Analysis of gene mutation in plant cell wall by dielectric relaxation

Arabidopsis Thaliana is a plant composed mainly of cellulose and lignin. Geneticists need techniques able to make differences at the molecular level between modified plants (DML6, CAD C/D) and non-modified ones. Thermo-stimulated current (TSC) analysis is a promising route to identify gene mutations. For the non-modified plant, at low temperatures, TSC thermograms highlight three dielectric relaxation modes. From −150 to −110 ◦C, γCellulose is attributed to CH2OH and –OH groups of cellulose. Between −110 and −80 ◦C, βLignin is detected. From −80 to −40 ◦C, βCellulose is characteristic of the molecular mobility of glycosidic linkages. For the CAD C/D modified plants, only γCellulose and βLignin are observed; due to analogous enthalpy values, those modes have the same molecular origin as in the non-modified plant. So, the βLignin mode is associated with the molecular mobility of the lignin-OH groups. The CAD C/D gene mutation changes the chemical structure of lignin, which promotes hydrogen bonds in the network and inhibits molecular mobility of glucosidic rings. It is also interesting to note that the DML6 gene mutation induces a higher cooperativity of this βCellulose relaxation than in wild vegetal composites. In fact, this mutation promotes molecular mobility of glycosidic rings thanks to β1–4 glycosidic linkages

Conifer R2R3-MYB transcription factors: sequence analyses and gene expression in wood-forming tissues of white spruce (Picea glauca)

BACKGROUND: Several members of the R2R3-MYB family of transcription factors act as regulators of lignin and phenylpropanoid metabolism during wood formation in angiosperm and gymnosperm plants. The angiosperm Arabidopsis has over one hundred R2R3-MYBs genes; however, only a few members of this family have been discovered in gymnosperms. RESULTS: We isolated and characterised full-length cDNAs encoding R2R3-MYB genes from the gymnosperms white spruce, Picea glauca (13 sequences), and loblolly pine, Pinus taeda L. (five sequences). Sequence similarities and phylogenetic analyses placed the spruce and pine sequences in diverse subgroups of the large R2R3-MYB family, although several of the sequences clustered closely together. We searched the highly variable C-terminal region of diverse plant MYBs for conserved amino acid sequences and identified 20 motifs in the spruce MYBs, nine of which have not previously been reported and three of which are specific to conifers. The number and length of the introns in spruce MYB genes varied significantly, but their positions were well conserved relative to angiosperm MYB genes. Quantitative RTPCR of MYB genes transcript abundance in root and stem tissues revealed diverse expression patterns; three MYB genes were preferentially expressed in secondary xylem, whereas others were preferentially expressed in phloem or were ubiquitous. The MYB genes expressed in xylem, and three others, were up-regulated in the compression wood of leaning trees within 76 hours of induction. CONCLUSION: Our survey of 18 conifer R2R3-MYB genes clearly showed a gene family structure similar to that of Arabidopsis. Three of the sequences are likely to play a role in lignin metabolism and/or wood formation in gymnosperm trees, including a close homolog of the loblolly pine PtMYB4, shown to regulate lignin biosynthesis in transgenic tobacco

Investigation of Argania spinosa L. (Skeels) polyphenols growing in arid and semi-arid conditions

Argania spinosa L. Skeels, belonging to the Argania genus of the Sapotaceae family, is a species native to Morocco and Algeria. Due to its perfect adaptation to soil and climate, this tree plays an important ecological role in a constantly threatened encroached desert region. To understand the biological role of polyphenols in making the argan tree adapts to its natural habitat, we conducted a comparative study in two of the tree development stations: Tindouf located in South-western Algeria and Stidia located in Northwest Algeria. High performance liquid chromatography (HPLC) and Gas chromatography-mass spectrometry (GC-MS) used for the analysis of flavonoids led to the identification of six flavonols (two types of myricetin, rutin, hyperoside, quercetin, kaempferol) in the leaves of Tindouf argan tree, and two molecules (myricetin and quercetin) in the leaves of Stidia argan tree. Other molecules presented are few. The determination of flavonoids by spectrophotometry revealed the richness of Tindouf argan in these compounds (20%) compared to that of Stidia argan tree (8.7%). Histolocalisation of the flavonoids, tannins and anthraquinone in the leaves and stems of the tree was done using fluorescence microscope to understand the role of these molecules in the protection of this tree in its environment.Key words: Aragania spinosa L. (Skeels), histolocalisation, polyphenols, high performance liquid chromatography (HPLC), gas chromatography-mass spectrometry (GC-MS)

