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Sorghum (Sorghum bicolor) is drought‐tolerant and has diverse germplasm for food, feed, forage and bioenergy. However, stalk diseases reduce quality and yield of biomass and grain, especially under drought. Previously, brown midrib (bmr) mutations in monolignol biosynthesis were shown to reduce lignin content and alter composition but were not more susceptible to stalk diseases than wild‐type lines. Recently characterized bmr mutations were shown to affect flavonoid biosynthesis (chalcone isomerase; bmr30‐1) or 1‐carbon metabolism (folylpolyglutamate synthase; bmr19‐1107, bmr19‐1168 and bmr19‐1937). Two other mutations, bmr29‐1 and bmr31‐1, have not yet been characterized. The six mutations were incorporated into elite genetic backgrounds (RTx430, BTx623 and BWheatland) to develop near‐isogenic lines containing each mutation. Using peduncle inoculations with Fusarium thapsinum and F. proliferatum (Fusarium stalk rot) and Macrophomina phaseolina (charcoal rot) under well‐watered conditions, most bmr lines were at least as resistant as the corresponding wild type, except for RTx430 bmr19‐1937 that had significantly longer mean lesion lengths when inoculated with M. phaseolina. Based on significantly reduced lesion lengths following inoculations with F. proliferatum and F. thapsinum, respectively, bmr29‐1 and bmr31‐1 lines were screened using basal stalk inoculations under well‐watered and water‐deficit conditions. The bmr lines were at least as resistant as the corresponding wild‐type lines. Wild‐type BTx623 was highly susceptible to M. phaseolina under water deficit, but near‐isogenic bmr29‐1 and bmr31‐1 lines had significantly shorter mean lesion lengths. Incorporation of these mutations can increase resistance to stalk pathogens in cultivar and hybrid development for feed, bioenergy and production of biomass‐based green chemicals. [ABSTRACT FROM AUTHOR]

Summary: Lignin is a key target for modifying lignocellulosic biomass for efficient biofuel production. Brown midrib 12 (bmr12) encodes the sorghum caffeic acid O‐methyltransferase (COMT) and is one of the key enzymes in monolignol biosynthesis. Loss of function mutations in COMT reduces syringyl (S) lignin subunits and improves biofuel conversion rate. Although lignin plays an important role in maintaining cell wall integrity of xylem vessels, physiological and molecular consequences due to loss of COMT on root growth and adaptation to water deficit remain unexplored.We addressed this gap by evaluating the root morphology, anatomy and transcriptome of bmr12 mutant. The mutant had reduced lateral root density (LRD) and altered root anatomy and response to water limitation. The wild‐type exhibits similar phenotypes under water stress, suggesting that bmr12 may be in a water deficit responsive state even in well‐watered conditions.bmr12 had increased transcript abundance of genes involved in (a)biotic stress response, gibberellic acid (GA) biosynthesis and signaling. We show that bmr12 is more sensitive to exogenous GA application and present evidence for the role of GA in regulating reduced LRD in bmr12.These findings elucidate the phenotypic and molecular consequences of COMT deficiency under optimal and water stress environments in grasses. [ABSTRACT FROM AUTHOR]

To grow and thrive plants must be able to adapt to both adverse environmental conditions and attack by a variety of pests. Elucidating the sophisticated mechanisms plants have developed to achieve this has been the focus of many studies. What is less well understood is how plants respond when faced with multiple stressors simultaneously. In this study, we assess the response of Zea mays (maize) to the combinatorial stress of flooding and infestation with the insect pest Spodoptera frugiperda (fall armyworm). This combined stress leads to elevated production of the defence hormone salicylic acid, which does not occur in the individual stresses, and the resultant salicylic acid‐dependent increase in S. frugiperda resistance. Remodelling of phenylpropanoid pathways also occurs in response to this combinatorial stress leading to increased production of the anti‐insect C‐glycosyl flavones (maysins) and the herbivore‐induced volatile phenolics, benzyl acetate, and phenethyl acetate. Furthermore, changes in cellular redox status also occur, as indicated by reductions in peroxidase and polyphenol oxidase activity. These data suggest that metabolite changes important for flooding tolerance and anti‐insect defence may act both additively and synergistically to provide extra protection to the plant. Flooding stress in Zea mays leads to a salicylic acid‐dependent increase in resistance to the insect pest Spodoptera frugiperda. This combinatorial stress results in phenylpropanoid remodelling, changes in maysin and herbivore‐induced volatile production, and altered cellular redox status. [ABSTRACT FROM AUTHOR]

