We report a reduction of lignin content in the woody plant Eucalyptus camaldulensis by the suppression of gene expression of the LIM domain transcription factor. Previously, we identified a cDNA encoding the tobacco (Nicotiana tabacum) LIM domain transcription factor, Nt1iml, involved in lignin biosynthesis and that specifically binds to an important cis-acting element, the PALbox sequence. The orthologous Eucalyptus gene of Ntliml, namely Ecliml, was isolated from the E. camaldulensis cDNA library (84% amino acid identity). The antisense Ntliml construct with a kanamycin-resistant gene was introduced into E. camaldulensis. The transgenic Eucalyptus plants grown in the greenhouse showed decreased expression levels of severallignin biosynthesis genes, phenylalanine ammonialyase (PAL), cinnamate-4-hydroxylase (C4H) and 4-hydroxycinnamate CoA ligase (4CL). The abnormal phenotypic changes and a 29% reduction of lignin content were observed in the line LG12, in which the transcript level of Ecliml was most1y suppressed. Ec1iml is one of the key transcription factors involved in lignin biosynthesis.

Wood basic density (WBD), the biomass of plant cell walls per unit volume, is an important trait for elite tree selection in kraft pulp production. Here, we investigated the correlation between WBD and wood volumes or wood properties using 98 open-pollinated, 2.4 to 2.8 year-old hybrid Eucalyptus (Eucalyptus urophylla x E. grandis). Transcript levels of lignocellulose biosynthesis-related genes were studied. The progeny plants had average WBD of 516 kg/m3 with normal distribution and did not show any correlations between WBD and wood volume or components of α-cellulose, hemicellulose and Klason lignin content. Transcriptomic analysis of two groups of five plants each with high (570–609 kg/m3) or low (378–409 kg/m3) WBD was carried out by RNA-Seq analysis with total RNAs extracted from developing xylem tissues at a breast height. Lignocellulose biosynthesis-related genes, such as cellulose synthase, invertase, cinnamate-4-hydroxylase and cinnamoyl-CoA reductase showed higher transcript levels in the high WBD group. Among plant cell wall modifying genes, increased transcript levels of several expansin and xyloglucan endo-transglycosylase/hydrolase genes were also found in high WBD plants. Interestingly, strong transcript levels of several cytoskeleton genes encoding tubulin, actin and myosin were observed in high WBD plants. Furthermore, we also found elevated transcript levels of genes encoding NAC, MYB, basic helix-loop-helix, homeodomain, WRKY and LIM transcription factors in the high WBD plants. All these results indicate that the high WBD in plants has been associated with the increased transcription of many genes related to lignocellulose formation. Most lignocellulose biosynthesis related genes exhibited a tendency to transcribe at relatively higher level in high WBD plants. These results suggest that lignocellulose biosynthesis-related genes may be associated with WBD.

Novel transgenic Eucalyptus camaldulensis trees expressing the bacterial choline oxidase A (codA) gene by the Cauliflower mosaic virus (CaMV) 35S promoter and the Arabidopsis thaliana heat shock protein (HSP) terminator was developed. To evaluate the codA transcription level and the metabolic products and abiotic stress tolerance of the transgenic trees, a six-month semi-confined screen house cultivation trial was conducted under a moderate-stringency salt-stress condition. The transcription level of the CaMV 35S promoter driven-codA was more than fourfold higher, and the content of glycine betaine, the metabolic product of codA, was twofold higher, with the HSP terminator than with the nopaline synthase (NOS) terminator. Moreover, the screen house cultivation revealed that the growth of transgenic trees under the salt stress condition was alleviated in correlation with the glycine betaine concentration. These results suggest that the enhancement of codA transcription by the HSP terminator increased the abiotic stress tolerance of Eucalyptus plantation trees.