NUCLEAR FACTOR Y Transcription Factors Have Both Opposing and Additive Roles in ABA-Mediated Seed Germination Roderick W. Kumimoto, Chamindika L. Siriwardana, Krystal K. Gayler, Jan R. Risinger, Nicholas Siefers, Ben F. Holt III* Department of Microbiology and Plant Biology, University of Oklahoma, Norman, Oklahoma, United States of America Abstract In the model organism Arabidopsis thaliana the heterotrimeric transcription factor NUCLEAR FACTOR Y (NF-Y) has been shown to play multiple roles in facilitating plant growth and development. Although NF-Y itself represents a multi-protein transcriptional complex, recent studies have shown important interactions with other transcription factors, especially those in the bZIP family. Here we add to the growing evidence that NF-Y and bZIP form common complexes to affect many processes. We carried out transcriptional profiling on nf-yc mutants and through subsequent analyses found an enrichment of bZIP binding sites in the promoter elements of misregulated genes. Using NF-Y as bait, yeast two hybrid assays yielded interactions with bZIP proteins that are known to control ABA signaling. Accordingly, we find that plants mutant for several NF-Y subunits show characteristic phenotypes associated with the disruption of ABA signaling. While previous reports have shown additive roles for NF-YC family members in photoperiodic flowering, we found that they can have opposing roles in ABA signaling. Collectively, these results demonstrated the importance and complexity of NF-Y in the integration of environmental and hormone signals. Citation: Kumimoto RW, Siriwardana CL, Gayler KK, Risinger JR, Siefers N, et al. (2013) NUCLEAR FACTOR Y Transcription Factors Have Both Opposing and Additive Roles in ABA-Mediated Seed Germination. PLoS ONE 8(3): e59481. doi:10.1371/journal.pone.0059481 Editor: Enamul Huq, University of Texas at Austin, United States of America Received September 20, 2012; Accepted February 14, 2013; Published March 19, 2013 Copyright: ß 2013 Kumimoto 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: This material is based upon work supported by the National Science Foundation under Grant No. IOS-0920258. Additional assistance was provided by the Oklahoma Center for the Advancement of Science and Technology under Grant No. PSB10-013. 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: benholt@ou.edu Introduction Successful acclimation of plants to temporal environmental changes requires the integration of multiple intersecting signals. These environmental signals, such as changing light and water availability, profoundly modify growth and developmental pro- grams. Phytohormones often play a central role in these responses and recent studies identified several integrators of light and hormone signaling pathways (reviewed in [1] and [2]). For example, the basic leucine zipper (bZIP) transcription factor ELONGATED HYPOCOTYL 5 (HY5) has been intensely studied for its roles in light regulated development, but was only recently found to additionally mediate abscisic acid (ABA) signaling in germinating seeds [3]. Thus, pathway integrators can act as hubs for multiple environmental inputs, but how they coordinate these variable inputs to generate unique transcriptional outputs remains unknown [2]. One possibility for coordinating multiple environmental inputs is through interactions with combinatorial transcription factors. Increasing evidence suggests that the combinatorial transcription factor NUCLEAR FACTOR Y (NF-Y) plays important roles in facilitating plant responses to various environmental signals and hormones [4–7]. In fact, like HY5, NF-Y complexes can regulate both light signaling (blue light) and hormone perception (ABA, [7]). NF-Y transcription factors bind at CCAAT cis-elements and function as trimeric complexes consisting of three distinct protein subunits, NF-YA (NF-Y, subunit A), NF-YB, and NF-YC [8]. These individual subunits are each encoded by small gene families found throughout the plant lineage – e.g., Arabidopsis thaliana (Arabidopsis) has 36 NF-Y encoding genes (10 NF-YA, 10 NF-YB, and 10 NF-YC [9–11]). Because the mature DNA-binding NF-Y complex is thought to contain only one of each subunit type, hundreds of unique combinations are theoretically possible. Thus, unique combinations of NF-Y transcription factors may provide a flexible system for fine-tuning the integration of environmental signals to transcriptional outputs [9,12,13]. NF-Y are known to regulate a variety of developmental phenotypes and stress responses. For example, NF-YB and NF- YC subunits regulate photoperiod-dependent flowering [5,6,13– 16] and overexpression of both NF-YA and NF-YB proteins can confer drought tolerance in plants [17,18]. In the legumes Phaseolus vulgaris and Medicago truncatula, specific NF-YA and NF-YC subunits are necessary for the development of nitrogen fixing nodules [19–21]. Other studies have implicated NF-Ys in embryogenesis, light perception, unfolded protein responses (UPR), and photosynthesis [7,22–27]. Although NF-Y itself represents a multi-protein complex that can independently integrate multiple signals, recent studies have shown additional interactions with other transcription factor families, especially those in the bZIP family. Yamamoto et al. [4] PLOS ONE | www.plosone.org 1 March 2013 | Volume 8 | Issue 3 | e59481