Molecular Characterization of Quinate and Shikimate Metabolism in Populus trichocarpa

The shikimate pathway leads to the biosynthesis of aromatic amino acids essential for protein biosynthesis and the production of a wide array of plant secondary metabolites. Among them, quinate is an astringent feeding deterrent that can be formed in a single step reaction from 3-dehydroquinate catalyzed by quinate dehydrogenase (QDH). 3-Dehydroquinate is also the substrate for shikimate biosynthesis through the sequential actions of dehydroquinate dehydratase (DQD) and shikimate dehydrogenase (SDH) contained in a single protein in plants. The reaction mechanism of QDH resembles that of SDH. The poplar genome encodes five DQD/SDH-like genes (Poptr1 to Poptr5), which have diverged into two distinct groups based on sequence analysis and protein structure prediction. In vitro biochemical assays proved that Poptr1 and -5 are true DQD/SDHs, whereas Poptr2 and -3 instead have QDH activity with only residual DQD/SDH activity. Poplar DQD/SDHs have distinct expression profiles suggesting separate roles in protein and lignin biosynthesis. Also, the QDH genes are differentially expressed. In summary, quinate (secondary metabolism) and shikimate (primary metabolism) metabolic activities are encoded by distinct members of the same gene family, each having different physiological functions.     

Biosynthesis of salicylic acid in plants

Salicylic acid (SA) is an important signal molecule in plants. Two pathways of SA biosynthesis have been proposed in plants. Biochemical studies using isotope feeding have suggested that plants synthesize SA from cinnamate produced by the activity of phenylalanine ammonia lyase (PAL). Silencing of PAL genes in tobacco or chemical inhibition of PAL activity in Arabidopsis, cucumber and potato reduces pathogen-induced SA accumulation. Genetic studies, on the other hand, indicate that the bulk of SA is produced from isochorismate. In bacteria, SA is synthesized from chorismate through two reactions catalyzed by isochorismate synthase (ICS) and isochorismate pyruvate lyase (IPL). Arabidopsis contains two ICS genes but has no gene encoding proteins similar to the bacterial IPL. Thus, how SA is synthesized in plants is not fully elucidated. Two recently identified Arabidopsis genes, PBS3 and EPS1, are important for pathogen-induced SA accumulation. PBS3 encodes a member of the acyl-adenylate/thioester-forming enzyme family and EPS1 encodes a member of the BAHD acyltransferase superfamily. PBS3 and EPS1 may be directly involved in the synthesis of an important precursor or regulatory molecule for SA biosynthesis. The pathways and regulation of SA biosynthesis in plants may be more complicated than previously thought.Key words: salicylic acid biosynthesis, isochorismate synthase, phenylalanine ammonia lyase     

Integrative transcriptome analysis identifies new oxidosqualene cyclase genes involved in ginsenoside biosynthesis in Jilin ginseng

BackgroundJilin ginseng, Panax ginseng, is a valuable medicinal herb whose ginsenosides are its major bioactive components. The ginseng oxidosqualene cyclase (PgOSC) gene family is known to play important roles in ginsenoside biosynthesis, but few members of the gene family have been functionally studied.MethodsThe PgOSC gene family has been studied by an integrated analysis of gene expression-ginsenoside content correlation, gene mutation-ginsenoside content association and gene co-expression network, followed by functional analysis through gene regulation.ResultsWe found that five of the genes in the PgOSC gene family, including two published ginsenoside biosynthesis genes and three new genes, were involved in ginsenoside biosynthesis. Not only were the expressions of these genes significantly correlated with ginsenoside contents, but also their nucleotide mutations significantly influenced ginsenoside contents. These results were further verified by regulation analysis of the genes by methyl jasmonate (MeJA) in ginseng hairy roots. Four of these five PgOSC genes were mapped to the ginsenoside biosynthesis pathway. These PgOSC genes expressed differently across tissues, but relatively consistent across developmental stages. These PgOSC genes formed a single co-expression network with those published ginsenoside biosynthesis genes, further confirming their roles in ginsenoside biosynthesis. When the network varied, ginsenoside biosynthesis was significantly influenced, thus revealing the molecular mechanism of ginsenoside biosynthesis.ConclusionAt least five of the PgOSC genes, including the three newly identified and two published PgOSC genes, are involved in ginsenoside biosynthesis. These results provide gene resources and knowledge essential for enhanced research and applications of ginsenoside biosynthesis in ginseng.     

