The IAA17.1/HSFA5a module enhances salt tolerance in Populus tomentosa by regulating flavonol biosynthesis and ROS levels in lateral roots.

Auxin signaling provides a promising approach to controlling root system architecture and improving stress tolerance in plants. However, how the auxin signaling is transducted in this process remains unclear. The Aux indole-3-acetic acid (IAA) repressor IAA17.1 is stabilized by salinity, and primarily expressed in the lateral root (LR) primordia and tips in poplar. Overexpression of the auxin-resistant form of IAA17.1 (IAA17.1m) led to growth inhibition of LRs, markedly reduced salt tolerance, increased reactive oxygen species (ROS) levels, and decreased flavonol content. We further identified that IAA17.1 can interact with the heat shock protein HSFA5a, which was highly expressed in roots and induced by salt stress. Overexpression of HSFA5a significantly increased flavonol content, reduced ROS accumulation, enhanced LR growth and salt tolerance in transgenic poplar. Moreover, HSFA5a could rescue the defective phenotypes caused by IAA17.1m. Expression analysis showed that genes associated with flavonol biosynthesis were altered in IAA17.1m- and HAFA5a-overexpressing plants. Furthermore, we identified that HSFA5a directly activated the expression of key enzyme genes in the flavonol biosynthesis pathway, while IAA17.1 suppressed HSFA5a-mediated activation of these genes. Collectively, the IAA17.1/HSFA5a module regulates flavonol biosynthesis, controls ROS accumulation, thereby modulating the root system of poplar to adapt to salt stress.

Integrative transcriptomic and metabolomic analyses provide insight into the long-term submergence response mechanisms of young Salix variegata stems.

MAIN CONCLUSION: The mechanisms underlying long-term complete submergence tolerance in S. variegata involve enhanced oxidative stress responses, strengthened ethylene and ABA signaling, synthesis of raffinose family oligosaccharides, unsaturated fatty acids, and specific stress-related amino acids. Salix variegata Franch. is a riparian shrub species that can tolerate long-term complete submergence; however, the molecular mechanisms underlying this trait remain to be elucidated. In this study, we subjected S. variegata plants to complete submergence for 60 d and collected stems to perform transcriptomic and metabolomic analyses, as well as quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assays. Results revealed that photosynthesis and the response to light stimulus were inhibited during submergence and recovered after desubmergence. Ethylene and abscisic acid (ABA) signaling could be important for the long-term submergence tolerance of S. variegata. Jasmonic acid (JA) signaling also participated in the response to submergence. Raffinose family oligosaccharides, highly unsaturated fatty acids, and specific stress-related amino acids accumulated in response to submergence, indicating that they may protect plants from submergence damage, as they do in response to other abiotic stressors. Several organic acids were produced in S. variegata plants after submergence, which may facilitate coping with the toxicity induced by submergence. After long-term submergence, cell wall reorganization and phenylpropanoid metabolic processes (the synthesis of specific phenolics and flavonoids) were activated, which may contribute to long-term S. variegata submergence tolerance; however, the detailed mechanisms require further investigation. Several transcription factors (TFs), such as MYB, continuously responded to submergence, indicating that they may play important roles in the responses and adaption to submergence. Genes related to oxidative stress tolerance were specifically expressed after desubmergence, potentially contributing to recovery of S. variegata plants within a short period of time.

MicroRNA6443-mediated regulation of FERULATE 5-HYDROXYLASE gene alters lignin composition and enhances saccharification in Populus tomentosa.

Ferulate 5-hydroxylase (F5H) is a limiting enzyme involved in biosynthesizing sinapyl (S) monolignol in angiosperms. Genetic regulation of F5H can influence S monolignol synthesis and therefore improve saccharification efficiency and biofuel production. To date, little is known about whether F5H is post-transcriptionally regulated by endogenous microRNAs (miRNAs) in woody plants. Here, we report that a microRNA, miR6443, specifically regulates S lignin biosynthesis during stem development in Populus tomentosa. In situ hybridization showed that miR6443 is preferentially expressed in vascular tissues. We further identified that F5H2 is the direct target of miR6443. Overexpression of miR6443 decreased the transcript level of F5H2 in transgenic plants, resulting in a significant reduction in S lignin content. Conversely, reduced miR6443 expression by short tandem target mimics (STTM) elevated F5H2 transcripts, therefore increasing S lignin composition. Introduction of a miR6443-resistant form of F5H2 into miR6443-overexpression plants restored lignin ectopic composition, supporting that miR6443 specifically regulated S lignin biosynthesis by repressing F5H2 in P. tomentosa. Furthermore, saccharification assays revealed decreased hexose yields by 7.5-24.5% in miR6443-overexpression plants compared with the wild-type control, and increased hexoses yields by 13.2-14.6% in STTM6443-overexpression plants. Collectively, we demonstrate that miR6443 modulates S lignin biosynthesis by specially regulating F5H2 in P. tomentosa.

