Identification of genetic and environmental factors influencing aerial root traits that support biological nitrogen fixation in sorghum.

Plant breeding and genetics play a major role in the adaptation of plants to meet human needs. The current requirement to make agriculture more sustainable can be partly met by a greater reliance on biological nitrogen fixation by symbiotic diazotrophic microorganisms that provide crop plants with ammonium. Select accessions of the cereal crop sorghum (Sorghum bicolor (L.) Moench) form mucilage-producing aerial roots that harbor nitrogen-fixing bacteria. Breeding programs aimed at developing sorghum varieties that support diazotrophs will benefit from a detailed understanding of the genetic and environmental factors contributing to aerial root formation. A genome-wide association study of the sorghum minicore, a collection of 242 landraces, and 30 accessions from the sorghum association panel was conducted in Florida and Wisconsin and under 2 fertilizer treatments to identify loci associated with the number of nodes with aerial roots and aerial root diameter. Sequence variation in genes encoding transcription factors that control phytohormone signaling and root system architecture showed significant associations with these traits. In addition, the location had a significant effect on the phenotypes. Concurrently, we developed F2 populations from crosses between bioenergy sorghums and a landrace that produced extensive aerial roots to evaluate the mode of inheritance of the loci identified by the genome-wide association study. Furthermore, the mucilage collected from aerial roots contained polysaccharides rich in galactose, arabinose, and fucose, whose composition displayed minimal variation among 10 genotypes and 2 fertilizer treatments. These combined results support the development of sorghums with the ability to acquire nitrogen via biological nitrogen fixation.

A Sorghum F-Box Protein Induces an Oxidative Burst in the Defense Against Colletotrichum sublineola.

The hemibiotrophic fungal pathogen Colletotrichum sublineola is the causal agent of anthracnose in sorghum (Sorghum bicolor), resulting in leaf blight, stalk rot, and head blight in susceptible genotypes, with yield losses of up to 50%. The development of anthracnose-resistant cultivars can reduce reliance on fungicides and provide a more sustainable and economical means for disease management. A previous genome-wide association study of the sorghum association panel identified the candidate resistance gene Sobic.005G172300 encoding an F-box protein. To better understand the role of this gene in the defense against C. sublineola, gene expression following infection with C. sublineola was monitored by RNA sequencing in seedlings of sorghum accession SC110, which harbored the resistance allele, and three accessions that harbored a susceptible allele. Only in SC110 did the expression of Sobic.005G172300 increase during the biotrophic phase of infection. Subsequent transcriptome analysis, gene co-expression networks, and gene regulatory networks of inoculated and mock-inoculated seedlings of resistant and susceptible accessions suggest that the increase in expression of Sobic.005G172300 induces an oxidative burst by lowering the concentration of ascorbic acid during the biotrophic phase of infection. Based on gene regulatory network analysis, the protein encoded by Sobic.005G172300 is proposed to target proteins involved in the biosynthesis of ascorbic acid for polyubiquitination through the SCF E3 ubiquitin ligase, causing their degradation via the proteasome.

Genetic diversity, population structure and anthracnose resistance response in a novel sweet sorghum diversity panel.

