Dissection of Symbiosis and Organ Development by Integrated Transcriptome Analysis of Lotus japonicus Mutant and Wild-Type Plants Niels Høgslund 1,2 , Simona Radutoiu 1 , Lene Krusell 1 , Vera Voroshilova 3¤a , Matthew A. Hannah 3¤b , Nicolas Goffard 4¤c , Diego H. Sanchez 3 , Felix Lippold 3 , Thomas Ott 3¤d , Shusei Sato 5 , Satoshi Tabata 5 , Poul Liboriussen 2 , Gitte V. Lohmann 1 , Leif Schauser 2¤e , Georg F. Weiller 4 , Michael K. Udvardi 3,6 , Jens Stougaard 1 * 1 Centre for Carbohydrate Recognition and Signalling, MBI, Aarhus University, Aarhus C, Denmark, 2 Bioinformatics Research Center (BiRC), Aarhus University, Aarhus C, Denmark, 3 Max-Planck-Institute for Molecular Plant Physiology, Potsdam, Germany, 4 ARC Centre of Excellence for Integrative Legume Research, Genomic Interactions Group, Research School of Biological Sciences, Australian National University, Canberra, Australian Capital Territory, Australia, 5 Kazusa DNA Research Institute, Kisarazu, Chiba, Japan, 6 Plant Biology Division, The Samuel Roberts Noble Foundation, Ardmore, Oklahoma, United States of America Abstract Genetic analyses of plant symbiotic mutants has led to the identification of key genes involved in Rhizobium-legume communication as well as in development and function of nitrogen fixing root nodules. However, the impact of these genes in coordinating the transcriptional programs of nodule development has only been studied in limited and isolated studies. Here, we present an integrated genome-wide analysis of transcriptome landscapes in Lotus japonicus wild-type and symbiotic mutant plants. Encompassing five different organs, five stages of the sequentially developed determinate Lotus root nodules, and eight mutants impaired at different stages of the symbiotic interaction, our data set integrates an unprecedented combination of organ- or tissue-specific profiles with mutant transcript profiles. In total, 38 different conditions sampled under the same well-defined growth regimes were included. This comprehensive analysis unravelled new and unexpected patterns of transcriptional regulation during symbiosis and organ development. Contrary to expectations, none of the previously characterized nodulins were among the 37 genes specifically expressed in nodules. Another surprise was the extensive transcriptional response in whole root compared to the susceptible root zone where the cellular response is most pronounced. A large number of transcripts predicted to encode transcriptional regulators, receptors and proteins involved in signal transduction, as well as many genes with unknown function, were found to be regulated during nodule organogenesis and rhizobial infection. Combining wild type and mutant profiles of these transcripts demonstrates the activation of a complex genetic program that delineates symbiotic nitrogen fixation. The complete data set was organized into an indexed expression directory that is accessible from a resource database, and here we present selected examples of biological questions that can be addressed with this comprehensive and powerful gene expression data set. Citation: Høgslund N, Radutoiu S, Krusell L, Voroshilova V, Hannah MA, et al. (2009) Dissection of Symbiosis and Organ Development by Integrated Transcriptome Analysis of Lotus japonicus Mutant and Wild-Type Plants. PLoS ONE 4(8): e6556. doi:10.1371/journal.pone.0006556 Editor: Nicholas James Provart, University of Toronto, Canada Received March 17, 2009; Accepted June 18, 2009; Published August 7, 2009 Copyright: ß 2009 Høgslund et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Danish National Research Foundation (Research Centre CARB), Danish Cancer Society, Danish Agency for Science Technology and Innovation- SJVF 2113-04-0018, Danish Agency for Nature and Univers- FNU-272-05-0023. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: stougaard@mb.au.dk ¤a Current address: All-Russia Research Institute for Agricultural Microbiology, Laboratory of Genetics of Plant-Microbe Interactions, St Petersburg, Russia ¤b Current address: Bayer BioScience N.V., Gent, Belgium ¤c Current address: Institut Louis Malarde ´, Papeete Tahiti, French Polynesia ¤d Current address: Ludwig-Maximilians-University, Department Biologie 1, Institute of Genetics Maria-Ward-Str. 1a, Mu ¨ nchen, Germany ¤e Current address: Interdisciplinary Nanoscience Center, Aarhus University, Ny Munkegade, Aarhus C, Denmark Introduction Legumes constitute the third largest family (Fabaceae) of flowering plants and they are second only to grasses in their economic and nutritional importance. Several legumes, including soybean, common bean and alfalfa, are major crops producing protein and oil for food and feed. A key trait of legumes is the competence for symbiotic nitrogen fixation, which is the result of an intimate relationship with a group of soil living bacteria collectively called rhizobia. Initial signal exchange between the symbiotic partners triggers a plant morphogenetic program leading to the formation of root nodules, inside which bacteria is hosted and which reduce gaseous nitrogen into ammonium. This eliminates the need for nitrogen fertilizer in crop legumes. Not only does the understanding of this mutualistic association hold the key to a better exploitation of a trait important in agriculture, it also provides insights into molecular processes controlling microbe recognition, pathogen defense and plant organogenesis. Providing PLoS ONE | www.plosone.org 1 August 2009 | Volume 4 | Issue 8 | e6556