Uncovering cold disruption of the circadian clock in poplar

Dormancy is an adaptive mechanism that allows woody plants to survive at low temperatures during the winter. Disruption of circadian clock genes in winter or under low temperatures, both in long days as in short days, were described in our group few years ago (Ramos et al., 2005). Basic mechanisms of the circadian clock function are similar in herbaceous as well as in woody plants although there are differences in their response to low temperatures (Bieniawska et al., 2008). Woody plants growing in daylight conditions should have a specific transcriptional control above the circadian clock genes, which is responsible of their constitutive transcriptional activation observed under low temperatures conditions. In order to understand this regulatory process, we are analyzing the behavior of a circadian clock gene in poplar. To this aim, we have isolated its promoter region and fused to the luciferase reporter gene. This construct has been transformed into Populus tremula x P. alba 717-1B4 INRA clone. Here we present the characterization of these transgenic lines under different conditions of light and temperature.

CsRAV1: a candidate gene for improving lignocellulosic biomass yield

Fast-growing tree species of Populus spp.,Salix spp. and Eucalyptus spp. are cultivated to produce wood in a short time. Poplars are cultivated with cycles of 15-18 years to obtain saw timber and peeler logs, but when grown as short -rotation coppice(SRC) to produce biomass, planting density increases and rotation is considerably reduced (3-5 years). In this regard, research efforts are focused in the identification of traits and loci that allow the generation of improved SRC biomass-yielding genotypes. Biomass yield is a highly complex trait as it is the combined outcome of many other complex traits, each under separate polygenic control. Among profitable biomass yield-related traits are the amount of sylleptic branching and the length of winter dormancy. In poplar and in a few other Salicaceae species some lateral buds grow out sylleptically, the same season in which they form without the need of an intervening rest period. Sylleptic branching in poplar increases branch number, leaf area and general growth of the tree in its early years, and is a reasonable predictor of coppice yield. On the other hand, the length of winter dormancy determines the extent of the growth period. Our group has characterized the RAV1 gene of Castanea sativa (CsRAV1), encoding a transcription factor of the subfamily RAV (Related to ABI3/VP1). CsRAV1 expression shows a marked seasonal pattern, being higher in autumn and winter both in stems and buds. We generated transgenic lines of the hybrid clone Populus tremulax P. alba INRA 717 1B4 constitutively expressing CsRAV 1. These CsRAV1-expressing poplars develop sylleptic branches only a few weeks after potting. In addition to the sylleptic branching phenotype, these trees show phenological features that could give rise to an extended growth period. We are currently assessing the phenotype and behavior of these transgenic trees in a field trial, and ultimately, we will evaluate the impact on lignocellulosic biomass quality and production.

Characterization of four bifunctional plant IAM/PAM-amidohydrolases capable of contributing to auxin biosynthesis.

Amidases [EC 3.5.1.4] capable of converting indole-3-acetamide (IAM) into the major plant growth hormone indole-3-acetic acid (IAA) are assumed to be involved in auxin de novo biosynthesis. With the emerging amount of genomics data, it was possible to identify over forty proteins with substantial homology to the already characterized amidases from Arabidopsis and tobacco. The observed high conservation of amidase-like proteins throughout the plant kingdom may suggest an important role of theses enzymes in plant development. Here, we report cloning and functional analysis of four, thus far, uncharacterized plant amidases from Oryza sativa, Sorghum bicolor, Medicago truncatula, and Populus trichocarpa. Intriguingly, we were able to demonstrate that the examined amidases are also capable of converting phenyl-2-acetamide (PAM) into phenyl-2-acetic acid (PAA), an auxin endogenous to several plant species including Arabidopsis. Furthermore, we compared the subcellular localization of the enzymes to that of Arabidopsis AMI1, providing further evidence for similar enzymatic functions. Our results point to the presence of a presumably conserved pathway of auxin biosynthesis via IAM, as amidases, both of monocot, and dicot origins, were analyzed.