Arabidopsis CBP1 Is a Novel Regulator of Transcription Initiation in Central Cell-Mediated Pollen Tube Guidance OPEN Hong-Ju Li, a,1 Shan-Shan Zhu, a,1 Meng-Xia Zhang, a,b,1 Tong Wang, a,b Liang Liang, a,b Yong Xue, a,b Dong-Qiao Shi, a Jie Liu, a and Wei-Cai Yang a,2 a State Key Laboratory of Molecular and Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China b University of Chinese Academy of Sciences, Beijing 100049, China ORCID IDs: 0000-0001-9798-9115 (L.L.); 0000-0003-0252-7079 (W.-C.Y.) In flowering plants, sperm cells are delivered to the embryo sac by a pollen tube guided by female signals. Both the gametic and synergid cells contribute to pollen tube attraction. Synergids secrete peptide signals that lure the tube, while the role of the gametic cells is unknown. Previously, we showed that CENTRAL CELL GUIDANCE (CCG) is essential for pollen tube attraction in Arabidopsis thaliana, but the molecular mechanism is unclear. Here, we identified CCG BINDING PROTEIN1 (CBP1) and demonstrated that it interacts with CCG, Mediator subunits, RNA polymerase II (Pol II), and central cell-specific AGAMOUS-like transcription factors. In addition, CCG interacts with TATA-box Binding Protein 1 and Pol II as a TFIIB-like transcription factor. CBP1-knockdown ovules are defective in pollen tube attraction. Expression profiling revealed that cysteine-rich peptide (CRP) transcripts were downregulated in ccg ovules. CCG and CBP1 coregulate a subset of CRPs in the central cell and the synergids, including the attractant LURE1. CBP1 is extensively expressed in multiple vegetative tissues and specifically in the central cell in reproductive growth. We propose that CBP1, via interaction with CCG and the Mediator complex, connects transcription factors and the Pol II machinery to regulate pollen tube attraction. INTRODUCTION Chemotactic guidance is a common mechanism for male-female recognition. Flowering plants have evolved unique multicellular gametophytes and a double fertilization mechanism. The female gametophyte (embryo sac) is composed of seven cells, of which the egg and the central cell are fertilized by two sperm cells from the same male gametophyte (pollen grain) to form the embryo and endosperm of a seed, respectively. The pollen grain consists of two sperm cells and a large vegetative cell, which germinates a tubular structure—the pollen tube—to deliver the immobile sperms to the embryo sac, a phenomenon called siphonogamy. The pollen tube grows to the embryo sac in a polar fashion and is guided by signals from the female, a process called pollen tube guidance, that requires intimate interaction between the embryo sac and the tube. Pollen tube guidance is regulated by the spo- rophytic tissues when growing in the style and transmitting tract, then by the female gametophyte when approaching the ovule, and finally it enters the embryo sac through the micropylar opening (Palanivelu et al., 2003; Palanivelu and Tsukamoto, 2012). During the gametophytic guidance process, two short-range signals are defined: the funicular and the micropylar signals (Higashiyama et al., 2003). The funicular signal attracts the pollen tube from the surface of the septum to the ovule, and the micropylar signal guides the pollen tube from the funicular surface to the receptive synergid cells at the micropyle of the embryo sac (Shimizu and Okada, 2000; Takeuchi and Higashiyama, 2011). The embryo sac has been shown to actively regulate the mi- cropylar pollen tube guidance process by laser ablation and genetic studies in Torenia fournieri and Arabidopsis thaliana (Shimizu and Okada, 2000; Higashiyama et al., 2001; Shimizu et al., 2008). Mutation of the synergid-specific transcription factor MYB98 abolishes micropylar guidance (Kasahara et al., 2005). The LURE family is defensin-like polypeptides derived from the syn- ergids and has been shown to be the chemotactic guidance signals for the pollen tube (Okuda et al., 2009; Takeuchi and Higashiyama, 2012). In maize (Zea mays), the small diffusible peptide EGG APPARATUS1 specifically expressed in the egg apparatus functions as the attractant (Márton et al., 2005, 2012). GEX3, an egg-expressed gene, is also required for pollen tube guidance (Alandete-Saez et al., 2008). magatama3 (maa3), a mutant defective in central cell maturation, exhibits pollen tube attraction defect as well (Shimizu et al., 2008). In the central cell guidance (ccg) mutant, embryo sac development is normal, but micropylar pollen tube guidance is abolished (Chen et al., 2007). CCG is expressed specifically in the central cell, and the central cell-specific expression of CCG driven by the FERTILIZATION- INDEPENDENT SEED2 promoter rescues the guidance defect indicating the indispensable role of the central cell in pollen tube guidance. Immunostaining analysis showed that Arabidopsis LURE1 peptides are not detected in myb98, maa3, and ccg ovules (Takeuchi and Higashiyama, 2012), suggesting that there might be intercellular interactions between the central cell and the syn- ergids to coordinate LURE1 production. CCG encodes a nuclear protein with a conserved N-terminal zinc ribbon domain that is typical for the transcription factor TFIIB family (Knutson, 2013). It 1 These authors contributed equally to this work. 2 Address correspondence to wcyang@genetics.ac.cn. The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantcell.org) is: Wei-Cai Yang (wcyang@ genetics.ac.cn). OPEN Articles can be viewed online without a subscription. www.plantcell.org/cgi/doi/10.1105/tpc.15.00370 The Plant Cell, Vol. 27: 2880–2893, October 2015, www.plantcell.org ã 2015 American Society of Plant Biologists. All rights reserved.