Hierarchical classification of switchgrass genotypes using SSR and chloroplast sequences: ecotypes, ploidies, gene pools, and cultivars.

Switchgrass (Panicum virgatum L.) is an important crop for bioenergy feedstock development. Switchgrass has two main ecotypes: the lowland ecotype being exclusively tetraploid (2n = 4x = 36) and the upland ecotype being mainly tetraploid and octaploid (2n = 8x = 72). Because there is a significant difference in ploidy, morphology, growth pattern, and zone of adaptation between and within the upland and lowland ecotypes, it is important to discriminate switchgrass plants belonging to different genetic pools. We used 55 simple sequence repeats (SSR) loci and six chloroplast sequences to identify patterns of variation between and within 18 switchgrass cultivars representing seven lowland and 11 upland cultivars from different geographic regions and of varying ploidy levels. We report consistent discrimination of switchgrass cultivars into ecotype membership and demonstrate unambiguous molecular differentiation among switchgrass ploidy levels using genetic markers. Also, SSR and chloroplast markers identified genetic pools related to the geographic origin of the 18 cultivars with respect to ecotype, ploidy, and geographical, and cultivar sources. SSR loci were highly informative for cultivar fingerprinting and to classify plants of unknown origin. This classification system is the first step toward developing switchgrass complementary gene pools that can be expected to provide a significant heterotic increase in biomass yield.

Functional studies of the bacterial avirulence protein AvrPto by mutational analysis.

Pseudomonas syringae pathovars expressing avrPto are avirulent on plants expressing the resistance gene Pto. Over 85 mutants of avrPto were generated with multiple strategies, and several assays were used to characterize AvrPto function. Only a core of 95 amino acids of the 164 residues was sufficient for binding Pto in the yeast two-hybrid system. Only nine of 65 mutant proteins of AvrPto with amino acid substitutions, created in planta and in vitro, did not interact with Pto in the Gal4 yeast two-hybrid system, suggesting that AvrPto can tolerate many nonconservative substitutions and still interact with Pto. These nine and 12 additional substitution mutants of AvrPto were characterized further. The ability to elicit a hypersensitive response and the effect on pathogenesis in planta for these 21 mutants of AvrPto were strongly correlated with recognition by Pto in the yeast two-hybrid system. Analyses of two proteins with substitutions H54P or D52G/L65P indicated that these residues may be required for delivery into the host cell and protein stability in the bacterial cytoplasm, respectively. The mutants that no longer interacted with Pto and had modified activities in planta were predicted to have changes in their secondary structure.

An S-locus-related gene in Arabidopsis encodes a functional kinase and produces two classes of transcripts.

ARK1 is a vegetatively expressed receptor protein kinase gene isolated from Arabidopsis thaliana based on its sequence similarity to Brassica genes involved in pollen-stigma signaling and the self-incompatibility response. This paper shows that the kinase domain of ARK1 autophosphorylates on serine and threonine residues when expressed as a recombinant fusion protein. ARK1 produces a 2.9 kb transcript encoding a transmembrane receptor protein kinase and a 1.4 kb transcript encoding the receptor domain alone. Constitutive high-level expression of ARK1 transcripts in transgenic Arabidopsis resulted in severe stunting and also disrupted normal cellular expansion and differentiation.

An Arabidopsis thaliana Gene with Sequence Similarity to the S-Locus Receptor Kinase of Brassica oleracea: Sequence and Expression.

Primary signal transduction plays a vital role in the way plants react to environmental and developmental signals. We report the sequence and expression of a putative receptor kinase gene, ARK1, in Arabidopsis thaliana that may be important in this regard. This Arabidopsis gene encodes a transmembrane protein with a cytoplasmic kinase catalytic domain, a transmembrane region, and an extracellular domain with sequence similarity to the secreted S-locus glycoprotein (SLG) gene of Brassica oleracea. This structure is similar to the S-locus receptor kinase (SRK) gene of Brassica and to the receptor kinase ZmPK1 gene of maize. RNA blots indicate that transcripts accumulate predominantly in leaf tissue, with limited amounts in stem and floral bud tissue and no detectable transcripts accumulating in root tissue. A smaller transcript that could be an alternative transcript of ARK1 also accumulates in leaf tissue. This transcript possibly encodes a secreted SLG-like glycoprotein that lacks transmembrane and kinase domains. The predominantly vegetative expression of ARK1 indicates that this gene is not primarily involved in pollen/pistil interactions in Arabidopsis.

Plants expressing the Pto disease resistance gene confer resistance to recombinant PVX containing the avirulence gene AvrPto.

Elicitation of hypersensitive cell death and induction of plant disease resistance by Pseudomonas syringae pv. tomato (Pst) is dependent on activity of the Pst Hrp secretion system and the gene-for-gene interaction between the tomato resistance gene Pto and the bacterial avirulence gene avrPto. AvrPto was expressed transiently in resistant or susceptible plant lines via a potato virus X (PVX) vector. We found that while PVX is normally virulent on tomato, a PVX derivative expressing avrPto was only capable of infecting plants lacking a functional Pto resistance pathway. Mutations in either the Pto or Prf genes allowed systemic spread of the recombinant virus. These results indicate that recognition of AvrPto by Pto in resistant plant lines triggers a plant defense response that can confer resistance to a viral as well as a bacterial pathogen.