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Reduced lignin content and altered lignin composition in the warm season forage grass Paspalum dilatatum by down-regulation of a Cinnamoyl CoA reductase gene.

ABSTRACT: C4 grasses are favoured as forage crops in warm, humid climates. The use of C4 grasses in pastures is expected to increase because the tropical belt is widening due to global climate change. While the forage quality of Paspalum dilatatum (dallisgrass) is higher than that of other C4 forage grass species, digestibility of warm-season grasses is, in general, poor compared with most temperate grasses. The presence of thick-walled parenchyma bundle-sheath cells around the vascular bundles found in the C4 forage grasses are associated with the deposition of lignin polymers in cell walls. High lignin content correlates negatively with digestibility, which is further reduced by a high ratio of syringyl (S) to guaiacyl (G) lignin subunits. Cinnamoyl-CoA reductase (CCR) catalyses the conversion of cinnamoyl CoA to cinnemaldehyde in the monolignol biosynthetic pathway and is considered to be the first step in the lignin-specific branch of the phenylpropanoid pathway. We have isolated three putative CCR1 cDNAs from P. dilatatum and demonstrated that their spatio-temporal expression pattern correlates with the developmental profile of lignin deposition. Further, transgenic P. dilatatum plants were produced in which a sense-suppression gene cassette, delivered free of vector backbone and integrated separately to the selectable marker, reduced CCR1 transcript levels. This resulted in the reduction of lignin, largely attributable to a decrease in G lignin.

SUBMITTER: Giordano A

PROVIDER: S-EPMC4010725 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND: Paspalum (Poaceae) is an important genus of the tribe Paniceae, which includes several species of economic importance for foraging, turf and ornamental purposes, and has a complex taxonomical classification. Because of the widespread interest in several species of this genus, many accessions have been conserved in germplasm banks and distributed throughout various countries around the world, mainly for the purposes of cultivar development and cytogenetic studies. Correct identification of germplasms and quantification of their variability are necessary for the proper development of conservation and breeding programs. Evaluation of microsatellite markers in different species of Paspalum conserved in a germplasm bank allowed assessment of the genetic differences among them and assisted in their proper botanical classification. RESULTS: Seventeen new polymorphic microsatellites were developed for Paspalum atratum Swallen and Paspalum notatum Flüggé, twelve of which were transferred to 35 Paspalum species and used to evaluate their variability. Variable degrees of polymorphism were observed within the species. Based on distance-based methods and a Bayesian clustering approach, the accessions were divided into three main species groups, two of which corresponded to the previously described Plicatula and Notata Paspalum groups. In more accurate analyses of P. notatum accessions, the genetic variation that was evaluated used thirty simple sequence repeat (SSR) loci and revealed seven distinct genetic groups and a correspondence of these groups to the three botanical varieties of the species (P. notatum var. notatum, P. notatum var. saurae and P. notatum var. latiflorum). CONCLUSIONS: The molecular genetic approach employed in this study was able to distinguish many of the different taxa examined, except for species that belong to the Plicatula group, which has historically been recognized as a highly complex group. Our molecular genetic approach represents a valuable tool for species identification in the initial assessment of germplasm as well as for characterization, conservation and successful species hybridization.