Sweet sorghum is an attractive feedstock for the production of renewable chemicals and fuels due to the readily available fermentable sugars that can be extracted from the juice, and the additional stream of fermentable sugars that can be obtained from the cell wall polysaccharides in the bagasse. An important selection criterion for new sweet sorghum germplasm is resistance to anthracnose, a disease caused by the fungal pathogen Colletotrichum sublineolum. The identification of novel anthracnose-resistance sources present in sweet sorghum germplasm offers a fast track towards the development of new resistant sweet sorghum germplasm. We established a sweet sorghum diversity panel (SWDP) of 272 accessions from the USDA-ARS National Plant Germplasm (NPGS) collection that includes landraces from 22 countries and advanced breeding material, and that represents ~15% of the NPGS sweet sorghum collection. Genomic characterization of the SWDP identified 171,954 single nucleotide polymorphisms (SNPs) with an average of one SNP per 4,071 kb. Population structure analysis revealed that the SWDP could be stratified into four populations and one admixed group, and that this population structure could be aligned to sorghum's racial classification. Results from a two-year replicated trial of the SWDP for anthracnose resistance response in Texas, Georgia, Florida, and Puerto Rico showed 27 accessions to be resistant across locations, while 145 accessions showed variable resistance response against local pathotypes. A genome-wide association study identified 16 novel genomic regions associated with anthracnose resistance. Four resistance loci on chromosomes 3, 6, 8 and 9 were identified against pathotypes from Puerto Rico, and two resistance loci on chromosomes 3 and 8 against pathotypes from Texas. In Georgia and Florida, three resistance loci were detected on chromosomes 4, 5, 6 and four on chromosomes 4, 5 (two loci) and 7, respectively. One resistance locus on chromosome 2 was effective against pathotypes from Texas and Puerto Rico and a genomic region of 41.6 kb at the tip of chromosome 8 was associated with resistance response observed in Georgia, Texas, and Puerto Rico. This publicly available SWDP and the extensive evaluation of anthracnose resistance represent a valuable genomic resource for the improvement of sorghum.

Structural Similarities and Overlapping Activities among Dihydroflavonol 4-Reductase, Flavanone 4-Reductase, and Anthocyanidin Reductase Offer Metabolic Flexibility in the Flavonoid Pathway.

Flavonoids are potent antioxidants that play a role in defense against pathogens, UV-radiation, and the detoxification of reactive oxygen species. Dihydroflavonol 4-reductase (DFR) and flavanone 4-reductase (FNR) reduce dihydroflavonols and flavanones, respectively, using NAD(P)H to produce flavan-(3)-4-(di)ols in flavonoid biosynthesis. Anthocyanidin reductase (ANR) reduces anthocyanidins to flavan-3-ols. In addition to their sequences, the 3D structures of recombinant DFR, FNR and ANR from sorghum and switchgrass showed a high level of similarity. The catalytic mechanism, substrate-specificity and key residues of three reductases were deduced from crystal structures, site-directed mutagenesis, molecular docking, kinetics, and thermodynamic ana-lyses. Although DFR displayed its highest activity against dihydroflavonols, it also showed activity against flavanones and anthocyanidins. It was inhibited by the flavonol quercetin and high concentrations of dihydroflavonols/flavonones. SbFNR1 and SbFNR2 did not show any activity against dihydroflavonols. However, SbFNR1 displayed activity against flavanones and ANR activity against two anthocyanidins, cyanidin and pelargonidin. Therefore, SbFNR1 and SbFNR2 could be specific ANR isozymes without delphinidin activity. Sorghum has high concentrations of 3-deoxyanthocyanidins in vivo, supporting the observed high activity of SbDFR against flavonols. Mining of expression data indicated substantial induction of these three reductase genes in both switchgrass and sorghum in response to biotic stress. Key signature sequences for proper DFR/ANR classification are proposed and could form the basis for future metabolic engineering of flavonoid metabolism.

Seasonal Variation in Cell Wall Composition and Carbohydrate Metabolism in the Seagrass Posidonia oceanica Growing at Different Depths.

Posidonia oceanica is a common seagrass in the Mediterranean Sea that is able to sequester large amounts of carbon. The carbon assimilated during photosynthesis can be partitioned into non-structural sugars and cell-wall polymers. In this study, we investigated the distribution of carbon in starch, soluble carbohydrates and cell-wall polymers in leaves and rhizomes of P. oceanica. Analyses were performed during summer and winter in meadows located south of the Frioul archipelago near Marseille, France. The leaves and rhizomes were isolated from plants collected in shallow (2 m) and deep water (26 m). Our results showed that P. oceanica stores more carbon as starch, sucrose and cellulose in summer and that this is more pronounced in rhizomes from deep-water plants. In winter, the reduction in photoassimilates was correlated with a lower cellulose content, compensated with a greater lignin content, except in rhizomes from deep-water plants. The syringyl-to-guaiacyl (S/G) ratio in the lignin was higher in leaves than in rhizomes and decreased in rhizomes in winter, indicating a change in the distribution or structure of the lignin. These combined data show that deep-water plants store more carbon during summer, while in winter the shallow- and deep-water plants displayed a different cell wall composition reflecting their environment.

