Supplemental Data. Nakata et al. Plant Cell. (2013). · PDF filemyc3 280 --- arv nng nn snsn...

Supplemental Figure 1. JA-insensitive root growth of Pro35S:MYC2-SRDX plants.coi1-16 mutant (left), Col-0 (middle), and Pro35S:MYC2-SRDX (right) plants grown for 17 days on MS medium supplemented with 50 µM MeJA.

coi1-16 Col-0Pro35S:MYC2

-SRDX

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Supplemental Data. Nakata et al. Plant Cell. (2013). 10.1105/tpc.113.111112

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Supplemental Figure 2. Expression pattern of JAM1.(A) Expression of JA and wound-inducible gene. RNAs were isolated from rosette leaves of four-week-old plants grown on soil before (control) and one hour after wounding and from 11-day-old seedlings one hour after mock or MeJA (50 µM) treatment. Value of mock-treated or control samples were set to 1. Error bars represent SD of results from three biological replicates. Asterisks indicate significant difference from mock-treated or control samples (Welch’s t test, P < 0.05).(B) to (D) Analyses of promoter activity of JAM1 by ProJAM1:GUS reporter gene construct. (B) Two-week-old ProJAM1:GUS plants grown on MS plates were transferred to MS liquid medium supplemented with (right panel) or without (left panel) 50 µM MeJA for one hour. (C) Rosette leaves of five-week-old ProJAM1:GUS plants one hour after wounding (right panel) and with no treatment (left panel). (D) Inflorescence of seven-week-old ProJAM1:GUS plants. Bars indicate 5 mm in (B) and (C), and 1 mm in (D).

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Supplemental Figure 3. Reduced expression of anthocyanin-related genes in JAM1 CRES-T plants.Relative expression of anthocyanin related genes, DFR (A), LDOX (B) and PAP1 (C). RNAs were purified from seven-day-old seedlings grown in liquid MS medium supplemented with (gray bar) or without (black bar) 50 µM MeJA. Error bars indicate SD of results from three biological replicates. Asterisks indicate significant difference from Col-0 in each treatment (Welch’s t test, P < 0.05).

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Supplemental Figure 4. Reduced trichome phenotype of ProJAM1:JAM1-SRDX plants.Rosette leaves of 12-day-old seedlings of Col-0 (left panel) and ProJAM1:JAM1-SRDX (right panel) plants.

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Supplemental Figure 5. Effect of JAM1-SRDX on floral development.(A) Inflorescences of six-week-old Col-0 (left) and Pro35S:JAM1-SRDX (right) plants.(B) Opening flowers of six-week-old Col-0 (left) and Pro35S:JAM1-SRDX (right) plants. Front petals and sepals were removed.(C) and (D) Scanning electron microscopy images of anthers of Col-0 (C) and Pro35S:JAM1-SRDX(D) plants of the same stage flowers as shown in (B).(E) Flowers of five-week-old plants. Front petals and sepals were removed. Col-0 and Pro35S:JAM1-SRDX #5 showed dehisced anthers and pollination of pollen onto stigma (red arrowheads). The anthers of Pro35S:JAM1-SRDX #9 and #35 plants were neither dehisced nor pollinated (blue arrowheads).(F) Quantification of the expression of JAM1. RNAs were extracted from rosette leaves. The expression of JAM1 (including endogenous JAM1) was examined by quantitative RT-PCR. The expression level of JAM1 in Col-0 was set to 1 and relative expression levels are shown. Pro35S:JAM1-SRDX plants with reduced fertility (#9, #35, and #41) showed higher expression levels than normally fertile plants (#5 and #8).(G) Inflorescence of seven-week-old ProMYC2:MYC2-SRDX.

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Supplemental Figure 6. Filament length in ProJAM1:JAM1-SRDX flowers.Filament length of stage 12 flower in wild type (Col-0) and ProJAM1:JAM1-SRDX plants. Error bars indicate SD (n > 20). Asterisks indicate significant difference (Welch’s t test, P < 0.001).

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Supplemental Figure 7. The level of wound-induced JA-Ile production is similar in JAM1 CRES-T and jam1 plants.Leaf extracts from wounded and unwounded rosette leaves of 24-day-old plants were subject to JA-Ile measurement by LC-MS/MS. (A) JA-Ile levels in Col-0 (black bars), Pro35S:JAM1-SRDX (gray bars) and ProJAM1:JAM1-SRDX(white bars) plants.(B) JA-Ile level in Ler (black bars) and jam1 (gray bars) plants.Error bars indicate SD of results from three biological replicates.

