Crassulacean acid metabolism (CAM) is a water-use efficient adaptation of photosynthesis that has evolved independently many times in diverse lineages of flowering plants. We hypothesize that convergent evolution of protein sequence and temporal gene expression underpins the independent emergences of CAM from C3 photosynthesis. To test this hypothesis, we generate a de novo genome assembly and genome-wide transcript expression data for Kalanchoë fedtschenkoi, an obligate CAM species within the core eudicots with a relatively small genome (~260 Mb). Our comparative analyses identify signatures of convergence in protein sequence and re-scheduling of diel transcript expression of genes involved in nocturnal CO2 fixation, stomatal movement, heat tolerance, circadian clock, and carbohydrate metabolism in K. fedtschenkoi and other CAM species in comparison with non-CAM species. These findings provide new insights into molecular convergence and building blocks of CAM and will facilitate CAM-into-C3 photosynthesis engineering to enhance water-use efficiency in crops.

F-box proteins are generally responsible for substrate recognition in the Skp1-Cullin-F-box complexes that are involved in protein degradation via the ubiquitin-26S proteasome pathway. In plants, F-box genes influence a variety of biological processes, such as leaf senescence, branching, self-incompatibility, and responses to biotic and abiotic stresses. The number of F-box genes in Populus (Populus trichocarpa; approximately 320) is less than half that found in Arabidopsis (Arabidopsis thaliana; approximately 660) or Oryza (Oryza sativa; approximately 680), even though the total number of genes in Populus is equivalent to that in Oryza and 1.5 times that in Arabidopsis. We performed comparative genomics analysis between the woody perennial plant Populus and the herbaceous annual plants Arabidopsis and Oryza in order to explicate the functional implications of this large gene family. Our analyses reveal interspecific differences in genomic distribution, orthologous relationship, intron evolution, protein domain structure, and gene expression. The set of F-box genes shared by these species appear to be involved in core biological processes essential for plant growth and development; lineage-specific differences primarily occurred because of an expansion of the F-box genes via tandem duplications in Arabidopsis and Oryza. The number of F-box genes in the newly sequenced woody species Vitis (Vitis vinifera; 156) and Carica (Carica papaya; 139) is similar to that in Populus, supporting the hypothesis that the F-box gene family is expanded in herbaceous annual plants relative to woody perennial plants. This study provides insights into the relationship between the structure and composition of the F-box gene family in herbaceous and woody species and their associated developmental and physiological features.

Leaf area index (LAI) and its seasonal dynamics are key determinants of terrestrial productivity and, therefore, of the response of ecosystems to a rising atmospheric CO(2) concentration. Despite the central importance of LAI, there is very little evidence from which to assess how forest LAI will respond to increasing [CO(2)]. We assessed LAI and related leaf indices of a closed-canopy deciduous forest for 4 years in 25-m-diameter plots that were exposed to ambient or elevated CO(2) (542 ppm) in a free-air CO(2) enrichment (FACE) experiment. LAI of this Liquidambar styraciflua (sweetgum) stand was about 6 and was relatively constant year-to-year, including the 2 years prior to the onset of CO(2) treatment. LAI throughout the 1999-2002 growing seasons was assessed through a combination of data on photosynthetically active radiation (PAR) transmittance, mass of litter collected in traps, and leaf mass per unit area (LMA). There was no effect of [CO(2)] on any expression of leaf area, including peak LAI, average LAI, or leaf area duration. Canopy mass and LMA, however, were significantly increased by CO(2) enrichment. The hypothesized connection between light compensation point (LCP) and LAI was rejected because LCP was reduced by [CO(2)] enrichment only in leaves under full sun, but not in shaded leaves. Data on PAR interception also permitted calculation of absorbed PAR (APAR) and light use efficiency (LUE), which are key parameters connecting satellite assessments of terrestrial productivity with ecosystem models of future productivity. There was no effect of [CO(2)] on APAR, and the observed increase in net primary productivity in elevated [CO(2)] was ascribed to an increase in LUE, which ranged from 1.4 to 2.4 g MJ(-1). The current evidence seems convincing that LAI of non-expanding forest stands will not be different in a future CO(2)-enriched atmosphere and that increases in LUE and productivity in elevated [CO(2)] are driven primarily by functional responses rather than by structural changes. Ecosystem or regional models that incorporate feedbacks on resource use through LAI should not assume that LAI will increase with CO(2) enrichment of the atmosphere.

