Nitrogen-driven stem elongation in poplar is linked with wood modification and gene clusters for stress, photosynthesis and cell wall formation

Euring et al. BMC Plant Biology (2014) 14:391 DOI 10.1186/s12870-014-0391-3 RESEARCH ARTICLE Open Access Nitrogen-driven stem elongation in poplar is linked with wood modification and gene clusters for stress, photosynthesis and cell wall formation Dejuan Euring, Hua Bai, Dennis Janz and Andrea Polle* Abstract Background: Nitrogen is an important nutrient, often limiting plant productivity and yield. In poplars, woody crops used as feedstock for renewable resources and bioenergy, nitrogen fertilization accelerates growth of the young, expanding stem internodes. The underlying molecular mechanisms of nitrogen use for extension growth in poplars are not well understood. The aim of this study was to dissect the nitrogen-responsive transcriptional network in the elongation zone of Populus trichocarpa in relation to extension growth and cell wall properties. Results: Transcriptome analyses in the first two internodes of P. trichocarpa stems grown without or with nitrogen fertilization (5 mM NH4NO3) revealed 1037 more than 2-fold differentially expressed genes (DEGs). Co-expression analysis extracted a network containing about one-third of the DEGs with three main complexes of strongly clustered genes. These complexes represented three main processes that were responsive to N-driven growth: Complex 1 integrated growth processes and stress suggesting that genes with established functions in abiotic and biotic stress are also recruited to coordinate growth. Complex 2 was enriched in genes with decreased transcript abundance and functionally annotated as photosynthetic hub. Complex 3 was a hub for secondary cell wall formation connecting well-known transcription factors that control secondary cell walls with genes for the formation of cellulose, hemicelluloses, and lignin. Anatomical and biochemical analysis supported that N-driven growth resulted in early secondary cell wall formation in the elongation zone with thicker cell walls and increased lignin. These alterations contrasted the N influence on the secondary xylem, where thinner cell walls with lower lignin contents than in unfertilized trees were formed. Conclusion: This study uncovered that nitrogen-responsive elongation growth of poplar internodes is linked with abiotic stress, suppression of photosynthetic genes and stimulation of genes for cell wall formation. Anatomical and biochemical analysis supported increased accumulation of cell walls and secondary metabolites in the elongation zone. The finding of a nitrogen-responsive cell wall hub may have wider implications for the improvement of tree nitrogen use efficiency and opens new perspectives on the enhancement of wood composition as a feedstock for biofuels. Keywords: Development, Metaxylem, Nitrogen use, Populus trichocarpa, Stress, Transcriptome, Wood, Xylem Background Woody biomass is a valuable resource for the generation of renewable energy and an important feedstock for fiber, pulp and cellulose production [1-3]. It is formed during the process of secondary growth. The molecular regulation of secondary growth is intensively being studied in poplar and in the model plant Arabidopsis thaliana [4-9]. * Correspondence: Forest Botany and Tree Physiology, Georg-August Universität Göttingen, Büsgenweg 2, 37077 Göttingen, Germany For example, cell differentiation in the vascular cambium is determined by auxin, auxin transporters, and auxinresponsive transcription factors [7,10]. Furthermore, transcriptional regulation involves members of the AUXIN RESPONSE FACTOR (ARF), MYB, NAC, and WRKY gene families [11-14] whose interplay eventually determines the amounts of cellulose, hemicellulose, and lignin produced during secondary cell wall formation [7]. The prerequisite for secondary growth is primary growth and shoot elongation. The molecular regulation of cell division and differentiation have mainly been addressed in © 2014 Euring et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Euring et al. BMC Plant Biology (2014) 14:391 Arabidopsis [15,16]. In the shoot apical meristem the transcription factors WUSCHEL (WUS) [17], CLAVATA (CLV), SHOOT MERISTEMLESS (STM) [18], and KNOX [19] have been identified as key actors in the control of the size of stem cell population and production of new cell files. They are regulated by hormones, like cytokinins, gibberellin and auxin [20]. Gradients of auxin and signaling peptides are important during the early steps of vascular development [7]. During primary growth, proto- and metaxylem elements are formed. Their differentiation is controlled by transcription factors of the VND (VASCULAR-RELATED NAC DOMAIN) family, VND7 and VND6 [21]. VNDs regulate down-stream transcription factors, especially MYB46 which plays a major role for the orchestration of biosynthetic genes for secondary cell wall formation [22-26]. Although primary growth that drives the elongation of the newly formed internodes is as important for wood production as secondary growth, very little is known about the molecular regulation underlying these developmental processes in poplars. With regard to yield improvement, molecular links between primary growth and nitrogen (N) are of particular interest. Low N frequently limits productivity and consequently, fertilization can enhance yield [27]. Increased N availability results in enhanced leaf area production, increased photosynthesis and higher stem biomass production in poplars [28,29]. However, the wood of fertilized poplars is often characterized by thinner cell walls, less lignification, and increased amounts of tension wood [30-35]. In the developing xylem, key transcription factors for wood formation such as WKRY and NAC domain factors were decreased in hybrid poplars exposed to high (7.5 mM NH4NO3) compared with those grown with adequate N supply (0.75 mM NH4NO3, [36]). Furthermore, the expression levels of several genes involved in hemicellulose and lignin biosynthesis were also reduced, while cellulose synthase increased under high compared with adequate N [36]. The observed transcriptional changes matched alterations in cell wall properties, for example the shift to lower lignin and higher cellulose concentrations in the wood of fertilized compared with nonfertilized poplars [36]. In contrast to radial growth, the influence of N on gene regulation during stem elongation has not been investigated. It is unknown whether high N mainly accelerates primary growth processes such as extension or whether it also impacts on cell wall properties. Understanding the molecular mec (...truncated)