Breeding grasses for capacity to biofuel production or silage feeding value: an updated list of genes involved in maize secondary cell wall biosynthesis and assembly

In the near future, maize, sorghum, or switchgrass stovers and cereal straws will be a significant source of carbohydrates for sustainable biofuel production, in addition to the current use of grass silage in cattle feeding. However, cell wall properties, including the enzymatic degradability of structural polysaccharides in industrial fermenters or animal rumen, is greatly influenced by the embedding of cell wall carbohydrates in lignin matrix, and the linkages between lignins, p-hydroxycinnamic acids, and arabinoxylans. Breeding for higher and cheaper biofuel or silage production will thus be based on the discovery of genetic traits involved in each cell wall component biosynthesis and deposition in each lignified tissue. Due to its considerable genetic and genomic backgrounds, maize is the relevant model species for identifying traits underlying cell wall degradability variations in grasses. Maize genes involved or putatively involved in the biosynthesis of cell wall phenolic compounds, cell wall carbohydrates and regulation factors were therefore searched for using data available in grass, Arabidopsis, and woody species (mostly poplar and eucalyptus). All maize ortholog genes were searched for using protein sequences and a “blastp” strategy against data available in the www.maizesequence.org database. Genes were also mapped in silico considering their physical position in the same database. Finally, 409 candidate genes putatively involved in secondary cell wall biosynthesis and assembly were shown in the maize genome, out of which 130 were related to phenolic compound biosynthesis, 81 were related to cell wall carbohydrate biosynthesis, and 198 were involved in more or less known regulation mechanisms. Most probable candidate genes involved in regulation and assembly of secondary cell wall belonged to the MYB (45 genes) and NAC (38 genes) families, but also included zinc finger and HDZipIII encoding genes. While genes involved in ferulic acid cross-linkages with other cell wall components were little known, several families putatively involved in (arabino)-xylan chain biosynthesis and in feruloyl transfer were shown, including especially arabinosyl-CoA-acyltransferases, feruloyl-AX b-1,2-xylosyl transferases, and xylan-O-3-arabinosyl transferases. This candidate gene list, which focused on genes and orthologs known to be involved in cell wall component biosynthesis and regulation, cannot be considered as exhaustive. Other genes, whose role in cell wall lignification and deposition have not yet been defined, should very likely be added to the list of candidates required for secondary cell wall assembly. Genes encoding proteins of still unknown function should also be added to the list, as several of the latter are probably involved in lignified tissue biosynthesis and deposition