3-O-caffeoylquinic acid, also known as chlorogenic acid (CGA), functions as an intermediate in lignin biosynthesis in the phenylpropanoid pathway. It is widely distributed among numerous plant species and acts as an antioxidant in both plants and animals. Using GC-MS, we discovered consistent and extreme variation in CGA content across a population of 739 4-yr-old Populus trichocarpa accessions. We performed genome-wide association studies (GWAS) from 917 P. trichocarpa accessions and expression-based quantitative trait loci (eQTL) analyses to identify key regulators. The GWAS and eQTL analyses resolved an overlapped interval encompassing a hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase 2 (PtHCT2) that was significantly associated with CGA and partially characterized metabolite abundances. PtHCT2 leaf expression was significantly correlated with CGA abundance and it was regulated by cis-eQTLs containing W-box for WRKY binding. Among all nine PtHCT homologs, PtHCT2 is the only one that responds to infection by the fungal pathogen Sphaerulina musiva (a Populus pathogen). Validation using protoplast-based transient expression system suggests that PtHCT2 is regulated by the defense-responsive WRKY. These results are consistent with reports of CGA functioning as an antioxidant in response to biotic stress. This study provides insights into data-driven and omics-based inference of gene function in woody species.

BackgroundQTL cloning for the discovery of genes underlying polygenic traits has historically been cumbersome in long-lived perennial plants like Populus. Linkage disequilibrium-based association mapping has been proposed as a cloning tool, and recent advances in high-throughput genotyping and whole-genome resequencing enable marker saturation to levels sufficient for association mapping with no a priori candidate gene selection. Here, multiyear and multienvironment evaluation of cell wall phenotypes was conducted in an interspecific P. trichocarpa x P. deltoides pseudo-backcross mapping pedigree and two partially overlapping populations of unrelated P. trichocarpa genotypes using pyrolysis molecular beam mass spectrometry, saccharification, and/ or traditional wet chemistry. QTL mapping was conducted using a high-density genetic map with 3,568 SNP markers. As a fine-mapping approach, chromosome-wide association mapping targeting a QTL hot-spot on linkage group XIV was performed in the two P. trichocarpa populations. Both populations were genotyped using the 34 K Populus Infinium SNP array and whole-genome resequencing of one of the populations facilitated marker-saturation of candidate intervals for gene identification.ResultsFive QTLs ranging in size from 0.6 to 1.8 Mb were mapped on linkage group XIV for lignin content, syringyl to guaiacyl (S/G) ratio, 5- and 6-carbon sugars using the mapping pedigree. Six candidate loci exhibiting significant associations with phenotypes were identified within QTL intervals. These associations were reproducible across multiple environments, two independent genotyping platforms, and different plant growth stages. cDNA sequencing for allelic variants of three of the six loci identified polymorphisms leading to variable length poly glutamine (PolyQ) stretch in a transcription factor annotated as an ANGUSTIFOLIA C-terminus Binding Protein (CtBP) and premature stop codons in a KANADI transcription factor as well as a protein kinase. Results from protoplast transient expression assays suggested that each of the polymorphisms conferred allelic differences in the activation of cellulose, hemicelluloses, and lignin pathway marker genes.ConclusionThis study illustrates the utility of complementary QTL and association mapping as tools for gene discovery with no a priori candidate gene selection. This proof of concept in a perennial organism opens up opportunities for discovery of novel genetic determinants of economically important but complex traits in plants.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1215-z) contains supplementary material, which is available to authorized users.