Sucrose Synthase Enhances Hull Size and Grain Weight by Regulating Cell Division and Starch Accumulation in Transgenic Rice.

Grain size and weight are two important determinants of grain yield in rice. Although overexpression of sucrose synthase (SUS) genes has led to several improvements on cellulose and starch-based traits in transgenic crops, little is reported about SUS enhancement of hull size and grain weight in rice. In this study, we selected transgenic rice plants that overexpressed OsSUS1-6 genes driven with the maize Ubi promoter. Compared to the controls (wild type and empty vector line), all independent OsSUS homozygous transgenic lines exhibited considerably increased grain yield and grain weights. Using the representative OsSUS3 overexpressed transgenic plants, four independent homozygous lines showed much raised cell numbers for larger hull sizes, consistent with their enhanced primary cell wall cellulose biosynthesis and postponed secondary wall synthesis. Accordingly, the OsSUS3 transgenic lines contained much larger endosperm volume and higher starch levels than those of the controls in the mature grains, leading to increased brown grain weights by 15-19%. Hence, the results have demonstrated that OsSUS overexpression could significantly improve hull size and grain weight by dynamically regulating cell division and starch accumulation in the transgenic rice.

Three AtCesA6-like members enhance biomass production by distinctively promoting cell growth in Arabidopsis.

Cellulose is an abundant biopolymer and a prominent constituent of plant cell walls. Cellulose is also a central component to plant morphogenesis and contributes the bulk of a plant's biomass. While cellulose synthase (CesA) genes were identified over two decades ago, genetic manipulation of this family to enhance cellulose production has remained difficult. In this study, we show that increasing the expression levels of the three primary cell wall AtCesA6-like genes (AtCesA2, AtCesA5, AtCesA6), but not AtCesA3, AtCesA9 or secondary cell wall AtCesA7, can promote the expression of major primary wall CesA genes to accelerate primary wall CesA complex (cellulose synthase complexes, CSCs) particle movement for acquiring long microfibrils and consequently increasing cellulose production in Arabidopsis transgenic lines, as compared with wild-type. The overexpression transgenic lines displayed changes in expression of genes related to cell growth and proliferation, perhaps explaining the enhanced growth of the transgenic seedlings. Notably, overexpression of the three AtCesA6-like genes also enhanced secondary cell wall deposition that led to improved mechanical strength and higher biomass production in transgenic mature plants. Hence, we propose that overexpression of certain AtCesA genes can provide a biotechnological approach to increase cellulose synthesis and biomass accumulation in transgenic plants.

Ectopic expression of a novel OsExtensin-like gene consistently enhances plant lodging resistance by regulating cell elongation and cell wall thickening in rice.

Plant lodging resistance is an important integrative agronomic trait of grain yield and quality in crops. Although extensin proteins are tightly associated with plant cell growth and cell wall construction, little has yet been reported about their impacts on plant lodging resistance. In this study, we isolated a novel extensin-like (OsEXTL) gene in rice, and selected transgenic rice plants that expressed OsEXTL under driven with two distinct promoters. Despite different OsEXTL expression levels, two-promoter-driven OsEXTL-transgenic plants, compared to a rice cultivar and an empty vector, exhibited significantly reduced cell elongation in stem internodes, leading to relatively shorter plant heights by 7%-10%. Meanwhile, the OsEXTL-transgenic plants showed remarkably thickened secondary cell walls with higher cellulose levels in the mature plants, resulting in significantly increased detectable mechanical strength (extension and pushing forces) in the mature transgenic plants. Due to reduced plant height and increased plant mechanical strength, the OsEXTL-transgenic plants were detected with largely enhanced lodging resistances in 3 years field experiments, compared to those of the rice cultivar ZH11. In addition, despite relatively short plant heights, the OsEXTL-transgenic plants maintain normal grain yields and biomass production, owing to their increased cellulose levels and thickened cell walls. Hence, this study demonstrates a largely improved lodging resistance in the OsEXTL-transgenic rice plants, and provides insights into novel extensin functions in plant cell growth and development, cell wall network construction and wall structural remodelling.

