The role of ascorbate metabolism and cell wall degradation in atemoya peel browning: insights from multi-omics analysis under modified atmosphere packaging

The identification of DNA polymorphisms associated to increased cell wall (CW) degradability could accelerate the development of alfalfa (Medicago sativa L.) cultivars with superior ethanol conversion yields. Genotypes with high (D+) or low (D−) CW degradability were recently identified within a biomass-type and three winter-hardy-type populations using near-infrared reflectance spectroscopy (NIRS) prediction of CW glucose released by enzyme saccharification. In this report, we used sequence-related amplified polymorphism to search for DNA variations associated to differences in enzyme-released glucose. A bulk segregant analysis (BSA) of pooled DNA samples (20 plants/bulk) from D+, D−, and randomly chosen genotypes uncovered polymorphisms associated to CW degradability. Polymorphisms that increase or decrease in intensity between D+ and D− bulks indicated the presence of genomic regions with either positive or negative effects on CW degradability. A primer pair (Me4-R14) generated a fragment, which increased in intensity in the D+ bulk of the biomass-type population. Conversely, the amplification of that fragment declined in the D+ bulks of the winter-hardy-type populations. Interestingly, these populations differ in their degradability. Assessment of the genotypic occurrence of this fragment confirmed that polymorphism detected with BSA reflects changes in the frequency of occurrence within populations. Sequence analysis of the Me4-R14 fragment revealed homologies with sequences from Medicago truncatula, a model species for legumes with documented synteny with M. sativa. Our results show that genomic regions associated to CW degradability can be identified using the combination of BSA of genotypes with contrasted degradability and a PCR-based amplification technique.

Efficient and cost-effective conversion of plant biomass to usable forms of energy requires a thorough understanding of cell wall biosynthesis, modification and degradation. To elucidate these processes, we assessed the expression dynamics during enzymatic removal and regeneration of rice cell walls in suspension cells over time. In total, 928 genes exhibited significant up-regulation during cell wall removal, whereas, 79 genes were up-regulated during cell wall regeneration. Both gene sets are enriched for kinases, transcription factors and genes predicted to be involved in cell wall-related functions. Integration of the gene expression datasets with a catalog of known and/or predicted biochemical pathways from rice, revealed metabolic and hormonal pathways involved in cell wall degradation and regeneration. Rice lines carrying Tos17 mutations in genes up-regulated during cell wall removal exhibit dwarf phenotypes. Many of the genes up-regulated during cell wall development are also up-regulated in response to infection and environmental perturbations indicating a coordinated response to diverse types of stress.

In order to contribute to the inventory of genomic areas involved in maize cell wall lignification and degradability, QTL analyses were investigated in a RIL progeny between an old Minnesota13 dent line (WM13) and a modern Iodent line (RIo). Significant variation for agronomic- and cell wall-related traits was observed for the RIL per se (plants without ears) and topcross (whole plants) experiments after crossing with both old (Ia153) and modern tester (RFl) lines. Three QTLs for stover (plant without ear) yield were observed in per se experiments, with alleles increasing yield originating from RIo in two genomic locations with the highest effects. However, no QTL for whole plant yield was detected in topcross experiments, despite the fact that two QTLs for starch content were shown with increasing alleles originating from the modern RIo line. Fifteen lignin QTLs were shown, including a QTL for Klason lignins in per se experiments, located in bin 2.04, which explained 43 % of the observed genetic variation. Thirteen QTLs for p-hydroxycinnamic acid contents and nine QTLs related to the monomeric composition of lignin were shown in per se experiments, with syringaldehyde and diferulate QTLs explaining nearly 25 % of trait variations. Nine and seven QTLs for cell wall digestibility were mapped in per se and topcross experiments, respectively. Five of the per se QTLs explained more than 15 % of the variation, up to nearly 25 %. QTL positions in bins 2.06, 5.04, 5.08 and 8.02 for ADL/NDF, IVNDFD, lignin structure and/or p-hydroxycinnamic acid contents have not been previously shown and were thus first identified in the RIo × WM13 progeny. Based on QTL colocalizations, differences in cell wall degradability between RIo and WM13 were less often related to acid detergent lignin (ADL) content than in previous RIL investigations. QTL colocalizations then highlighted the probable importance of ferulate cross linkages in variation for cell wall digestibility. No colocalizations of QTL for cell wall phenolic-related traits were shown with genes involved in monolignol biosynthesis or polymerization. In contrast, colocalizations were most often shown with MYB and NAC transcription factors, including orthologs of master genes involved in Arabidopsis secondary wall assembly. QTL colocalizations also strengthened the probable involvement of members of the CoA-dependent acyltransferase PF02458 family in the feruloylation of arabinoxylan chains.