The MIXTA-like Transcription factor MYB16 is a major regulator of cuticle formation in vegetative organs

Cuticle secreted on the surface of the epidermis of aerial organs protects plants from the external environment. We recently found that Arabidopsis MIXTA-like R2R3-MYB family members MYB16 and MYB106 regulate cuticle formation in reproductive organs and trichomes. However, the artificial miRNA (amiRNA)-mediated knockdown plants showed no clear phenotypic abnormality in vegetative tissues. In this study, we used RNA interference (RNAi) targeting MYB16 to produce plants with reduced expression of both MYB16 and MYB106. The rosette leaves of RNAi plants showed more severe permeable cuticle phenotypes than the myb106 mutants expressing the MYB16 amiRNA in the previous study. The RNAi plants also showed reduced expression of cuticle biosynthesis genes LACERATA and ECERIFERUM1. By contrast, expression of a gain-of-function MYB16 construct induced over-accumulation of waxy substances on leaves. These results suggest that MYB16 functions as a major regulator of cuticle formation in vegetative organs, in addition to its effect in reproductive organs and trichomes.     

Species-Specific Expansion and Molecular Evolution of the 3-hydroxy-3-methylglutaryl Coenzyme A Reductase (HMGR) Gene Family in Plants

The terpene compounds represent the largest and most diverse class of plant secondary metabolites which are important in plant growth and development. The 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR; EC 1.1.1.34) is one of the key enzymes contributed to terpene biosynthesis. To better understand the basic characteristics and evolutionary history of the HMGR gene family in plants, a genome-wide analysis of HMGR genes from 20 representative species was carried out. A total of 56 HMGR genes in the 14 land plant genomes were identified, but no genes were found in all 6 algal genomes. The gene structure and protein architecture of all plant HMGR genes were highly conserved. The phylogenetic analysis revealed that the plant HMGRs were derived from one ancestor gene and finally developed into four distinct groups, two in the monocot plants and two in dicot plants. Species-specific gene duplications, caused mainly by segmental duplication, led to the limited expansion of HMGR genes in Zea mays, Gossypium raimondii, Populus trichocarpa and Glycine max after the species diverged. The analysis of Ka/Ks ratios and expression profiles indicated that functional divergence after the gene duplications was restricted. The results suggested that the function and evolution of HMGR gene family were dramatically conserved throughout the plant kingdom.     

Genome-wide analysis and expression profiling of PP2C clade D under saline and alkali stresses in wild soybean and <Emphasis Type="Italic">Arabidopsis</Emphasis>

Protein phosphatase 2Cs (PP2Cs) belong to the largest protein phosphatase family in plants. Some members have been described as being negative modulators of plant growth and development, as well as responses to hormones and environmental stimuli. However, little is known about the members of PP2C clade D, which may be involved in the regulation of signaling pathways, especially in response to saline and alkali stresses. Here, we identified 13 PP2C orthologs from the wild soybean (Glycine soja) genome. We examined the sequence characteristics, chromosome locations and duplications, gene structures, and promoter cis-elements of the PP2C clade D genes in Arabidopsis and wild soybean. Our results showed that GsPP2C clade D (GsAPD) genes exhibit more gene duplications than AtPP2C clade D genes. Plant hormone and abiotic stress-responsive elements were identified in the promoter regions of most PP2C genes. Moreover, we investigated their expression patterns in roots, stems, and leaves. Quantitative real-time PCR analyses revealed that the expression levels of representative GsPP2C and AtPP2C clade D genes were significantly influenced by alkali and salt stresses, suggesting that these genes might be associated with or directly involved in the relevant stress signaling pathways. Our results established a foundation for further functional characterization of PP2C clade D genes in the future.     

miR482 Regulation of NBS-LRR Defense Genes during Fungal Pathogen Infection in Cotton

In this study, we characterized the miR482 family in cotton using existing small RNA datasets and the recently released draft genome sequence of Gossypium raimondii, a diploid cotton species whose progenitor is the putative contributor of the D_t (representing the D genome of tetraploid) genome of the cultivated tetraploid cotton species G. hirsutum and G. barbadense. Of the three ghr-miR482 members reported in G. hirsutum, ghr-miR482a has no homolog in G. raimondii, ghr-miR482b and ghr-miR482c each has a single homolog in G. raimondii. Gra-miR482d has five homologous loci (gra-miR482d, f-i) in G. raimondii and also exists in G. hirsutum (ghr-miR482d). A variant, miR482.2 that is a homolog of miR2118 in other species, is produced from several GHR-MIR482 loci in G. hirsutum. Approximately 12% of the G. raimondii NBS-LRR genes were predicted targets of various members of the gra-miR482 family. Based on the rationale that the regulatory relationship between miR482 and NBS-LRR genes will be conserved in G. raimondii and G. hirsutum, we investigated this relationship using G. hirsutum miR482 and G. raimondii NBS-LRR genes, which are not currently available in G. hirsutum. Ghr-miR482/miR482.2-mediated cleavage was confirmed for three of the four NBS-LRR genes analysed. As in tomato, miR482-mediated cleavage of NBS-LRR genes triggered production of phased secondary small RNAs in cotton. In seedlings of the susceptible cultivar Sicot71 (G. hirsutum) infected with the fungal pathogen Verticillium dahliae, the expression levels of ghr-miR482b/miR482b.2, ghr-miR482c and ghr-miR482d.2 were down-regulated, and several NBS-LRR targets of ghr-miR482c and ghr-miR482d were up-regulated. These results imply that, like tomato plants infected with viruses or bacteria, cotton plants are able to induce expression of NBS-LRR defence genes by suppression of the miRNA-mediated gene silencing pathway upon fungal pathogen attack.     