AtCSLD3 and GhCSLD3 mediate root growth and cell elongation downstream of the ethylene response pathway in Arabidopsis.

CSLD3, a gene of the cellulose synthase-like D family, affects root hair elongation, but its interactions with ethylene signaling and phosphate-starvation are poorly understood. Here, we aim to understand the role of CSLD3 in the context of the ethylene signaling and phosphate starvation pathways in Arabidopsis plant growth. Therefore, we performed a comparative analysis of the csld3-1 mutant, CSLD3-overexpressing lines, and ethylene-response mutants, such as the constitutive ethylene-response mutant i-ctr1. We found that CSLD3 overexpression enhanced root and hypocotyl growth by increasing cell elongation, and that the root growth was highly sensitive to ethylene treatment (1 µM ACC), in particular under phosphate starvation. However, the CSLD3-mediated hypocotyl elongation occurred independently of the ethylene signaling pathway. Notably, the typical induction of root hair and root elongation by ethylene and phosphate-starvation was completely abolished in the csld3-1 mutant. Furthermore, i-ctr1 csld3-1 double-mutants were hairless like the csld3-1 parent, confirming that CSLD3 acts downstream of the ethylene signaling pathway during root growth. Moreover, the CSLD3 levels positively correlated with cellulose levels, indicating a role of CSLD3 in cellulose synthesis, which may explain the observed growth effects. Our results establish how CSLD3 works in the context of the ethylene signaling and phosphate-starvation pathways during root hair growth, cell elongation, and cell wall biosynthesis.

Analysis of five rice 4-coumarate:coenzyme A ligase enzyme activity and stress response for potential roles in lignin and flavonoid biosynthesis in rice.

4-Coumarate:coenzyme A ligase (4CL) catalyzes the conversion of hydroxycinnamates into corresponding CoA esters for biosynthesis of flavonoids and lignin. In this study, five members of the 4CL gene family from rice were cloned and analyzed. Recombinant 4CL data revealed that 4-coumaric acid and ferulic acid were the two main substrates of 4CL (Os4CL1/3/4/5) for monolignol biosynthesis in rice. Os4CL2 was specifically expressed in the anther and was strongly activated by UV irradiation, suggesting its potential involvement in flavonoid formation. Moreover, bioinformatics analysis showed that the existence of valine residue at the substrate-binding pocket may mainly affect rice 4CL activities toward sinapic acid.

Overexpression of PtoMYB115 improves lignocellulose recalcitrance to enhance biomass digestibility and bioethanol yield by specifically regulating lignin biosynthesis in transgenic poplar.

BACKGROUND: Woody plants provide the most abundant biomass resource that is convertible for biofuels. Since lignin is a crucial recalcitrant factor against lignocellulose hydrolysis, genetic engineering of lignin biosynthesis is considered as a promising solution. Many MYB transcription factors have been identified to involve in the regulation of cell wall formation or phenylpropanoid pathway. In a previous study, we identified that PtoMYB115 contributes to the regulation of proanthocyanidin pathway, however, little is known about its role in lignocellulose biosynthesis and biomass saccharification in poplar. RESULTS: Here, we detected the changes of cell wall features and examined biomass enzymatic saccharification for bioethanol production under various chemical pretreatments in PtoMYB115 transgenic plants. We reported that PtoMYB115 might specifically regulate lignin biosynthesis to affect xylem development. Overexpression of PtoMYB115 altered lignin biosynthetic gene expression, resulting in reduced lignin deposition, raised S/G and beta-O-4 linkage, resulting in a significant reduction in cellulase adsorption with lignin and an increment in cellulose accessibility. These alterations consequently improved lignocellulose recalcitrance for significantly enhanced biomass saccharification and bioethanol yield in the PtoMYB115-OE transgenic lines. In contrast, the knockout of PtoMYB115 by CRISPR/Cas9 showed reduced woody utilization under various chemical pretreatments. CONCLUSIONS: This study shows that PtoMYB115 plays an important role in specifically regulating lignin biosynthesis and improving lignocellulose features. The enhanced biomass saccharification and bioethanol yield in the PtoMYB115-OE lines suggests that PtoMYB115 is a candidate gene for genetic modification to facilitate the utilization of biomass.