A new genomic resource dedicated to wood formation in Eucalyptus

<p>Abstract</p> <p>Background</p> <p>Renowned for their fast growth, valuable wood properties and wide adaptability, <it>Eucalyptus </it>species are amongst the most planted hardwoods in the world, yet they are still at the early stages of domestication because conventional breeding is slow and costly. Thus, there is huge potential for marker-assisted breeding programs to improve traits such as wood properties. To this end, the sequencing, analysis and annotation of a large collection of expressed sequences tags (ESTs) from genes involved in wood formation in <it>Eucalyptus </it>would provide a valuable resource.</p> <p>Results</p> <p>We report here the normalization and sequencing of a cDNA library from developing <it>Eucalyptus </it>secondary xylem, as well as the construction and sequencing of two subtractive libraries (juvenile <it>versus </it>mature wood and <it>vice versa</it>). A total of 9,222 high quality sequences were collected from about 10,000 cDNA clones. The EST assembly generated a set of 3,857 wood-related unigenes including 2,461 contigs (Cg) and 1,396 singletons (Sg) that we named 'EUCAWOOD'. About 65% of the EUCAWOOD sequences produced matches with poplar, grapevine, <it>Arabidopsis </it>and rice protein sequence databases. BlastX searches of the Uniref100 protein database allowed us to allocate gene ontology (GO) and protein family terms to the EUCAWOOD unigenes. This annotation of the EUCAWOOD set revealed key functional categories involved in xylogenesis. For instance, 422 sequences matched various gene families involved in biosynthesis and assembly of primary and secondary cell walls. Interestingly, 141 sequences were annotated as transcription factors, some of them being orthologs of regulators known to be involved in xylogenesis. The EUCAWOOD dataset was also mined for genomic simple sequence repeat markers, yielding a total of 639 putative microsatellites. Finally, a publicly accessible database was created, supporting multiple queries on the EUCAWOOD dataset.</p> <p>Conclusion</p> <p>In this work, we have identified a large set of wood-related <it>Eucalyptus </it>unigenes called EUCAWOOD, thus creating a valuable resource for functional genomics studies of wood formation and molecular breeding in this economically important genus. This set of publicly available annotated sequences will be instrumental for candidate gene approaches, custom array development and marker-assisted selection programs aimed at improving and modulating wood properties.</p

Comparative expression of cell wall related genes in four maize RILs and one parental line of variable lignin content and cell wall degradability

A comparison of gene expression in maize between the parental line F271 and four RILs derived from the cross F288 x F271 was investigated based on hybridization on the 17,555 probes Affymetrix micro-array, targeting nearly one third of the genes present in maize genomes. The parental line had unfavorable alleles for cell wall degradability traits at the major QTL position in bin 6.06, while the set of RILs had both the favorable allele and high cell wall degradability. 360 genes were differentially expressed in the four RIL in comparison to F271, including nine genes underlying the major QTL position and 36 underlying two other QTL positions. However, their proposed function (whenever is described) do not allow us to firmly consider their involvement in the observed variation of cell wall related traits. Only a few genes involved in monolignol biosynthesis or polymerization located elsewhere in the genome were differentially expressed between the four RILs and F271, corroborating with the fact that these genes are probably not involved in major determinants of cell wall degradability in the studied set of lines. Among the investigated regulation factors, three ZmMYB, one NAC and one C3HC4 zinc finger were differentially expressed between the four RILs and F271, but they were not located in bin 6.06. Notwithstanding, the obtained results especially strengthened the probable involvement of these genes in maize secondary wall assembly and/ or lignification

Structural, evolutionary and functional analysis of the NAC domain protein family in Eucalyptus

NAC domain transcription factors regulate many developmental processes and stress responses in plants and vary widely in number and family structure. We analysed the characteristics and evolution of the NAC gene family of Eucalyptus grandis, a fastgrowing forest tree in the rosid order Myrtales. NAC domain genes identified in the E. grandis genome were subjected to amino acid sequence, phylogenetic and motif analyses. Transcript abundance in developing tissues and abiotic stress conditions in E. grandis and E. globulus was quantified using RNA-seq and RT-qPCR.189 E. grandis NAC (EgrNAC) proteins, arranged into 22 subfamilies, are extensively duplicated in subfamilies associated with stress response. Most EgrNAC genes form tandem duplicate arrays that frequently carry signatures of purifying selection. Sixteen amino acid motifs were identified in EgrNAC proteins, eight of which are enriched in, or unique to, Eucalyptus. New candidates for the regulation of normal and tension wood development and cold responses were identified.This first description of a Myrtales NAC domain family reveals a unique history of tandem duplication in stress-related subfamilies that has likely contributed to the adaptation of eucalypts to the challenging Australian environment. Several new candidates for the regulation of stress, wood formation and tree-specific development are reported.ANR (Project “Tree For Joules” ANR-2010-KBBE-007-01; Labex Tulip ANR-10-LABX-41), the CNRS, and the University Toulouse III (UPS). Bioinformatics and Functional Genomics Programme of the National Research Foundation, South Africa (UID 71255).http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1469-81372016-06-30hb201