Project description:Cinnamoyl-CoA reductase (CCR) is the first committed enzyme in the monolignol pathway for lignin biosynthesis and catalyzes the conversion of hydroxycinnamoyl-CoAs into hydroxycinnamaldehydes. In the rice genome, 33 genes are annotated as CCR and CCR-like genes, collectively called OsCCRs. To elucidate the functions of OsCCRs, their phylogenetic relationships, expression patterns at the transcription levels and biochemical characteristics were thoroughly analyzed. Of the 33 OsCCRs, 24 of them encoded polypeptides of lengths similar to those of previously identified plant CCRs. The other nine OsCCRs had much shorter peptide lengths. Phylogenetic tree and sequence similarities suggested OsCCR4, 5, 17, 18, 19, 20, and 21 as likely candidates for functional CCRs in rice. To elucidate biochemical functions, OsCCR1, 5, 17, 19, 20, 21, and 26 were heterologously expressed in Escherichia coli and the resulting recombinant OsCCRs were purified to apparent homogeneity. Activity assays of the recombinant OsCCRs with hydroxycinnamoyl-CoAs revealed that OsCCR17, 19, 20, and 21 were biochemically active CCRs, in which the NAD(P)-binding and NADP-specificity motifs as well as the CCR signature motif were fully conserved. The kinetic parameters of enzyme reactions revealed that feruloyl-CoA, a precursor for the guaiacyl (G)-unit of lignin, is the most preferred substrate of OsCCR20 and 21. This result is consistent with a high content (about 70%) of G-units in rice lignins. Phylogenetic analysis revealed that OsCCR19 and 20 were grouped with other plant CCRs involved in developmental lignification, whereas OsCCR17 and 21 were closely related to stress-responsible CCRs identified from other plant species. In agreement with the phylogenetic analysis, expression analysis demonstrated that OsCCR20 was constitutively expressed throughout the developmental stages of rice, showing particularly high expression levels in actively lignifying tissues, such as roots and stems. These results suggest that OsCCR20 is primarily involved in developmental deposition of lignins in secondary cell walls. As expected, the expressions of OsCCR17 and 21 were induced in response to biotic and abiotic stresses, such as Magnaporthe grisea and Xanthomonas oryzae pv. oryzae (Xoo) infections, UV-irradiation and high salinity, suggesting that these genes play a role in defense-related processes in rice.

Project description:BackgroundDiet is a key driver of equine hindgut microbial community structure and composition. The aim of this study was to characterize shifts in the fecal microbiota of grazing horses during transitions between forage types within integrated warm- (WSG) and cool-season grass (CSG) rotational grazing systems (IRS). Eight mares were randomly assigned to two IRS containing mixed cool-season grass and one of two warm-season grasses: bermudagrass [Cynodon dactylon (L.) Pers.] or crabgrass [Digitaria sanguinalis (L.) Scop.]. Fecal samples were collected during transitions from CSG to WSG pasture sections (C-W) and WSG to CSG (W-C) on days 0, 2, 4, and 6 following pasture rotation and compared using 16S rRNA gene sequencing.ResultsRegardless of IRS or transition (C-W vs. W-C), species richness was greater on day 4 and 6 in comparison to day 0 (P < 0.05). Evenness, however, did not differ by day. Weighted UniFrac also did not differ by day, and the most influential factor impacting β-diversity was the individual horse (R2 ≥ 0.24; P = 0.0001). Random forest modeling was unable to accurately predict days within C-W and W-C, but could predict the individual horse based on microbial composition (accuracy: 0.92 ± 0.05). Only three differentially abundant bacterial co-abundance groups (BCG) were identified across days within all C-W and W-C for both IRS (W ≥ 126). The BCG differing by day for all transitions included amplicon sequence variants (ASV) assigned to bacterial groups with known fibrolytic and butyrate-producing functions including members of Lachnospiraceae, Clostridium sensu stricto 1, Anaerovorax the NK4A214 group of Oscillospiraceae, and Sarcina maxima. In comparison, 38 BCG were identified as differentially abundant by horse (W ≥ 704). The ASV in these groups were most commonly assigned to genera associated with degradation of structural carbohydrates included Rikenellaceae RC9 gut group, Treponema, Christensenellaceae R-7 group, and the NK4A214 group of Oscillospiraceae. Fecal pH also did not differ by day.ConclusionsOverall, these results demonstrated a strong influence of individual horse on the fecal microbial community, particularly on the specific composition of fiber-degraders. The equine fecal microbiota were largely stable across transitions between forages within IRS suggesting that the equine gut microbiota adjusted at the individual level to the subtle dietary changes imposed by these transitions. This adaptive capacity indicates that horses can be managed in IRS without inducing gastrointestinal dysfunction.

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Reduced lignin content and altered lignin composition in the warm season forage grass Paspalum dilatatum by down-regulation of a Cinnamoyl CoA reductase gene.

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