LACCASE14 is required for the deposition of guaiacyl lignin and affects cell wall digestibility in poplar.

BACKGROUND: The recalcitrance of lignocellulosic biomass provided technical and economic challenges in the current biomass conversion processes. Lignin is considered as a crucial recalcitrance component in biomass utilization. An in-depth understanding of lignin biosynthesis can provide clues to overcoming the recalcitrance. Laccases are believed to play a role in the oxidation of lignin monomers, leading to the formation of higher-order lignin. In plants, functions of only a few laccases have been evaluated, so little is known about the effect of laccases on cell wall structure and biomass saccharification. RESULTS: In this study, we screened a gain-of-function mutant with a significant increase in lignin content from Arabidopsis mutant lines overexpressing a full-length poplar cDNA library. Further analysis confirmed that a Chinese white poplar (Populus tomentosa) laccase gene PtoLAC14 was inserted into the mutant, and PtoLAC14 could functionally complement the Arabidopsis lac4 mutant. Overexpression of PtoLAC14 promoted the lignification of poplar and reduced the proportion of syringyl/guaiacyl. In contrast, the CRISPR/Cas9-generated mutation of PtLAC14 results in increased the syringyl/guaiacyl ratios, which led to integrated enhancement on biomass enzymatic saccharification. Notably, the recombinant PtoLAC14 protein showed higher oxidized efficiency to coniferyl alcohol (precursor of guaiacyl unit) in vitro. CONCLUSIONS: This study shows that PtoLAC14 plays an important role in the oxidation of guaiacyl deposition on cell wall. The reduced recalcitrance of the PtoLAC14-KO lines suggests that PtoLAC14 is an elite target for cell wall engineering, and genetic manipulation of this gene will facilitate the utilization of lignocellulose.

Brassinosteroid overproduction improves lignocellulose quantity and quality to maximize bioethanol yield under green-like biomass process in transgenic poplar.

BACKGROUND: As a leading biomass feedstock, poplar plants provide enormous lignocellulose resource convertible for biofuels and bio-chemicals. However, lignocellulose recalcitrance particularly in wood plants, basically causes a costly bioethanol production unacceptable for commercial marketing with potential secondary pollution to the environment. Therefore, it becomes important to reduce lignocellulose recalcitrance by genetic modification of plant cell walls, and meanwhile to establish advanced biomass process technology in woody plants. Brassinosteroids, plant-specific steroid hormones, are considered to participate in plant growth and development for biomass production, but little has been reported about brassinosteroids roles in plant cell wall assembly and modification. In this study, we generated transgenic poplar plant that overexpressed DEETIOLATED2 gene for brassinosteroids overproduction. We then detected cell wall feature alteration and examined biomass enzymatic saccharification for bioethanol production under various chemical pretreatments. RESULTS: Compared with wild type, the PtoDET2 overexpressed transgenic plants contained much higher brassinosteroids levels. The transgenic poplar also exhibited significantly enhanced plant growth rate and biomass yield by increasing xylem development and cell wall polymer deposition. Meanwhile, the transgenic plants showed significantly improved lignocellulose features such as reduced cellulose crystalline index and degree of polymerization values and decreased hemicellulose xylose/arabinose ratio for raised biomass porosity and accessibility, which led to integrated enhancement on biomass enzymatic saccharification and bioethanol yield under various chemical pretreatments. In contrast, the CRISPR/Cas9-generated mutation of PtoDET2 showed significantly lower brassinosteroids level for reduced biomass saccharification and bioethanol yield, compared to the wild type. Notably, the optimal green-like pretreatment could even achieve the highest bioethanol yield by effective lignin extraction in the transgenic plant. Hence, this study proposed a mechanistic model elucidating how brassinosteroid regulates cell wall modification for reduced lignocellulose recalcitrance and increased biomass porosity and accessibility for high bioethanol production. CONCLUSIONS: This study has demonstrated a powerful strategy to enhance cellulosic bioethanol production by regulating brassinosteroid biosynthesis for reducing lignocellulose recalcitrance in the transgenic poplar plants. It has also provided a green-like process for biomass pretreatment and enzymatic saccharification in poplar and beyond.

Distinct cellulose and callose accumulation for enhanced bioethanol production and biotic stress resistance in OsSUS3 transgenic rice.