Few transcription factors have been identified in C4 grasses that either positively or negatively regulate monolignol biosynthesis., Previously, the overexpression of SbMyb60 in sorghum ( Sorghum bicolor) has been shown to induce monolignol biosynthesis, which leads to elevated lignin deposition and altered cell wall composition. To determine how SbMyb60 overexpression impacts other metabolic pathways, RNA-Seq and metabolite profiling were performed on stalks and leaves., 35S::SbMyb60 was associated with the transcriptional activation of genes involved in aromatic amino acid, S-adenosyl methionine ( SAM) and folate biosynthetic pathways. The high coexpression values between SbMyb60 and genes assigned to these pathways indicate that SbMyb60 may directly induce their expression. In addition, 35S::SbMyb60 altered the expression of genes involved in nitrogen (N) assimilation and carbon (C) metabolism, which may redirect C and N towards monolignol biosynthesis. Genes linked to UDP-sugar biosynthesis and cellulose synthesis were also induced, which is consistent with the observed increase in cellulose deposition in the internodes of 35S::SbMyb60 plants. However, SbMyb60 showed low coexpression values with these genes and is not likely to be a direct regulator of cell wall polysaccharide biosynthesis., These findings indicate that SbMyb60 can activate pathways beyond monolignol biosynthesis, including those that synthesize the substrates and cofactors required for lignin biosynthesis. [ABSTRACT FROM AUTHOR]

Switchgrass ( Panicum virgatum), a perennial, polyploid, C4 warm-season grass is among the foremost herbaceous species being advanced as a source of biomass for biofuel end uses. At the end of every growing season, the aerial tissues senesce, and the below-ground rhizomes become dormant. Future growth is dependent on the successful over-wintering of the rhizomes. Although the importance of rhizome health to overall year-upon-year plant productivity has been long recognized, there is limited information on seasonal changes occurring during dormancy at both the transcriptome and metabolite levels. Here, global changes in transcriptomes and metabolites were investigated over two growing seasons in rhizomes harvested from field-grown plants. The objectives were: (a) synthesize information on cellular processes that lead to dormancy; and (b) provide models that could account for major metabolic pathways present in dormant switchgrass rhizomes. Overall, metabolism during dormancy appeared to involve discrete but interrelated events. One was a response to abscisic acid that resulted in dehydration, increases in osmolytes and upregulation of autophagic processes, likely through the target of rapamycin complex and sucrose non-fermentative-related kinase-based signaling cascades. Another was a recalibration of energy transduction through apparent reductions in mitochondrial oxidative phosphorylation, increases in substrate level generation of ATP and reducing equivalents, and recycling of N and possibly CO2 through refixation. Lastly, transcript abundances indicated that cold-related signaling was also occurring. Altogether, these data provide a detailed overview of rhizome metabolism, especially during dormancy, which can be exploited in the future to improve winter survival in switchgrass. [ABSTRACT FROM AUTHOR]

The phenylpropanoid biosynthetic pathway that generates lignin subunits represents a significant target for altering the abundance and composition of lignin. The global regulators of phenylpropanoid metabolism may include MYB transcription factors, whose expression levels have been correlated with changes in secondary cell wall composition and the levels of several other aromatic compounds, including anthocyanins and flavonoids. While transcription factors correlated with downregulation of the phenylpropanoid biosynthesis pathway have been identified in several grass species, few transcription factors linked to activation of this pathway have been identified in C4 grasses, some of which are being developed as dedicated bioenergy feedstocks. In this study we investigated the role of SbMyb60 in lignin biosynthesis in sorghum ( Sorghum bicolor), which is a drought-tolerant, high-yielding biomass crop. Ectopic expression of this transcription factor in sorghum was associated with higher expression levels of genes involved in monolignol biosynthesis, and led to higher abundances of syringyl lignin, significant compositional changes to the lignin polymer and increased lignin concentration in biomass. Moreover, transgenic plants constitutively overexpressing SbMyb60 also displayed ectopic lignification in leaf midribs and elevated concentrations of soluble phenolic compounds in biomass. Results indicate that overexpression of SbMyb60 is associated with activation of monolignol biosynthesis in sorghum. SbMyb60 represents a target for modification of plant cell wall composition, with the potential to improve biomass for renewable uses. [ABSTRACT FROM AUTHOR]