Metabolic link between auxin production and specialized metabolites in Sorghum bicolor.

Aldoximes are amino acid derivatives that serve as intermediates for numerous specialized metabolites including cyanogenic glycosides, glucosinolates, and auxins. Aldoxime formation is mainly catalyzed by cytochrome P450 monooxygenases of the 79 family (CYP79s) that can have broad or narrow substrate specificity. Except for SbCYP79A1, aldoxime biosynthetic enzymes in the cereal sorghum (Sorghum bicolor) have not been characterized. This study identified nine CYP79-encoding genes in the genome of sorghum. A phylogenetic analysis of CYP79 showed that SbCYP79A61 formed a subclade with maize ZmCYP79A61, previously characterized to be involved in aldoxime biosynthesis. Functional characterization of this sorghum enzyme using transient expression in Nicotiana benthamiana and stable overexpression in Arabidopsis thaliana revealed that SbCYP79A61 catalyzes the production of phenylacetaldoxime (PAOx) from phenylalanine but, unlike the maize enzyme, displays no detectable activity against tryptophan. Additionally, targeted metabolite analysis after stable isotope feeding assays revealed that PAOx can serve as a precursor of phenylacetic acid (PAA) in sorghum and identified benzyl cyanide as an intermediate of PAOx-derived PAA biosynthesis in both sorghum and maize. Taken together, our results demonstrate that SbCYP79A61 produces PAOx in sorghum and may serve in the biosynthesis of other nitrogen-containing phenylalanine-derived metabolites involved in mediating biotic and abiotic stresses.

A sorghum ascorbate peroxidase with four binding sites has activity against ascorbate and phenylpropanoids.

In planta, H2O2 is produced as a by-product of enzymatic reactions and during defense responses. Ascorbate peroxidase (APX) is a key enzyme involved in scavenging cytotoxic H2O2. Here, we report the crystal structure of cytosolic APX from sorghum (Sorghum bicolor) (Sobic.001G410200). While the overall structure of SbAPX was similar to that of other APXs, SbAPX uniquely displayed four bound ascorbates rather than one. In addition to the ɣ-heme pocket identified in other APXs, ascorbates were bound at the δ-meso and two solvent-exposed pockets. Consistent with the presence of multiple binding sites, our results indicated that the H2O2-dependent oxidation of ascorbate displayed positive cooperativity. Bound ascorbate at two surface sites established an intricate proton network with ascorbate at the ɣ-heme edge and δ-meso sites. Based on crystal structures, steady-state kinetics, and site-directed mutagenesis results, both ascorbate molecules at the ɣ-heme edge and the one at the surface are expected to participate in the oxidation reaction. We provide evidence that the H2O2-dependent oxidation of ascorbate by APX produces a C2-hydrated bicyclic hemiketal form of dehydroascorbic acid at the ɣ-heme edge, indicating two successive electron transfers from a single-bound ascorbate. In addition, the δ-meso site was shared with several organic compounds, including p-coumaric acid and other phenylpropanoids, for the potential radicalization reaction. Site-directed mutagenesis of the critical residue at the ɣ-heme edge (R172A) only partially reduced polymerization activity. Thus, APX removes stress-generated H2O2 with ascorbates, and also uses this same H2O2 to potentially fortify cell walls via oxidative polymerization of phenylpropanoids in response to stress.

Dysregulation of Cell Envelope Homeostasis in Staphylococcus aureus Exposed to Solvated Lignin.