Genetic modification of plant cell walls is an effective approach to reduce lignocellulose recalcitrance in biofuel production, but it may affect plant stress response. Hence, it remains a challenge to reduce biomass recalcitrance and simultaneously enhance stress resistance. In this study, the OsSUS3-transgenic plants exhibited increased cell wall polysaccharides deposition and reduced cellulose crystallinity and xylose/arabinose proportion of hemicellulose, resulting in largely enhanced biomass saccharification and bioethanol production. Additionally, strengthening of the cell wall also contributed to plant biotic resistance. Notably, the transgenic plants increased stress-induced callose accumulation, and promoted the activation of innate immunity, leading to greatly improved multiple resistances to the most destructive diseases and a major pest. Hence, this study demonstrates a significant improvement both in bioethanol production and biotic stress resistance by regulating dynamic carbon partitioning for cellulose and callose biosynthesis in OsSUS3-transgenic plants. Meanwhile, it also provides a potential strategy for plant cell wall modification.

AtCesA8-driven OsSUS3 expression leads to largely enhanced biomass saccharification and lodging resistance by distinctively altering lignocellulose features in rice.

BACKGROUND: Biomass recalcitrance and plant lodging are two complex traits that tightly associate with plant cell wall structure and features. Although genetic modification of plant cell walls can potentially reduce recalcitrance for enhancing biomass saccharification, it remains a challenge to maintain a normal growth with enhanced biomass yield and lodging resistance in transgenic plants. Sucrose synthase (SUS) is a key enzyme to regulate carbon partitioning by providing UDP-glucose as substrate for cellulose and other polysaccharide biosynthesis. Although SUS transgenic plants have reportedly exhibited improvement on the cellulose and starch based traits, little is yet reported about SUS impacts on both biomass saccharification and lodging resistance. In this study, we selected the transgenic rice plants that expressed OsSUS3 genes when driven by the AtCesA8 promoter specific for promoting secondary cell wall cellulose synthesis in Arabidopsis. We examined biomass saccharification and lodging resistance in the transgenic plants and detected their cell wall structures and wall polymer features. RESULTS: During two-year field experiments, the selected AtCesA8::SUS3 transgenic plants maintained a normal growth with slightly increased biomass yields. The four independent transgenic lines exhibited much higher biomass enzymatic saccharification and bioethanol production under chemical pretreatments at P < 0.01 levels, compared with the controls of rice cultivar and empty vector transgenic line. Notably, all transgenic lines showed a consistently enhanced lodging resistance with the increasing extension and pushing forces. Correlation analysis suggested that the reduced cellulose crystallinity was a major factor for largely enhanced biomass saccharification and lodging resistance in transgenic rice plants. In addition, the cell wall thickenings with the increased cellulose and hemicelluloses levels should also contribute to plant lodging resistance. Hence, this study has proposed a mechanistic model that shows how OsSUS3 regulates cellulose and hemicelluloses biosyntheses resulting in reduced cellulose crystallinity and increased wall thickness, thereby leading to large improvements of both biomass saccharification and lodging resistance in transgenic rice plants. CONCLUSIONS: This study has demonstrated that the AtCesA8::SUS3 transgenic rice plants exhibited largely improved biomass saccharification and lodging resistance by reducing cellulose crystallinity and increasing cell wall thickness. It also suggests a powerful genetic approach for cell wall modification in bioenergy crops.

Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology.

Distinct Geographical Distribution of the Miscanthus Accessions with Varied Biomass Enzymatic Saccharification.

Miscanthus is a leading bioenergy candidate for biofuels, and it thus becomes essential to characterize the desire natural Miscanthus germplasm accessions with high biomass saccharification. In this study, total 171 natural Miscanthus accessions were geographically mapped using public database. According to the equation [P(H/L| East) = P(H/L∩East)/P(East)], the probability (P) parameters were calculated on relationships between geographical distributions of Miscanthus accessions in the East of China, and related factors with high(H) or low(L) values including biomass saccahrification under 1% NaOH and 1% H2SO4 pretreatments, lignocellulose features and climate conditions. Based on the maximum P value, a golden cutting line was generated from 42°25' N, 108°22' E to 22°58' N, 116°28' E on the original locations of Miscanthus accessions with high P(H|East) values (0.800-0.813), indicating that more than 90% Miscanthus accessions were originally located in the East with high biomass saccharification. Furthermore, the averaged insolation showed high P (H|East) and P(East|H) values at 0.782 and 0.754, whereas other climate factors had low P(East|H) values, suggesting that the averaged insolation is unique factor on Miscanthus distributions for biomass saccharification. In terms of cell wall compositions and wall polymer features, both hemicelluloses level and cellulose crystallinity (CrI) of Miscanthus accessions exhibited relative high P values, suggesting that they should be the major factors accounting for geographic distributions of Miscanthus accessions with high biomass digestibility.

