental process that allows perennial plants to survive extreme seasonal variations in climate. Dormancy transitions in underground crown buds of the model herbaceous perennial weed leafy spurge were investigated using a 23 K element cDNA microarray. These data represent the first large-scale transcriptome analysis of dormancy in underground buds of an herbaceous perennial species. Crown buds collected monthly from August through December, over a five year period, were used to monitor the changes in the transcriptome during dormancy transitions.ResultsNearly 1,000 genes were differentially-expressed through seasonal dormancy transitions. Expected patterns of gene expression were observed for previously characterized genes and physiological processes indicated that resolution in our analysis was sufficient for identifying shifts in global gene expression.ConclusionGene ontology of differentially-expressed genes suggests dormancy transitions require specific alterations in transport functions (including induction of a series of mitochondrial substrate carriers, and sugar transporters), ethylene, jasmonic acid, auxin, gibberellic acid, and abscisic acid responses, and responses to stress (primarily oxidative and cold/drought). Comparison to other dormancy microarray studies indicated that nearly half of the genes identified in our study were also differentially expressed in at least two other plant species during dormancy transitions. This comparison allowed us to identify a particular MADS-box transcription factor related to the DORMANCY ASSOCIATED MADS-BOX genes from peach and hypothesize that it may play a direct role in dormancy induction and maintenance through regulation of FLOWERING LOCUS T.

Characterization, expression and function of DORMANCY ASSOCIATED MADS-BOX genes from leafy spurge

DORMANCY ASSOCIATED MADS-BOX (DAM) genes are related to AGAMOUS-LIKE 24 and SHORT VEGETATIVE PHASE genes of arabidopsis and are differentially regulated coordinately with endodormancy induction and release in buds of several perennial plant species. DAM genes were first shown to directly impact endodormancy in peach where a deletion of a series of DAM resulted in loss of endodormancy induction. We have cloned and characterized several MADS box genes from the model perennial weed leafy spurge. Leafy spurge DAM genes are preferentially expressed in shoot tips and buds in response to cold temperatures and day length in a manner that is relative to the level of endodormancy induced by various environmental conditions. Over-expression of one DAM gene in arabidopsis delays flowering. Additionally, we show that at least one DAM gene is differentially regulated by chromatin remodeling. Comparisons of the DAM gene promoters between poplar and leafy spurge have identified several conserved sequences that may be important for their expression patterns in response to dormancy-inducing stimuli.

Gene-based microsatellites for cassava (Manihot esculenta Crantz): prevalence, polymorphisms, and cross-taxa utility

BackgroundCassava (Manihot esculenta Crantz), a starchy root crop grown in tropical and subtropical climates, is the sixth most important crop in the world after wheat, rice, maize, potato and barley. The repertoire of simple sequence repeat (SSR) markers for cassava is limited and warrants a need for a larger number of polymorphic SSRs for germplasm characterization and breeding applications.ResultsA total of 846 putative microsatellites were identified in silico from an 8,577 cassava unigene set with an average density of one SSR every 7 kb. One hundred and ninety-two candidate SSRs were screened for polymorphism among a panel of cassava cultivars from Africa, Latin America and Asia, four wild Manihot species as well as two other important taxa in the Euphorbiaceae, leafy spurge (Euphorbia esula) and castor bean (Ricinus communis). Of 168 markers with clean amplification products, 124 (73.8%) displayed polymorphism based on high resolution agarose gels. Of 85 EST-SSR markers screened, 80 (94.1%) amplified alleles from one or more wild species (M epruinosa, M glaziovii, M brachyandra, M tripartita) whereas 13 (15.3%) amplified alleles from castor bean and 9 (10.6%) amplified alleles from leafy spurge; hence nearly all markers were transferable to wild relatives of M esculenta while only a fraction was transferable to the more distantly related taxa. In a subset of 20 EST-SSRs assessed by fluorescence-based genotyping the number of alleles per locus ranged from 2 to 10 with an average of 4.55 per locus. These markers had a polymorphism information content (PIC) from 0.19 to 0.75 with an average value of 0.55 and showed genetic relationships consistent with existing information on these genotypes.ConclusionA set of 124 new, unique polymorphic EST-SSRs was developed and characterized which extends the repertoire of SSR markers for cultivated cassava and its wild relatives. The markers show high PIC values and therefore will be useful for cultivar identification, taxonomic studies, and genetic mapping. The study further shows that mining ESTs is a highly efficient strategy for polymorphism detection within the cultivated cassava gene pool.