Lignin is an aromatic plant cell wall polymer that facilitates water transport through the vasculature of plants and is generated in large quantities as an inexpensive by-product of pulp and paper manufacturing and biorefineries. Although lignin's ability to reduce bacterial growth has been reported previously, its hydrophobicity complicates the ability to examine its biological effects on living cells in aqueous growth media. We recently described the ability to solvate lignin in Good's buffers with neutral pH, a breakthrough that allowed examination of lignin's antimicrobial effects against the human pathogen Staphylococcus aureus. These analyses showed that lignin damages the S. aureus cell membrane, causes increased cell clustering, and inhibits growth synergistically with tunicamycin, a teichoic acid synthesis inhibitor. In the present study, we examined the physiological and transcriptomic responses of S. aureus to lignin. Intriguingly, lignin restored the susceptibility of genetically resistant S. aureus isolates to penicillin and oxacillin, decreased intracellular pH, impaired normal cell division, and rendered cells more resistant to detergent-induced lysis. Additionally, transcriptome sequencing (RNA-Seq) differential expression (DE) analysis of lignin-treated cultures revealed significant gene expression changes (P < 0.05 with 5% false discovery rate [FDR]) related to the cell envelope, cell wall physiology, fatty acid metabolism, and stress resistance. Moreover, a pattern of concurrent up- and downregulation of genes within biochemical pathways involved in transmembrane transport and cell wall physiology was observed, which likely reflects an attempt to tolerate or compensate for lignin-induced damage. Together, these results represent the first comprehensive analysis of lignin's antibacterial activity against S. aureus. IMPORTANCE S. aureus is a leading cause of skin and soft tissue infections. The ability of S. aureus to acquire genetic resistance to antibiotics further compounds its ability to cause life-threatening infections. While the historical response to antibiotic resistance has been to develop new antibiotics, bacterial pathogens are notorious for rapidly acquiring genetic resistance mechanisms. As such, the development of adjuvants represents a viable way of extending the life span of current antibiotics to which pathogens may already be resistant. Here, we describe the phenotypic and transcriptomic response of S. aureus to treatment with lignin. Our results demonstrate that lignin extracted from sugarcane and sorghum bagasse restores S. aureus susceptibility to ß-lactams, providing a premise for repurposing these antibiotics in treatment of resistant S. aureus strains, possibly in the form of topical lignin/ß-lactam formulations.

The maize low-lignin brown midrib3 mutant shows pleiotropic effects on photosynthetic and cell wall metabolisms in response to chilling.

Maize (Zea mays L.) is one of the major cereal crops in the world and is highly sensitive to low temperature. Here, changes in photosynthetic and cell wall metabolisms were investigated during a long chilling exposure in inbred line F2 and a low-lignin near-isogenic brown midrib3 mutant (F2bm3), which has a mutation in the caffeic acid O-methyltransferase (COMT) gene. Results revealed that the plant biomass was reduced, and this was more pronounced in F2bm3. Photosynthesis was altered in both lines with distinct changes in photosynthetic pigment content between F2bm3 and F2, indicating an alternative photoprotection mechanism between lines under chilling. Starch remobilization was observed in F2bm3 while concentrations of sucrose, fructose and starch increased in F2, suggesting a reduced sugar partitioning in F2. The cell wall was altered upon chilling, resulting in changes in the composition of glucuronorabinoxylan and a reduced cellulose level in F2. Chilling shifted lignin subunit composition in F2bm3 mutant to a higher proportion of p-hydroxyphenyl (H) units, whereas it resulted in lignin with a higher proportion of syringyl (S) residues in F2. On average, the total cell wall ferulic acid (FA) content increased in both genotypes, with an increase in ether-linked FA in F2bm3, suggesting a greater degree of cross-linking to lignin. The reinforcement of the cell wall with lignin enriched in H-units and a higher concentration in cell-wall-bound FA observed in F2bm3 as a response to chilling, could be a strategy to protect the photosystems.