Positive selection drives adaptive diversification of the 4-coumarate: CoA ligase (4CL) gene in angiosperms.

Lignin and flavonoids play a vital role in the adaption of plants to a terrestrial environment. 4-Coumarate: coenzyme A ligase (4CL) is a key enzyme of general phenylpropanoid metabolism which provides the precursors for both lignin and flavonoids biosynthesis. However, very little is known about how such essential enzymatic functions evolve and diversify. Here, we analyze 4CL sequence variation patterns in a phylogenetic framework to further identify the evolutionary forces that lead to functional divergence. The results reveal that lignin-biosynthetic 4CLs are under positive selection. The majority of the positively selected sites are located in the substrate-binding pocket and the catalytic center, indicating that nonsynonymous substitutions might contribute to the functional evolution of 4CLs for lignin biosynthesis. The evolution of 4CLs involved in flavonoid biosynthesis is constrained by purifying selection and maintains the ancestral role of the protein in response to biotic and abiotic factors. Overall, our results demonstrate that protein sequence evolution via positive selection is an important evolutionary force driving adaptive diversification in 4CL proteins in angiosperms. This diversification is associated with adaption to a terrestrial environment.

A near infrared spectroscopic assay for stalk soluble sugars, bagasse enzymatic saccharification and wall polymers in sweet sorghum.

In this study, 123 sweet sorghum (Sorghum bicolor L.) accessions and 50 mutants were examined with diverse stalk soluble sugars, bagasse enzymatic saccharification and wall polymers, indicating the potential near infrared spectroscopy (NIRS) assay for those three important parameters. Using the calibration and validation sets and modified squares method, nine calibration optimal equations were generated with high determination coefficient on the calibration (R(2)) (0.81-0.99), cross-validation (R(2)cv) (0.77-0.98), and the ratio performance deviation (RPD) (2.07-7.45), which were at first time applied by single spectra for simultaneous assay of stalk soluble sugars, bagasse hydrolyzed sugars, and three major wall polymers in bioenergy sweet sorghum.

Lignin extraction distinctively enhances biomass enzymatic saccharification in hemicelluloses-rich Miscanthus species under various alkali and acid pretreatments.

In this study, one- and two-step pretreatments with alkali and acid were performed in the three Miscanthus species that exhibit distinct hemicelluloses levels. As a result, one-step with 4% NaOH or two-step with 2% NaOH and 1% H2SO4 was examined to be optimal for high biomass saccharification, indicating that alkali was the main effecter of pretreatments. Notably, both one- and two-step pretreatments largely enhanced biomass digestibility distinctive in hemicelluloses-rich samples by effectively co-extracting hemicelluloses and lignin. However, correlation analysis further indicated that the effective lignin extraction, other than the hemicelluloses removals, predominately determined biomass saccharification under various alkali and acid pretreatments, leading to a significant alteration of cellulose crystallinity. Hence, this study has suggested the potential approaches in bioenergy crop breeding and biomass process technology.

Sugar-rich sweet sorghum is distinctively affected by wall polymer features for biomass digestibility and ethanol fermentation in bagasse.

Sweet sorghum has been regarded as a typical species for rich soluble-sugar and high lignocellulose residues, but their effects on biomass digestibility remain unclear. In this study, we examined total 63 representative sweet sorghum accessions that displayed a varied sugar level at stalk and diverse cell wall composition at bagasse. Correlative analysis showed that both soluble-sugar and dry-bagasse could not significantly affect lignocellulose saccharification under chemical pretreatments. Comparative analyses of five typical pairs of samples indicated that DP of crystalline cellulose and arabinose substitution degree of non-KOH-extractable hemicelluloses distinctively affected lignocellulose crystallinity for high biomass digestibility. By comparison, lignin could not alter lignocellulose crystallinity, but the KOH-extractable G-monomer predominately determined lignin negative impacts on biomass digestions, and the G-levels released from pretreatments significantly inhibited yeast fermentation. The results also suggested potential genetic approaches for enhancing soluble-sugar level and lignocellulose digestibility and reducing ethanol conversion inhibition in sweet sorghum.