Review: A current review on the regulation of dormancy in vegetative buds

Abstract In this review, we examine current techniques and recent advances directed toward understanding cellular mechanisms involved in controlling dormancy in vegetative propagules. Vegetative propagules (including stems, rhizomes, tubers, bulbs, stolons, creeping roots, etc.) contain axillary and adventitious buds capable of producing new stems/branches under permissive environments. Axillary and adventitious buds are distinct in that axillary buds are formed in the axil of leaves and are responsible for production of lateral shoots (branches). Adventitious buds refer to buds that arise on the plant at places (stems, roots, or leaves) other than leaf axils. Both axillary and adventitious buds generally undergo periods of dormancy. Dormancy has been described as a temporary suspension of visible growth of any plant structure containing a meristem (Lang et al. 1987). Dormancy can be subdivided into three categories: (1) ecodormancy-arrest is under the control of external environmental factors; (2) paradormancy-arrest is under the control of external physiological factors within the plant; and (3) endodormancy-arrest is under the control of internal physiological factors. One common feature in all of these processes is prevention of growth under conditions where growth should otherwise continue. There is growing evidence that lack of growth is due to blockage of cell division resulting from interactions between the signaling pathways controlling dormancy and those controlling the cell cycle. Nomenclature: Abscisic acid, ABA; CDK-activating kinase, CAK; cyclin-dependent protein kinase, CDK; extracellular-signal-regulated kinase, ERK; gibberellic acid, GA; growth factor receptor, GFR; mitogen-activated protein kinase, MAPK; underground adventitious buds, UABs; nuclear export signal, NES; nuclear localization signal, NLS; retinoblastoma, RB; virus-induced gene silencing, VIGS.

Association between seed dormancy and pericarp color is controlled by a pleiotropic gene that regulates abscisic acid and flavonoid synthesis in weedy red rice.

Seed dormancy has been associated with red grain color in cereal crops for a century. The association was linked to qSD7-1/qPC7, a cluster of quantitative trait loci for seed dormancy/pericarp color in weedy red rice. This research delimited qSD7-1/qPC7 to the Os07g11020 or Rc locus encoding a basic helix-loop-helix family transcription factor by intragenic recombinants and provided unambiguous evidence that the association arises from pleiotropy. The pleiotropic gene expressed in early developing seeds promoted expression of key genes for biosynthesis of abscisic acid (ABA), resulting in an increase in accumulation of the dormancy-inducing hormone; activated a conserved network of eight genes for flavonoid biosynthesis to produce the pigments in the lower epidermal cells of the pericarp tissue; and enhanced seed weight. Thus, the pleiotropic locus most likely controls the dormancy and pigment traits by regulating ABA and flavonoid biosynthetic pathways, respectively. The dormancy effect could be eliminated by a heat treatment, but could not be completely overcome by gibberellic acid or physical removal of the seed maternal tissues. The dormancy-enhancing alleles differentiated into two groups basically associated with tropical and temperate ecotypes of weedy rice. Of the pleiotropic effects, seed dormancy could contribute most to the weed adaptation. Pleiotropy prevents the use of the dormancy gene to improve resistance of white pericarp cultivars against pre-harvest sprouting through conventional breeding approaches.

Characterization of an 18,166 EST dataset for cassava (Manihot esculenta Crantz) enriched for drought-responsive genes

Cassava (Manihot esculenta Crantz) is a staple food for over 600 million people in the tropics and subtropics and is increasingly used as an industrial crop for starch production. Cassava has a high growth rate under optimal conditions but also performs well in drought-prone areas and on marginal soils. To increase the tools for understanding and manipulating drought tolerance in cassava, we generated expressed sequence tags (ESTs) from normalized cDNA libraries prepared from dehydration-stressed and control well-watered tissues. Analysis of a total of 18,166 ESTs resulted in the identification of 8,577 unique gene clusters (5,383 singletons and 3,194 clusters). Functional categories could be assigned to 63% of the unigenes, while another ∼11% were homologous to hypothetical genes with unclear functions. The remaining ∼26% were not significantly homologous to sequences in public databases suggesting that some may be novel and putatively specific to cassava. The dehydration-stressed library uncovered numerous ESTs with recognized roles in drought-responses, including those that encode late-embryogenesis-abundant proteins thought to confer osmoprotective functions during water stress, transcription factors, heat-shock proteins as well as proteins involved in signal transduction and oxidative stress. The unigene clusters were screened for short tandem repeats for further development as microsatellite markers. A total of 592 clusters contained 646 repeats, representing 3.3% of the ESTs queried. The ESTs presented here are the first dehydration stress transcriptome of cassava and can be utilized for the development of microarrays and gene-derived molecular markers to further dissect the molecular basis of drought tolerance in cassava.

Genomics of Compositae weeds: EST libraries, microarrays, and evidence of introgression.