Functional and structural insight into the flexibility of cytochrome P450 reductases from Sorghum bicolor and its implications for lignin composition.

Plant NADPH-dependent cytochrome P450 reductase (CPR) is a multidomain enzyme that donates electrons for hydroxylation reactions catalyzed by class II cytochrome P450 monooxygenases involved in the synthesis of many primary and secondary metabolites. These P450 enzymes include trans-cinnamate-4-hydroxylase, p-coumarate-3'-hydroxylase, and ferulate-5-hydroxylase involved in monolignol biosynthesis. Because of its role in monolignol biosynthesis, alterations in CPR activity could change the composition and overall output of lignin. Therefore, to understand the structure and function of three CPR subunits from sorghum, recombinant subunits SbCPR2a, SbCPR2b, and SbCPR2c were subjected to X-ray crystallography and kinetic assays. Steady-state kinetic analyses demonstrated that all three CPR subunits supported the oxidation reactions catalyzed by SbC4H1 (CYP73A33) and SbC3'H (CYP98A1). Furthermore, comparing the SbCPR2b structure with the well-investigated CPRs from mammals enabled us to identify critical residues of functional importance and suggested that the plant flavin mononucleotide-binding domain might be more flexible than mammalian homologs. In addition, the elucidated structure of SbCPR2b included the first observation of NADP+ in a native CPR. Overall, we conclude that the connecting domain of SbCPR2, especially its hinge region, could serve as a target to alter biomass composition in bioenergy and forage sorghums through protein engineering.

Delivery of DNA into Human Cells by Functionalized Lignin Nanoparticles.

Lignin is an aromatic plant cell wall polymer that is generated in large quantities as a low-value by-product by the pulp and paper industry and by biorefineries that produce renewable fuels and chemicals from plant biomass. Lignin structure varies among plant species and as a function of the method used for its extraction from plant biomass. We first explored the impact of this variation on the physico-chemical properties of lignin nanoparticles (LNPs) produced via a solvent exchange procedure and then examined whether LNPs produced from industrial sources of lignin could be used as delivery vehicles for DNA. Spherical LNPs were formed from birch and wheat BioLignin™ and from poplar thioglycolic acid lignin after dissolving the lignin in tetrahydrofuran (THF) and dialyzing it against water. Dynamic light scattering indicated that the diameter of these LNPs was dependent on the initial concentration of the lignin, while electrophoretic light scattering indicated that the LNPs had a negative zeta potential, which became less negative as the diameter increased. The dynamics of LNP formation as a function of the initial lignin concentration varied as a function of the source of the lignin, as did the absolute value of the zeta potential. After coating the LNPs with cationic poly-l-lysine, an electrophoretic mobility shift assay indicated that DNA could adsorb to LNPs. Upon transfection of human A549 lung carcinoma basal epithelial cells with functionalized LNPs carrying plasmid DNA encoding the enhanced green fluorescent protein (eGFP), green foci were observed under the microscope, and the presence of eGFP in the transfected cells was confirmed by ELISA. The low cytotoxicity of these LNPs and the ability to tailor diameter and zeta potential make these LNPs of interest for future gene therapy applications.

The Sorghum (Sorghum bicolor) Brown Midrib 30 Gene Encodes a Chalcone Isomerase Required for Cell Wall Lignification.

In sorghum (Sorghum bicolor) and other C4 grasses, brown midrib (bmr) mutants have long been associated with plants impaired in their ability to synthesize lignin. The brown midrib 30 (Bmr30) gene, identified using a bulk segregant analysis and next-generation sequencing, was determined to encode a chalcone isomerase (CHI). Two independent mutations within this gene confirmed that loss of its function was responsible for the brown leaf midrib phenotype and reduced lignin concentration. Loss of the Bmr30 gene function, as shown by histochemical staining of leaf midrib and stalk sections, resulted in altered cell wall composition. In the bmr30 mutants, CHI activity was drastically reduced, and the accumulation of total flavonoids and total anthocyanins was impaired, which is consistent with its function in flavonoid biosynthesis. The level of the flavone lignin monomer tricin was reduced 20-fold in the stem relative to wild type, and to undetectable levels in the leaf tissue of the mutants. The bmr30 mutant, therefore, harbors a mutation in a phenylpropanoid biosynthetic gene that is key to the interconnection between flavonoids and monolignols, both of which are utilized for lignin synthesis in the grasses.

