Genetic modification of the flavonoid pathway alters growth and reveals flexible responses to enhanced UVB - Role of foliar condensed tannins.

Accumulation of certain phenolics is a well-known response of plants to enhanced UVB radiation (280-315 nm), but few experiments have compared the relative importance of different phenolic groups for UVB resilience. To study how an altered phenolic profile affects the responses and resilience of silver birch (Betula pendula) to enhanced UVB, we used RNA interference (RNAi) targeting dihydroflavonol reductase (DFR), anthocyanidin synthase (ANS), or anthocyanidin reductase (ANR) to change the accumulation of phenolics. The unmodified control line and RNAi-modified plants were grown for 51 days under ambient or +32% enhanced UVB dose in a greenhouse. RNAi greatly affected phenolic profile and plant growth. There were no interactive effects of RNAi and UVB on growth or photosynthesis, which indicates that the RNAi and unmodified control plants were equally resilient. UVB enhancement led to an accumulation of foliar flavonoids and condensed tannins, and an increase in the density of stem glands and glandular trichomes on upper leaf surfaces in both the control and RNAi-modified plants. Our results do not indicate a photoprotective role for condensed tannins. However, decreased growth of high-flavonoid low-tannin DFRi and ANRi plants implies that the balance of flavonoids and condensed tannins might be important for normal plant growth.

Decreased anthocyanidin reductase expression strongly decreases silver birch (Betula pendula) growth and alters accumulation of phenolics.

Phenolics, formed via a complex phenylpropanoid pathway, are important defensive agents in plants and are strongly affected by nitrogen (N) fertilization. Proanthocyanidins (PAs) are one possible endpoint of the phenylpropanoid pathway, and anthocyanidin reductase (ANR) represents a key enzyme in PA biosynthesis. In this study, the expression of silver birch (Betula pendula) anthocyanidin reductase BpANR was inhibited using the RNA interference (RNAi) method, in three consequent BpANR RNAi (ANRi birches) lines. The growth, the metabolites of the phenylpropanoid pathway, and the number of resin glands of the ANRi birches were studied when grown at two N levels. ANRi birches showed decreased growth and reduction in PA content, while the accumulation of total phenolics in both stems and leaves increased. Moreover, ANRi birches produced more resin glands than did wild-type (WT) birches. The response of ANRi birches to N depletion varied compared with that of WT birches, and in particular, the concentrations of some phenolics in stems increased in WT birches and decreased in ANRi birches. Because the inhibition of PAs biosynthesis via ANR seriously affected birch growth and resulted in accumulation of the precursors, the native level of PAs in plant tissues is assumed to be the prerequisite for normal plant growth. This draws attention to the real plant developmental importance of PAs in plant tissues.

Effects of overproduction of condensed tannins and elevated temperature on chemical and ecological traits of genetically modified hybrid aspens (Populus tremula × P. tremuloides).

Gene transfer techniques offer new possibilities to study regulation of phenolic pathways and the defensive role of phenolics. Hybrid aspen lines (Populus tremula × tremuloides) that overexpress the PtMYB134 transcription factor were used to study the effects of condensed tannin production on plant physiology and plant defenses. The MYB134 protein activates all the known genes of the biosynthetic pathway for condensed tannins (CTs), so overexpression of MYB134 was expected to increase CT concentration in all tissues of the plants. Two out of three MYB134 overexpression lines (46 and 54) accumulated high levels of CTs and (+)-catechin, with a concomitant decrease in the levels of salicylates, but one transgenic line, MYB 61, failed to overproduce CTs. The concentrations of phenolic compounds generally were lower in the aspen leaves grown under elevated temperature than in those grown under ambient temperature. A specialist leaf beetle, Phratora vitellinae (Coleoptera: Chrysomelidae), was chosen to examine how over-expression of MYB134 and elevated temperature affect the food choice of a beetle adapted to feed on leaves rich in salicylates but containing little CT. Specialist beetles preferred the leaves grown at ambient temperatures possibly because these leaves had higher concentrations of salicylates, which are feeding stimulants. Beetles also preferred MYB line 61, which contained a normal level of CT but a slightly elevated level of salicylates. Our results show that transgenic plants are powerful tools, but that enhancing one secondary pathway may lead to unexpected effects on other pathways, and thus impact characteristics such as plant resistance against herbivores, especially under changing climatic conditions.

Identification of novel IL-4/Stat6-regulated genes in T lymphocytes.

IL-4, primarily produced by T cells, mast cells, and basophiles, is a cytokine which has pleiotropic effects on the immune system. IL-4 induces T cells to differentiate to Th2 cells and activated B lymphocytes to proliferate and to synthesize IgE and IgG1. IL-4 is particularly important for the development and perpetuation of asthma and allergy. Stat6 is the protein activated by signal transduction through the IL-4R, and studies with knockout mice demonstrate that Stat6 is critical for a number of IL-4-mediated functions including Th2 development and production of IgE. In the present study, novel IL-4- and Stat6-regulated genes were discovered by using Stat6(-/-) mice and Affymetrix oligonucleotide arrays. Genes regulated by IL-4 were identified by comparing the gene expression profile of the wild-type T cells induced to polarize to the Th2 direction (CD3/CD28 activation + IL-4) to gene expression profile of the cells induced to proliferate (CD3/CD28 activation alone). Stat6-regulated genes were identified by comparing the cells isolated from the wild-type and Stat6(-/-) mice; in this experiment the cells were induced to differentiate to the Th2 direction (CD3/CD28 activation + IL-4). Our study demonstrates that a number a novel genes are regulated by IL-4 through Stat6-dependent and -independent pathways. Moreover, elucidation of kinetics of gene expression at early stages of cell differentiation reveals several genes regulated rapidly during the process, suggesting their importance for the differentiation process.