Basic Helix-Loop-Helix Transcription Factors JASMONATE-ASSOCIATED MYC2-LIKE1 (JAM1), JAM2, and JAM3 Are Negative Regulators of Jasmonate Responses in Arabidopsis 1[W][OPEN] Yuko Sasaki-Sekimoto 2 , Yusuke Jikumaru, Takeshi Obayashi, Hikaru Saito, Shinji Masuda, Yuji Kamiya, Hiroyuki Ohta, and Ken Shirasu* RIKEN Plant Science Center, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan (Y.S.-S., Y.J., Y.K., K.S.); Graduate School of Information Sciences, Tohoku University, Aoba-ku, Sendai, Miyagi 980-8579, Japan (T.O.); Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Midori-ku, Yokohama, Kanagawa 226-8501, Japan (H.S.); Center for Biological Resources and Informatics, Tokyo Institute of Technology, Midori-ku, Yokohama, Kanagawa 226-8501, Japan (S.M., H.O.); Earth-Life Science Institute, Tokyo Institute of Technology, Meguro-ku, Tokyo 152-8550, Japan (S.M., H.O.); and RIKEN Center for Sustainable Resource Science, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan (K.S.) ORCID IDs: 0000-0003-1057-1870 (Y.S.-S.); 0000-0003-4415-520X (Y.K.); 0000-0002-0349-3870 (K.S.). Jasmonates regulate transcriptional reprogramming during growth, development, and defense responses. Jasmonoyl-isoleucine, an amino acid conjugate of jasmonic acid (JA), is perceived by the protein complex composed of the F-box protein CORONATINE INSENSITIVE1 (COI1) and JASMONATE ZIM DOMAIN (JAZ) proteins, leading to the ubiquitin-dependent degradation of JAZ proteins. This activates basic helix-loop-helix-type MYC transcription factors to regulate JA-responsive genes. Here, we show that the expression of genes encoding other basic helix-loop-helix transcription factors, JASMONATE ASSOCIATED MYC2-LIKE1 (JAM1), JAM2, and JAM3, is positively regulated in a COI1- and MYC2-dependent manner in Arabidopsis (Arabidopsis thaliana). However, contrary to myc2, the jam1jam2jam3 triple mutant exhibited shorter roots when treated with methyl jasmonate (MJ), indicating enhanced responsiveness to JA. Our genome-wide expression analyses revealed that key jasmonate metabolic genes as well as a set of genes encoding transcription factors that regulate the JA-responsive metabolic genes are negatively regulated by JAMs after MJ treatment. Consistently, loss of JAM genes resulted in higher accumulation of anthocyanin in MJ-treated plants as well as higher accumulation of JA and 12-hydroxyjasmonic acid in wounded plants. These results show that JAMs negatively regulate the JA responses in a manner that is mostly antagonistic to MYC2. Jasmonic acid (JA) and its cyclic precursors and de- rivatives, so called jasmonates, are widely distributed in the plant kingdom. Jasmonates regulate a wide spec- trum of plant processes, such as growth and develop- ment, as well as defense systems against biotic and abiotic stresses (Schilmiller and Howe, 2005; Browse and Howe, 2008; Zhang and Turner, 2008; Browse, 2009; Reinbothe et al., 2009). JA and jasmonoyl-L-isoleucine (JA-Ile), both derived from a-linolenic acid, accumulate in response to biotic and abiotic stress (Fig. 1; Wasternack, 2007; Gfeller et al., 2010). The accumulation of JA-Ile, which is the ligand for the CORONATINE INSENSI- TIVE1 (COI1)-JASMONATE ZIM DOMAIN (JAZ) re- ceptor complex (Chini et al., 2007; Thines et al., 2007; Sheard et al., 2010), is followed by the generation of 12- hydroxyjasmonoyl-L-isoleucine (12-OH-JA-Ile), which has a lesser binding ability to the complex (Kitaoka et al., 2011; Koo et al., 2011; Heitz et al., 2012). 12-Hydrox- yjasmonic acid (12-OH-JA) and 12-hydroxyjasmonic acid glucoside, on the other hand, have been described as inducers of tuber formation (Yoshihara et al., 1989). 12-OH-JA is also synthesized by the jasmonate pathway and has been detected in a wide range of plant species, especially after wounding (Glauser et al., 2008; Miersch et al., 2008). Biochemical pathways of JA-related com- pounds will be referred to as the jasmonate metabolic pathway in this paper (Fig. 1). Increased JA and JA-Ile levels lead to transcriptional reprogramming. Upon the perception of JA-Ile by the COI-JAZ receptor complex, SCF-type E3 ubiquitin li- gase (SCF COI1 ) presumably ubiquitinates and degrades JAZ proteins via the 26S proteasome system (Chini et al., 2007; Thines et al., 2007; Staswick, 2008). This is 1 This work was supported by Grants-in-Aid for Japan Society for the Promotion of Science Fellows (grant nos. 22–40129 and 19–8288 to Y.S.-S,) and The Ministry of Education, Culture, Sports, Science and Technology KAKENHI (grant no. 24228008 to K.S.). 2 Present address: Earth-Life Science Institute, Tokyo Institute of Technology, 2–12–1 Ookayama Meguro-ku, Tokyo 152-8550, Japan. * Address correspondence to ken.shirasu@riken.jp. The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy de- scribed in the Instructions for Authors (www.plantphysiol.org) is: Ken Shirasu (ken.shirasu@riken.jp). [W] The online version of this article contains Web-only data. [OPEN] Articles can be viewed online without a subscription. www.plantphysiol.org/cgi/doi/10.1104/pp.113.220129 Plant Physiology Ò , September 2013, Vol. 163, pp. 291–304, www.plantphysiol.org Ó 2013 American Society of Plant Biologists. All Rights Reserved. 291