The inheritance of anthracnose (Colletotrichum sublineola) resistance in sorghum differential lines QL3 and IS18760.

Anthracnose caused by the fungal pathogen C. sublineola is an economically important constraint on worldwide sorghum production. The most effective strategy to safeguard yield is through the introgression of resistance alleles. This requires elucidation of the genetic basis of the different resistance sources that have been identified. In this study, 223 recombinant inbred lines (RILs) derived from crossing anthracnose-differentials QL3 (96 RILs) and IS18760 (127 RILs) with the common susceptible parent PI609251 were evaluated at four field locations in the United States (Florida, Georgia, Texas, and Puerto Rico) for their anthracnose resistance response. Both RIL populations were highly susceptible to anthracnose in Florida and Georgia, while in Puerto Rico and Texas they were segregating for anthracnose resistance response. A genome scan using a composite linkage map of 982 single nucleotide polymorphisms (SNPs) detected two genomic regions of 4.31 and 0.85 Mb on chromosomes 4 and 8, respectively, that explained 10-27% of the phenotypic variation in Texas and Puerto Rico. In parallel, a subset of 43 RILs that contained 67% of the recombination events were evaluated against anthracnose pathotypes from Arkansas (2), Puerto Rico (2) and Texas (4) in the greenhouse. A genome scan showed that the 7.57 Mb region at the distal end of the short arm of chromosome 5 is associated with the resistance response against the pathotype AMP-048 from Arkansas. Comparative analysis identified the genomic region on chromosome 4 overlaps with an anthracnose resistance locus identified in another anthracnose-differential line, SC414-12E, indicating this genomic region is of interest for introgression in susceptible sorghum germplasm. Candidate gene analysis for the resistance locus on chromosome 5 identified an R-gene cluster that has high similarity to another R-gene cluster associated with anthracnose resistance on chromosome 9.

Elucidating Anthracnose Resistance Mechanisms in Sorghum-A Review.

Sorghum (Sorghum bicolor) is the fifth most cultivated cereal crop in the world, traditionally providing food, feed, and fodder, but more recently also fermentable sugars for the production of renewable fuels and chemicals. The hemibiotrophic fungal pathogen Colletotrichum sublineola, the causal agent of anthracnose disease in sorghum, is prevalent in the warm and humid climates where much of the sorghum is cultivated and poses a serious threat to sorghum production. The use of anthracnose-resistant sorghum germplasm is the most environmentally and economically sustainable way to protect sorghum against this pathogen. Even though multiple anthracnose resistance loci have been mapped in diverse sorghum germplasm in recent years, the diversity in C. sublineola pathotypes at the local and regional levels means that these resistance genes are not equally effective in different areas of cultivation. This review summarizes the genetic and cytological data underlying sorghum's defense response and describes recent developments that will enable a better understanding of the interactions between sorghum and C. sublineola at the molecular level. This includes releases of the sorghum genome and the draft genome of C. sublineola, the use of next-generation sequencing technologies to identify gene expression networks activated in response to infection, and improvements in methodologies to validate resistance genes, notably virus-induced and transgenic gene silencing approaches.

Structure and Function of the Cytochrome P450 Monooxygenase Cinnamate 4-hydroxylase from Sorghum bicolor.