Various perennial C4 grass species have tremendous potential for use as lignocellulosic biofuel feedstocks. Currently available grasses require costly pre-treatment and exogenous hydrolytic enzyme application to break down complex cell wall polymers into sugars that can then be fermented into ethanol. It has long been hypothesized that engineered feedstock production of cell wall degrading (CWD) enzymes would be an efficient production platform for of exogenous hydrolytic enzymes. Most research has focused on plant overexpression of CWD enzyme-coding genes from free-living bacteria and fungi that naturally break down plant cell walls. Recently, it has been found that insect digestive tracts harbor novel sources of lignocellulolytic biocatalysts that might be exploited for biofuel production. These CWD enzyme genes can be located in the insect genomes or in symbiotic microbes. When CWD genes are transformed into plants, negative pleiotropic effects are possible such as unintended cell wall digestion. The use of codon optimization along with organelle and tissue specific targeting improves CWD enzyme yields. The literature teaches several important lessons on strategic deployment of CWD genes in transgenic plants, which is the focus of this review.

Effect of gibberellic acid on cell wall degradation and softening in postharvest okras

Akebia trifoliata fruit is prone to crack after ripening, but little is known about the mechanism underlying the cracking process. This study integrated transcriptomic and metabolomic data, revealing significant changes in 398 metabolites and 8414 genes during ripening and cracking, mainly impacting cell-wall metabolism. Multi-omics joint analysis indicated that genes related to polygalacturonase, pectate lyase, α-amylase, and glycogen phosphorylase were up-regulated after cracking, degrading cell wall and starch. Concurrently, diminished photosynthetic metabolism and heightened phenylpropanoid metabolism suggested alterations in cuticle structure, potentially impacting cell-wall robustness. Numerous auxin and abscisic acid signaling-related genes were expressed, and we assume that they contributed to the promoting peel growth. These alterations collectively might compromise peel strength and elevate expanding pressure, potentially leading to A. trifoliata cracking. Transcription factors, predominantly ethylene response factors and helix-loop-helix family members, appeared to regulate these metabolic shifts. These findings provide valuable insights into A. trifoliata cracking mechanisms; however, direct experimental validation of these assumptions is necessary to strengthen these conclusions and expedite their commercial utilization.

Drought-induced cell wall degradation in the base of pedicel is associated with accelerated cotton square shedding

The enzymatic degradation of algal cell walls was investigated using two experimental approaches. Algal cell walls contain, among other compounds, cellulose and phospholipids. Phospholipids are also present in the membranes of some organelles, particularly the lipid globule of some algae. The desired reactions were the conversion of cell wall cellulose to glucose and the conversion of membrane phospholipids to fatty acids and glycerol. Enzyme degradation was carried out using whole cell dead algae and pre-lysed, pretreated (diluted in 1.5% sulfuric acid and autoclaved) mixed culture algae. The pre-lysed, pretreated feedstock showed the formation of glycerol when incubated with lipases. Formation of glucose was also observed when incubated with cellulases. Upon culturing in the presence of yeast, complete consumption of glucose was observed, however ethanol could not be detected. Such technology could be used in the production of ethanol or biodiesel.

Effect of the partial degradation of endosperm cell walls on the cooking properties of rice grains and also on the texture of cooked rice was examined by using two different enzyme preparations, i.e., crude enzyme from Aspergillus japonicus and purified endoxylanase from Streptomyces sp. E-86. Texturometer analysis showed that the partial degradation of the cell walls softened the cooked rice grains and made them more sticky. When the excess water was supplied, cooked rice grains expanded more after the enzyme treatment. Moreover, this treatment affected the amylogram of milled rice flour and lowered its viscosity, especially in the retrogradation process. Purified endoxylanase showed these effect without any other enzyme, suggesting the important role of hemicellulosic polysaccharides for the physical properties of endosperm cell walls.

Reserve mobilization dynamics and degradation pattern of mannan-rich cell walls in the recalcitrant seed of Mauritia flexuosa (Arecaceae)

Carbohydrate-active enzymes (CAZymes) are involved in the metabolism of glycoconjugates, oligosaccharides, and polysaccharides and, in the case of plant pathogens, in the degradation of the host cell wall and storage compounds. We performed an in silico analysis of CAZymes predicted from the genomes of seven Pythium species (Py. aphanidermatum, Py. arrhenomanes, Py. irregulare, Py. iwayamai, Py. ultimum var. ultimum, Py. ultimum var. sporangiiferum and Py. vexans) using the “CAZymes Analysis Toolkit” and “Database for Automated Carbohydrate-active Enzyme Annotation” and compared them to previously published oomycete genomes. Growth of Pythium spp. was assessed in a minimal medium containing selected carbon sources that are usually present in plants. The in silico analyses, coupled with our in vitro growth assays, suggest that most of the predicted CAZymes are involved in the metabolism of the oomycete cell wall with starch and sucrose serving as the main carbohydrate sources for growth of these plant pathogens. The genomes of Pythium spp. also encode pectinases and cellulases that facilitate degradation of the plant cell wall and are important in hyphal penetration; however, the species examined in this study lack the requisite genes for the complete saccharification of these carbohydrates for use as a carbon source. Genes encoding for xylan, xyloglucan, (galacto)(gluco)mannan and cutin degradation were absent or infrequent in Pythium spp.. Comparative analyses of predicted CAZymes in oomycetes indicated distinct evolutionary histories. Furthermore, CAZyme gene families among Pythium spp. were not uniformly distributed in the genomes, suggesting independent gene loss events, reflective of the polyphyletic relationships among some of the species.