PREMISE OF STUDY Weeds cause considerable environmental and economic damage. However, genomic characterization of weeds has lagged behind that of model plants and crop species. Here we describe the development of genomic tools and resources for 11 weeds from the Compositae family that will serve as a basis for subsequent population and comparative genomic analyses. Because hybridization has been suggested as a stimulus for the evolution of invasiveness, we also analyze these genomic data for evidence of hybridization. METHODS We generated 22 expressed sequence tag (EST) libraries for the 11 targeted weeds using Sanger, 454, and Illumina sequencing, compared the coverage and quality of sequence assemblies, and developed NimbleGen microarrays for expression analyses in five taxa. When possible, we also compared the distributions of Ks values between orthologs of congeneric taxa to detect and quantify hybridization and introgression. RESULTS Gene discovery was enhanced by sequencing from multiple tissues, normalization of cDNA libraries, and especially greater sequencing depth. However, assemblies from short sequence reads sometimes failed to resolve close paralogs. Substantial introgression was detected in Centaurea and Helianthus, but not in Ambrosia and Lactuca. CONCLUSIONS Transcriptome sequencing using next-generation platforms has greatly reduced the cost of genomic studies of nonmodel organisms, and the ESTs and microarrays reported here will accelerate evolutionary and molecular investigations of Compositae weeds. Our study also shows how ortholog comparisons can be used to approximately estimate the genome-wide extent of introgression and to identify genes that have been exchanged between hybridizing taxa.

Molecular Analysis of Signals Controlling Dormancy and Growth in Underground Adventitious Buds of Leafy Spurge

Dormancy and subsequent regrowth of adventitious buds is a critical physiological process for many perennial plants. We have used the expression of hormone and cell cycle-responsive genes as markers to follow this process in leafy spurge (Euphorbia esula). In conjunction with earlier studies, we show that loss of mature leaves results in decreased sugar levels and increased gibberellin perception in underground adventitious buds. Gibberellin is sufficient for induction of S phase-specific but not M phase-specific gene expression. Loss of both apical and axillary buds or inhibition of polar auxin transport did not result in induction of S phase- or M phase-specific gene expression. Loss of polar auxin transport was necessary for continuation of the cell cycle and further bud development if the S phase was previously initiated.

Potential model weeds to study genomics, ecology, and physiology in the 21st century

Abstract Plant model systems have contributed greatly to the dramatic progress in understanding the fundamental aspects of plant biology. Using model weeds will also help facilitate focused funding and research in the weed science community. Criteria for developing model weeds require attention to weedy characteristics that impart economic losses and a wide geographic distribution, attributes that present the potential for political and scientific support. Expressed sequence tag (EST) databases for model weeds are the most practical approach to identifying new genes and obtaining data on the gene expression underlying weedy characteristics. Weeds such as Canada thistle, eastern black nightshade, johnsongrass, jointed goatgrass, leafy spurge, waterhemp, and weedy rice are proposed as model systems. Nomenclature: Canada thistle, Cirsium arvense (L.) Scop CIRAR; common waterhemp, Amaranthus rudis Sauer AMATA; eastern black nightshade, Solanum ptycanthum Dun. SOLPT; johnsongrass, Sorghum halepense (L.) Pers SORHA; jointed goatgrass, Aegilops cylindrica Host. AEGCY; leafy spurge, Euphorbia esula L. EUPES; red rice (weedy rice), Oryza sativa L. ORYSA; tall waterhemp, Amaranthus tuberculatus (Moq.) J. D. Sauer AMATU.

Characterization of an EST Database for the Perennial Weed Leafy Spurge: An Important Resource for Weed Biology Research

Abstract Genomics programs in the weed science community have not developed as rapidly as that of other crop, horticultural, forestry, and model plant systems. Development of genomic resources for selected model weeds are expected to enhance our understanding of weed biology, just as they have in other plant systems. In this report, we describe the development, characteristics, and information gained from an expressed sequence tag (EST) database for the perennial weed leafy spurge. ESTs were obtained using a normalized cDNA library prepared from a comprehensive collection of tissues. During the EST characterization process, redundancy was minimized by periodic subtractions of the normalized cDNA library. A sequencing success rate of 88% yielded 45,314 ESTs with an average read length of 671 nucleotides. Using bioinformatic analysis, the leafy spurge EST database was assembled into 23,472 unique sequences representing 19,015 unigenes (10,293 clusters and 8,722 singletons). Blast similarity searches to the GenBank nonredundant protein database identified 18,186 total matches, of which 14,205 were nonredundant. These data indicate that 77.4% of the 23,472 unique sequences and 74.7% of the 19,015 unigenes are similar to other known proteins. Further bioinformatics analysis indicated that 2,950, or 15.5%, of the unigenes have previously not been identified suggesting that some may be novel to leafy spurge. Functional classifications assigned to leafy spurge unique sequences using Munich Information Center for Protein or Gene Ontology were proportional to functional classifications for genes of arabidopsis, with the exception of unclassified or unknowns and transposable elements which were significantly reduced in leafy spurge. Although these EST resources have been developed for the purpose of constructing high-density leafy spurge microarrays, they are already providing valuable information related to sugar metabolism, cell cycle regulation, dormancy, terpenoid secondary metabolism, and flowering. Nomenclature: Leafy spurge, Euphorbia esula L. EPHES, arabidopsis, Arabidopsis thaliana (L.) Heynh