Cinnamate 4-hydroxylase (C4H; CYP73A) is a cytochrome P450 monooxygenase associated externally with the endoplasmic reticulum of plant cells. The enzyme uses NADPH-cytochrome P450 reductase as a donor of electrons and hydroxylates cinnamic acid to form 4-coumaric acid in phenylpropanoid metabolism. In order to better understand the structure and function of this unique class of plant P450 enzymes, we have characterized the enzyme C4H1 from lignifying tissues of sorghum (Sorghum bicolor), encoded by Sobic.002G126600 Here we report the 1.7 Å resolution crystal structure of CYP73A33. The obtained structural information, along with the results of the steady-state kinetic analysis and the absorption spectroscopy titration, displays a high degree of similarity of the structural and functional features of C4H to those of other P450 proteins. Our data also suggest the presence of a putative allosteric substrate-binding site in a hydrophobic pocket on the enzyme surface. In addition, comparing the newly resolved structure with those of well-investigated cytochromes P450 from mammals and bacteria enabled us to identify those residues of critical functional importance and revealed a unique sequence signature that is potentially responsible for substrate specificity and catalytic selectivity of C4H.

Genomic Dissection of Anthracnose (Colletotrichum sublineolum) Resistance Response in Sorghum Differential Line SC112-14.

Sorghum production is expanding to warmer and more humid regions where its production is being limited by multiple fungal pathogens. Anthracnose, caused by Colletotrichum sublineolum, is one of the major diseases in these regions, where it can cause yield losses of both grain and biomass. In this study, 114 recombinant inbred lines (RILs) derived from resistant sorghum line SC112-14 were evaluated at four distinct geographic locations in the United States for response to anthracnose. A genome scan using a high-density linkage map of 3,838 single nucleotide polymorphisms (SNPs) detected two loci at 5.25 and 1.18 Mb on chromosomes 5 and 6, respectively, that explain up to 59% and 44% of the observed phenotypic variation. A bin-mapping approach using a subset of 31 highly informative RILs was employed to determine the disease response to inoculation with ten anthracnose pathotypes in the greenhouse. A genome scan showed that the 5.25 Mb region on chromosome 5 is associated with a resistance response to nine pathotypes. Five SNP markers were developed and used to fine map the locus on chromosome 5 by evaluating 1,500 segregating F2:3 progenies. Based on the genotypic and phenotypic analyses of 11 recombinants, the locus was narrowed down to a 470-kb genomic region. Following a genome-wide association study based on 574 accessions previously phenotyped and genotyped, the resistance locus was delimited to a 34-kb genomic interval with five candidate genes. All five candidate genes encode proteins associated with plant immune systems, suggesting they may act in synergy in the resistance response.

Cellulose-based injectable hydrogel composite for pH-responsive and controllable drug delivery.

Cellulose-based biocompatible, tunable and injectable hydrogels embedded with pH-responsive diblock copolymer micelles were constructed to achieve localized drug delivery with prolonged, stimuli-driven and slow-release function. First, we prepared two types of modified carboxymethyl cellulose (CMC) including hydrazide-modified carboxymethyl cellulose (CMC-NH2) and oxidized carboxymethyl cellulose (CMC-CHO) with varying degrees of oxidation. Then, pH-responsive poly (ethylene oxide)-block-poly (2-(diisopropylamino) ethyl methacrylate) (PEO-b-PDPA) copolymers as micelle cores to carry hydrophobic substances were also synthesized through atom transfer radical polymerization (ATRP). An injectable hydrogel composite system was finally obtained by mixing CMC-NH2 and CMC-CHO polymer suspensions containing PEO-b-PDPA copolymer micelles through a Schiff base reaction. This newly-synthesized, tunable, cellulose-based double barrier system exhibits a pH-triggered, prolonged, and slow-release profile based on the release test using both Nile Red dye and doxorubicin. The hydrogel system also exhibited comparable storage moduli and tunable degradation properties.

A new reference genome for Sorghum bicolor reveals high levels of sequence similarity between sweet and grain genotypes: implications for the genetics of sugar metabolism.

