Characterization of NF-Y heterotrimeric transcription factor complexes involved in the symbiotic interaction between M. truncatula and S. meliloti.

MAËL BAUDIN, TOM LALOUM, AGNÈS LEPAGE1, FERNANDA DE CARVALHO-NIEBEL1, CAROLINA RIPODAS2, FLAVIO BLANCO2, MARIA EUGENIA ZANETTI, SENJUTI SINHAROY, MICHAEL UDVARDI AND ANDREAS NIEBEL1

Tenerife

Congreso; 11th European Nitrogen Fixation Conference; 2014

Characterization of NF-Y heterotrimeric transcription factor complexes involved in the symbiotic interaction between M. truncatula and S. meliloti. Maël Baudin1, Tom Laloum1, Agnès Lepage1, Fernanda de Carvalho-Niebel1, Carolina Ripodas2, Flavio Blanco2, Maria-Eugenia Zanetti2, Senjuti Sinharoy3, Michael Udvardi3 and Andreas Niebel1 1 INRA/CNRS, Laboratoire des Interactions Plantes-Microorganismes (LIPM), UMR441/2594, F-31326 Castanet-Tolosan, (France).2 Instituto de Biotecnología y Biología Molecular, FCE UNLP, calle 115 y 49 s/n, CP 1900-La Plata, Argentina. 3 The Samuel Roberts Noble Foundation, Ardmore, Oklahoma 73401 E-mail: mael.baudin@toulouse.inra.fr The root endosymbiotic association between legumes and soil bacteria known as rhizobia leads to the formation of root nodules in which the differentiated bacteria convert atmospheric nitrogen into a form that can be assimilated by the host plant. Several specialized NF-Y (Nuclear Factor Y) transcription factors (TFs) are described to be involved in this symbiotic interaction (1-3). Among them, MtNF-YA1 was first shown to be a key regulator of nodule development in Medicago truncatula (1) and more recently, also of rhizobial infection (4). The NF-Y proteins are part of a transcriptional complex also called CBF (CCAAT-box Binding Factor) or HAP (Heme Activator Protein) present in all eukaryotic systems (5). This complex is composed by three unrelated proteins (NF-YA, NF-YB and NF-YC) which assemble together to bind DNA at CCAAT-boxes, a motif present in most eukaryotic promoters. Interestingly, the trimeric form of the NF-Y complex is required for DNA binding, the NF-YA subunit can thus not act as a TF alone (5). However, in most plant reports, NF-Y subunits are not studied as a trimeric complex thereby neglecting an important aspect of this regulator. In plants, NF-Y proteins are encoded by small multigene families with sometimes specialized expression patterns and functions, and show a certain degree of interaction specificity. In this study, we characterized the NF-YB and NF-YC proteins interacting with MtNF-YA1 during the rhizobium?legume symbiosis. For this purpose, we combined blind protein/protein interaction approaches in yeast and analysis of transcriptomic data to identify the most relevant symbiotic NF-Y complex. Two by two interactions and trimeric complex formation were confirmed using yeast-two and -three hybrid, BiFC and Co-IP experiments. In addition functional studies using RNAi approaches revealed partial redundancy among certain NF-Y subunits and similar functions to MtNF-YA1 for others. The identification and characterization of trimeric NF-Y complexes around MtNF-YA1 reveal an additional layer of regulation of this important symbiotic TF. Bibliography 1. Combier JP, et al. (2006) MtHAP2-1 is a key transcriptional regulator of symbiotic nodule development regulated by microRNA169 in Medicago truncatula. Genes & Development 20(22):3084-3088. 2. Soyano T, Kouchi H, Hirota A, & Hayashi M (2013) NODULE INCEPTION Directly Targets NF-Y Subunit Genes to Regulate Essential Processes of Root Nodule Development in Lotus japonicus. Plos Genetics 9(3). 3. Zanetti ME, Blanco FA, Beker MP, Battaglia M, & Aguilar OM (2010) A C Subunit of the Plant Nuclear Factor NF-Y Required for Rhizobial Infection and Nodule Development Affects Partner Selection in the Common Bean-Rhizobium etli Symbiosis. Plant Cell 22(12):4142-4157. 4. Laporte P, et al. (2014) The CCAAT box-binding transcription factor NF-YA1 controls rhizobial infection. J Exp Bot 65(2):481-494. 5. Laloum T, De Mita S, Gamas P, Baudin M, & Niebel A (2013) CCAAT-box binding transcription factors in plants: Y so many? Trends in Plant Science 18(3):157-166.