The presence of lignin reduces the quality of lignocellulosic biomass for forage materials and feedstock for biofuels. In C4 grasses, the brown midrib phenotype has been linked to mutations to genes in the monolignol biosynthesis pathway. For example, the Bmr6 gene in sorghum ( Sorghum bicolor) has been previously shown to encode cinnamyl alcohol dehydrogenase (CAD), which catalyzes the final step of the monolignol biosynthesis pathway. Mutations in this gene have been shown to reduce the abundance of lignin, enhance digestibility, and improve saccharification efficiencies and ethanol yields. Nine sorghum lines harboring five different bmr6 alleles were identified in an EMS-mutagenized TILLING population. DNA sequencing of Bmr6 revealed that the majority of the mutations impacted evolutionarily conserved amino acids while three-dimensional structural modeling predicted that all of these alleles interfered with the enzyme's ability to bind with its NADPH cofactor. All of the new alleles reduced in vitro CAD activity levels and enhanced glucose yields following saccharification. Further, many of these lines were associated with higher reductions in acid detergent lignin compared to lines harboring the previously characterized bmr6-ref allele. These bmr6 lines represent new breeding tools for manipulating biomass composition to enhance forage and feedstock quality. [ABSTRACT FROM AUTHOR]

Successful modification of plant cell-wall composition without compromising plant integrity is dependent on being able to modify the expression of specific genes, but this can be very challenging when the target genes are members of multigene families. 4-coumarate:CoA ligase (4CL) catalyzes the formation of 4-coumaroyl CoA, a precursor of both flavonoids and monolignols, and is an attractive target for transgenic down-regulation aimed at improving agro-industrial properties. Inconsistent phenotypes of transgenic plants have been attributed to variable levels of down-regulation of multiple 4CL genes. Phylogenetic analysis of the sorghum genome revealed 24 4CL(-like) proteins, five of which cluster with bona fide 4CLs from other species. Using a map-based cloning approach and analysis of two independent mutant alleles, the sorghum brown midrib2 ( bmr2) locus was shown to encode 4CL. In vitro enzyme assays indicated that its preferred substrate is 4-coumarate. Missense mutations in the two bmr2 alleles result in loss of 4CL activity, probably as a result of improper folding as indicated by molecular modeling. Bmr2 is the most highly expressed 4CL in sorghum stems, leaves and roots, both at the seedling stage and in pre-flowering plants, but the products of several paralogs also display 4CL activity and compensate for some of the lost activity. The contribution of the paralogs varies between developmental stages and tissues. Gene expression assays indicated that Bmr2 is under auto-regulatory control, as reduced 4CL activity results in over-expression of the defective gene. Several 4CL paralogs are also up-regulated in response to the mutation. [ABSTRACT FROM AUTHOR]

Summary Lignin, a complex heterogenous polymer present in virtually all plant cell walls, plays a critical role in protecting plants from various stresses. However, little is known about how lignin modifications in sorghum will impact plant defense against sugarcane aphids (SCA), a key pest of sorghum. We utilized the sorghum brown midrib (bmr) mutants, which are impaired in monolignol synthesis, to understand sorghum defense mechanisms against SCA. We found that loss of Bmr12 function and overexpression (OE) of Bmr12 provided enhanced resistance and susceptibility to SCA, respectively, as compared with wild‐type (WT; RTx430) plants. Monitoring of the aphid feeding behavior indicated that SCA spent more time in reaching the first sieve element phase on bmr12 plants compared with RTx430 and Bmr12‐OE plants. A combination of transcriptomic and metabolomic analyses revealed that bmr12 plants displayed altered auxin metabolism upon SCA infestation and specifically, elevated levels of auxin conjugate indole‐3‐acetic acid–aspartic acid (IAA–Asp) were observed in bmr12 plants compared with RTx430 and Bmr12‐OE plants. Furthermore, exogenous application of IAA–Asp restored resistance in Bmr12‐OE plants, and artificial diet aphid feeding trial bioassays revealed that IAA–Asp is associated with enhanced resistance to SCA. Our findings highlight the molecular underpinnings that contribute to sorghum bmr12‐mediated resistance to SCA. [ABSTRACT FROM AUTHOR]