BACKGROUND: The process of crop domestication often consists of two stages: initial domestication, where the wild species is first cultivated by humans, followed by diversification, when the domesticated species are subsequently adapted to more environments and specialized uses. Selective pressure to increase sugar accumulation in certain varieties of the cereal crop Sorghum bicolor is an excellent example of the latter; this has resulted in pronounced phenotypic divergence between sweet and grain-type sorghums, but the genetic mechanisms underlying these differences remain poorly understood. RESULTS: Here we present a new reference genome based on an archetypal sweet sorghum line and compare it to the current grain sorghum reference, revealing a high rate of nonsynonymous and potential loss of function mutations, but few changes in gene content or overall genome structure. We also use comparative transcriptomics to highlight changes in gene expression correlated with high stalk sugar content and show that changes in the activity and possibly localization of transporters, along with the timing of sugar metabolism play a critical role in the sweet phenotype. CONCLUSIONS: The high level of genomic similarity between sweet and grain sorghum reflects their historical relatedness, rather than their current phenotypic differences, but we find key changes in signaling molecules and transcriptional regulators that represent new candidates for understanding and improving sugar metabolism in this important crop.

Lignin-based polymers and nanomaterials.

Approximately 50 million tons of lignin are currently produced annually as a by-product of the pulp- and paper industry, and this amount is likely to double in the future with the anticipated production of renewable fuels and chemicals from lignocellulosic biomass, as a sustainable alternative to petroleum. The latter process can be expedited by valorizing lignin, which entails making products from lignin that generate additional revenues for biorefineries so that the production of biofuels becomes more competitive with gasoline. Industrially produced lignin is considered a low-value material that is used as a boiler fuel to generate heat and electricity, and as an ingredient of adhesives, cement, and drilling fluids for underwater oil wells. The aromatic nature of lignin, its ability to participate in radical-mediated cross-linking reactions, the many functional groups available for derivatization or chemical reactions, and its amenability to existing procedures for making thermoplastics, make it attractive as an additive to polymers to enhance UV-tolerance and/or other physico-chemical properties. Lignin can also be used as the basis for various nanomaterials, either per se or in combination with other polymers. This review summarizes recent developments in the synthesis of lignin-containing polymers and nanomaterials, whereby inherent variation in lignin subunit composition and structure, as a function of plant species and lignin extraction method, offer unique opportunities for fine-tuning material properties (e.g. tensile strength, hardness, elasticity) to match specific applications.

Biochemical and Structural Analysis of Substrate Specificity of a Phenylalanine Ammonia-Lyase.

Phenylalanine ammonia-lyase (PAL) is the first enzyme of the general phenylpropanoid pathway catalyzing the nonoxidative elimination of ammonia from l-phenylalanine to give trans-cinnamate. In monocots, PAL also displays tyrosine ammonia lyase (TAL) activity, leading to the formation of p-coumaric acid. The catalytic mechanism and substrate specificity of a major PAL from sorghum (Sorghum bicolor; SbPAL1), a strategic plant for bioenergy production, were deduced from crystal structures, molecular docking, site-directed mutagenesis, and kinetic and thermodynamic analyses. This first crystal structure of a monocotyledonous PAL displayed a unique conformation in its flexible inner loop of the 4-methylidene-imidazole-5-one (MIO) domain compared with that of dicotyledonous plants. The side chain of histidine-123 in the MIO domain dictated the distance between the catalytic MIO prosthetic group created from 189Ala-Ser-Gly191 residues and the bound l-phenylalanine and l-tyrosine, conferring the deamination reaction through either the Friedel-Crafts or E2 reaction mechanism. Several recombinant mutant SbPAL1 enzymes were generated via structure-guided mutagenesis, one of which, H123F-SbPAL1, has 6.2 times greater PAL activity without significant TAL activity. Additional PAL isozymes of sorghum were characterized and categorized into three groups. Taken together, this approach identified critical residues and explained substrate preferences among PAL isozymes in sorghum and other monocots, which can serve as the basis for the engineering of plants with enhanced biomass conversion properties, disease resistance, or nutritional quality.