Understanding of the role of CAD4, CAD5 and CAD6 gene redundancy during the lignification of Arabidopsis xylem.
Independent thesis Advanced level (degree of Master (Two Years)), 40 credits / 60 HE creditsStudent thesis
Lignin is a cell wall polyphenolic polymer constituting the second most abundant bio-polymer on earth. This polymer is mostly accumulated in wood or xylem where it covers the other cell wall polysaccharides. Thus the removal of lignin allows accessing the polysaccharidic cell wall polymers which can then be converted into bio-fuels, textile or paper. Understanding lignin biosynthesis is therefore important to improve the industrial processing of the woody biomass. Lignin derives from the oxidative polymerization of different types of monomers called monolignols including 4-hydroxyphenylpropene alcohols as the most common monomers. Monolignols derive from the amino acid phenylalanine which is converted by a biosynthetic pathway known as the phenylpropanoid pathway. The last step of the multiple enzymatic process use the enzyme cinnamyl alcohol dehydrogenase (CAD) to convert 4-hydroxyphenylpropene aldehydes into 4-hydroxyphenylpropene alcohols. CAD is encoded by a small multigenic family in Arabidopsis thaliana comprising 17 genes.
The aim of this master thesis is to understand the role of cinnamyl alcohol dehydrogenase gene redundancy during xylem lignification. CAD4, CAD5 and CAD6 have been associated in previous studies with xylem lignification and the main aim is to decipher if these genes are redundant or if they exhibit specificity in their expression (level, time and/or localization) and/or protein activity and structure. To do so, a genetic analysis of the single and double T-DNA insertional loss-of-function mutants in each of these genes were studied to compare their morphological characteristics, their biochemical structure (for both lignin quantification and composition) as well as their gene expression levels.
Although minor changes in the lignin quantity and composition were observed for all of the single mutants, double mutants exhibited significant reductions and changes. Gene expression analysis moreover showed that the loss-of-function in any one of the three CADs caused a reduction ranging from 48% to 95% of the expression of the other CADs independently of the gene mutated. CAD4 and CAD5 both catalyzed the reduction of classical 4-hydroxyphenylpropene aldehydes into their corresponding alcohols: CAD5 catalytic activity is more specific to doubly methoxylated 4-hydroxyphenylpropene aldehydes than CAD4. In contrast, CAD6 did not affect the classical monolignols incorporation into lignin, but instead appeared to assist the function of CAD4 and CAD5. A clear synergetic effect of the double mutants suggested that a potential interaction could occur between these CAD proteins. Overall, our analysis showed that these three CAD genes were not redundant, but instead exhibited distinct function during xylem lignin biosynthesis.
Place, publisher, year, edition, pages
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:umu:diva-113772OAI: oai:DiVA.org:umu-113772DiVA: diva2:890009
Pesquet, Edouard, Univ.Lektor
Bako, Laszlo, Lektor