Influence of soil properties on the distribution of Deschampsia antarctica on King George Island, Maritime Antarctica

The extremely cold and infertile Antarctic is one of the harshest terrestrial ecosystems for the growth of vegetation, except for the grass species Deschampsia antarctica. We examined the main soil variables that determine the distribution of D. antarctica in King George Island by using Bayesian analysis of variance and regression methods. This study compared the density of D. antarctica between 2 sites; the density remained relatively stable at site 1, whereas it severely decreased in site 2 over a period of 3?years. Although site 2 showed better soil conditions for the growth of D. antarctica such as organic matter content, available phosphorus, NO₃-N, and extractable cations, its poor drainage and low soil pH may affected the survival of D. antarctica by altering nutrition availability and inhibiting root respiration. Poisson analysis of covariance showed that the early melting of snow was also an important factor in the distribution of D. antarctica. The results also showed that seabirds and mammals might have greatly influenced the distribution of the grass species in King George Island by transferring nutrients from the sea onto land; thus, changing the chemical characteristics of the soil.     

Adaptation of the seed reproduction system to conditions of Maritime Antarctic in <Emphasis Type="Italic">Deschampsia antarctica</Emphasis> E. Desv.

Deschampsia antarctica3 E. Desv. is one of the two flowering plants that, along with Colobanthus quitensis (Kunth) Bartl., was able to settle the ice-free areas of Antarctica. In order to identify the possible adaptations of the D. antarctica reproductive system to adverse environmental conditions, comparative cytoembryological analysis of plants of this species growing on the Antarctic Peninsula with plants of the closely related species D. beringensis Hult. from the Kamchatka Peninsula was conducted. It was found that both species are characterized by sexual mode of reproduction, equal size of pollen grains (25.5 ± 2.2 and 26.2 ± 1.9 μm, respectively), same features of the embryo sac structure, and emryo- and endospermogenesis. Interspecies differences have been found in mature embryo sac size (326.8 ± 12.8 and 161.7 ± 10.4 μm), pollen sterility percentage (86.1 ± 8.9 and 35.3 ± 9.2%), and quantity of pollen in the anthers (140 ± 15.3 and 1578 ± 88.6). Possible causes and significance of these differences are discussed. No unique adaptations of seed reproduction system that are inherent exclusively to D. antarctica were found. The D. antarctica reproduction strategy is based on the combination of autogamy (and its extreme form cleistogamy) with production of excess pollen quantity for its mode of pollination.     

Alkaline ceramidase family: The first two decades

Ceramidases are a group of enzymes that catalyze the hydrolysis of ceramide, dihydroceramide, and phytoceramide into sphingosine (SPH), dihydrosphingosine (DHS), and phytosphingosine (PHS), respectively, along with a free fatty acid. Ceramidases are classified into the acid, neutral, and alkaline ceramidase subtypes according to the pH optima for their catalytic activity. YPC1 and YDC1 were the first alkaline ceramidase genes to be identified and cloned from the yeast Saccharomyces cerevisiae two decades ago. Subsequently, alkaline ceramidase genes were identified from other species, including one Drosophila melanogaster ACER gene (Dacer), one Arabidopsis thaliana ACER gene (AtACER), three Mus musculus ACER genes (Acer1, Acer2, and Acer3), and three Homo sapiens ACER genes (ACER1, ACER2, and ACER3). The protein products of these genes constitute a large protein family, termed the alkaline ceramidase (ACER) family. All the biochemically characterized members of the ACER family are integral membrane proteins with seven transmembrane segments in the Golgi complex or endoplasmic reticulum, and they each have unique substrate specificity. An increasing number of studies suggest that the ACER family has diverse roles in regulating sphingolipid metabolism and biological processes. Here we discuss the discovery of the ACER family, the biochemical properties, structures, and catalytic mechanisms of its members, and its role in regulating sphingolipid metabolism and biological processes in yeast, insects, plants, and mammals.