title: Effects of coumarate 3-hydroxylase down-regulation on lignin structure.
abstract: Down-regulation of the gene encoding 4-coumarate 3-hydroxylase (C3H) in alfalfa massively but predictably increased the proportion of p-hydroxyphenyl (P) units relative to the normally dominant guaiacyl (G) and syringyl (S) units. Stem levels of up to approximately 65% P (from wild-type levels of approximately 1%) resulting from down-regulation of C3H were measured by traditional degradative analyses as well as two-dimensional 13C-1H correlative NMR methods. Such levels put these transgenics well beyond the P:G:S compositional bounds of normal plants; p-hydroxyphenyl levels are reported to reach a maximum of 30% in gymnosperm severe compression wood zones but are limited to a few percent in dicots. NMR also revealed structural differences in the interunit linkage distribution that characterizes a lignin polymer. Lower levels of key beta-aryl ether units were relatively augmented by higher levels of phenylcoumarans and resinols. The C3H-deficient alfalfa lignins were devoid of beta-1 coupling products, highlighting the significant differences in the reaction course for p-coumaryl alcohol versus the two normally dominant monolignols, coniferyl and sinapyl alcohols. A larger range of dibenzodioxocin structures was evident in conjunction with an approximate doubling of their proportion. The nature of each of the structural units was revealed by long range 13C-1H correlation experiments. For example, although beta-ethers resulted from the coupling of all three monolignols with the growing polymer, phenylcoumarans were formed almost solely from coupling reactions involving p-coumaryl alcohol; they resulted from both coniferyl and sinapyl alcohol in the wild-type plants. Such structural differences form a basis for explaining differences in digestibility and pulping performance of C3H-deficient plants.
authors.affiliation: United States Dairy Forage Research Center, United States Department of Agriculture-Agricultural Research Service, Madison, Wisconsin 53706, USA. jralph@wisc.edu
authors.forename: John
authors.initials: J
authors.lastname: Ralph
authors.forename: Takuya
authors.initials: T
authors.lastname: Akiyama
authors.forename: Hoon
authors.initials: H
authors.lastname: Kim
authors.forename: Fachuang
authors.initials: F
authors.lastname: Lu
authors.forename: Paul F
authors.initials: PF
authors.lastname: Schatz
authors.forename: Jane M
authors.initials: JM
authors.lastname: Marita
authors.forename: Sally A
authors.initials: SA
authors.lastname: Ralph
authors.forename: M S Srinivasa
authors.initials: MS
authors.lastname: Reddy
authors.forename: Fang
authors.initials: F
authors.lastname: Chen
authors.forename: Richard A
authors.initials: RA
authors.lastname: Dixon
mesh_terms: D002473:Cell Wall / Q000737:chemistry; D015536:Down-Regulation; D008031:Lignin / Q000032:analysis* / Q000096:biosynthesis / Q000737:chemistry; D000455:Medicago sativa / Q000737:chemistry* / Q000201:enzymology* / Q000235:genetics; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D015394:Molecular Structure; D014442:Monophenol Monooxygenase; D019906:Nuclear Magnetic Resonance, Biomolecular; D030821:Plants, Genetically Modified / Q000737:chemistry / Q000201:enzymology
chemical_list: D008031:Lignin; D006899:Mixed Function Oxygenases; D014442:Monophenol Monooxygenase