Genomics and Molecular Genetics of Plant-Nematode Interactions

The Sigatoka disease complex, caused by the closely-related Dothideomycete fungi Pseudocercospora musae (yellow sigatoka), Pseudocercospora eumusae (eumusae leaf spot), and Pseudocercospora fijiensis (black sigatoka), is currently the most devastating disease on banana worldwide. The three species emerged on bananas from a recent common ancestor and show clear differences in virulence, with P. eumusae and P. fijiensis considered the most aggressive. In order to understand the genomic modifications associated with shifts in the species virulence spectra after speciation, and to identify their pathogenic core that can be exploited in disease management programs, we have sequenced and analyzed the genomes of P. eumusae and P. musae and compared them with the available genome sequence of P. fijiensis. Comparative analysis of genome architectures revealed significant differences in genome size, mainly due to different rates of LTR retrotransposon proliferation. Still, gene counts remained relatively equal and in the range of other Dothideomycetes. Phylogenetic reconstruction based on a set of 46 conserved single-copy genes strongly supported an earlier evolutionary radiation of P. fijiensis from P. musae and P. eumusae. However, pairwise analyses of gene content indicated that the more virulent P. eumusae and P. fijiensis share complementary patterns of expansions and contractions in core gene families related to metabolism and enzymatic degradation of plant cell walls, suggesting that the evolution of virulence in these two pathogens has, to some extent, been facilitated by convergent changes in metabolic pathways associated with nutrient acquisition and assimilation. In spite of their common ancestry and shared host-specificity, the three species retain fairly dissimilar repertoires of effector proteins, suggesting that they likely evolved different strategies for manipulating the host immune system. Finally, 234 gene families, including seven putative effectors, were exclusively present in the three Sigatoka species, and could thus be related to adaptation to the banana host.

The rumen microbial ecosystem provides ruminants a selective advantage, the ability to utilize forages, allowing them to flourish worldwide in various environments. For many years, our understanding of the ruminal microbial ecosystem was limited to understanding the microbes (usually only laboratory-amenable bacteria) grown in pure culture, meaning that much of our understanding of ruminal function remained a "black box." However, the ruminal degradation of plant cell walls is performed by a consortium of bacteria, archaea, protozoa, and fungi that produces a wide variety of carbohydrate-active enzymes (CAZymes) that are responsible for the catabolism of cellulose, hemicellulose, and pectin. The past 15 years have seen the development and implementation of numerous next-generation sequencing (NGS) approaches (e.g., pyrosequencing, Illumina, and shotgun sequencing), which have contributed significantly to a greater level of insight regarding the microbial ecology of ruminants fed a variety of forages. There has also been an increase in the utilization of liquid chromatography and mass spectrometry that revolutionized transcriptomic approaches, and further improvements in the measurement of fermentation intermediates and end products have advanced with metabolomics. These advanced NGS techniques along with other analytic approaches, such as metaproteomics, have been utilized to elucidate the specific role of microbial CAZymes in forage degradation. Other methods have provided new insights into dynamic changes in the ruminal microbial population fed different diets and how these changes impact the assortment of products presented to the host animal. As more omics-based data has accumulated on forage-fed ruminants, the sequence of events that occur during fiber colonization by the microbial consortium has become more apparent, with fungal populations and fibrolytic bacterial populations working in conjunction, as well as expanding understanding of the individual microbial contributions to degradation of plant cell walls and polysaccharide components. In the future, the ability to predict microbial population and enzymatic activity and end products will be able to support the development of dynamic predictive models of rumen forage degradation and fermentation. Consequently, it is imperative to understand the rumen's microbial population better to improve fiber degradation in ruminants and, thus, stimulate more sustainable production systems.

Flooding can cause root hypoxia and can lead to significant agricultural losses. Therefore, understanding plant adaptations to flooding, including root aerenchyma development, is one important avenue for insuring future global food security. We investigated cell wall modifications during root aerenchyma formation in response to the prolonged 0–48 h flooding of Phaseolus coccineus, Pisum sativum, and Cicer arietinum seedlings. Using transmission electron microscopy, toluidine blue O (TBO) staining, and immunolabeling with antibodies targeting de-methyl-esterified homogalacturonan (DMEH), partially DMEH, and methyl-esterified homogalacturonan (MEH), we examined changes in cell wall composition. Transmission electron microscopy and TBO staining revealed degradation of cell walls and middle lamella, with accumulation of DMEH near flooding-induced aerenchyma cavities. Immunolabeling indicated increased DMEH epitope availability in flooded roots, suggesting a role in cell wall remodeling. Enzyme pretreatments, used to “unmask” homogalacturonan by removing cellulose and hemicellulose, revealed that specific forms of homogalacturonan, particularly DMEH complexed with calcium and MEH, are masked by these cell wall components. The study highlights the complex interplay of pectin, cellulose, and hemicellulose in cell wall degradation during aerenchyma development, providing insights into legume flooding stress responses.