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JAM1 1 ----------------MNMSDLGWDD-E--DKSVVSAVLGHLASDFLRANSNSNQN--------------LFLVMGTDDTLNKKLSSLVDWPNSENFSWNYAIFWQQTMSR-SG------bHLH013 1 ----------------MNIGRLVWNE-D--DKAIVASLLGKRALDYLLSNSVSNAN--------------LLMTLGSDENLQNKLSDLVERPNASNFSWNYAIFWQISRSK-AG------bHLH003 1 ------------------MGQKFWENQE--DRAMVESTIGSEACDFFISTASASNTALS-----------KLVSPPSDSNLQQGLRHVV-----EGSDWDYALFWLASNVNSSD------MYC2 1 -------MTDYRL----QPTMNLWTTDD--NASMMEAFMSSSDISTLWPPASTTTTTATTETTPTPAMEIPAQAGFNQETLQQRLQALIEG---THEGWTYAIFWQPSYDFSGA------MYC3 1 -------------MNGTTSSINFLTSDDDASAAAMEAFIGTNHHSSLFPPP----------------PQQPPQPQFNEDTLQQRLQALIES---AGENWTYAIFWQISHDFDSST--GDNMYC4 1 MSPTNVQVTDYHLNQSKTDTTNLWSTDD--DASVMEAFIGGGSDHSSLFPP----------------LPPPPLPQVNEDNLQQRLQALIEG---ANENWTYAVFWQSSHGFAGEDNNNNN

JAM1 81 QQVLGWGDGCCREPNEEEESKVVRSYNFNNMGAEEETWQDMRKRVLQKLHRLFGGSDEDNYALSLEKVTATEIFFLASMYFFFNHG--EGGPGRCYSSGKHVWLSDAVNSESDYCFRSFMbHLH013 81 DLVLCWGDGYCREPKEGEKSEIVRI---LSMGREEETHQTMRKRVLQKLHDLFGGSEEENCALGLDRVTDTEMFLLSSMYFSFPRG--EGGPGKCFASAKPVWLSDVVNSGSDYCVRSFLbHLH003 79 GCVLIWGDGHCRVKKGAS-------------GEDYSQQDEIKRRVLRKLHLSFVGSDEDHRLVKSGALTDLDMFYLASLYFSFRCDTNKYGPAGTYVSGKPLWAADLPSCLSYYRVRSFLMYC2 99 -SVLGWGDGYYKGEEDKANPRRRSS----SPPFSTPADQEYRKKVLRELNSLISGGVAPSDDAVDEEVTDTEWFFLVSMTQSFACG--AGLAGKAFATGNAVWVSGSDQLSGSGCERAKQMYC3 87 TVILGWGDGYYKGEEDKEKKK--N--------NTNTAEQEHRKRVIRELNSLISGGIGVSDESNDEEVTDTEWFFLVSMTQSFVNG--VGLPGESFLNSRVIWLSGSGALTGSGCERAGQMYC4 100 TVLLGWGDGYYKGEEEKSRKKKSN--------PASAAEQEHRKRVIRELNSLISGGVGGGDEAGDEEVTDTEWFFLVSMTQSFVKG--TGLPGQAFSNSDTIWLSGSNALAGSSCERARQ

JAM1 199 AKSAGIRTIVMVPTDAGVLELGSVWSLPENIGLVKSVQALFMRRVTQ----PVM----VT SNTNMTGGIHKLFGQDLSGAHAYPKK------------LEVRRNLDER--------FTPQbHLH013 196 AKSAGIQTVVLVPTDLGVVELGSTSCLPESEDSILSIRSLFTSSLPP----VRAVALPVT VAEKIDDNRTKIFGKDLHNSGFLQHHQHHQQQQQQPPQQQQHRQFREKLTVRKMDDRAPKbHLH003 186 ARSAGFQTVLSVPVNSGVVELGSLRHIPEDKSVIEMVKSVFGGSDFV-----------------QAKEAPKIFGRQLSLGGAKPRSMSIN--------------------------FSP-MYC2 212 GGVFGMHTIACIPSANGVVEVGSTEPIRQSSDLINKVRILFNFDGGAGDL----SGLNWN LDPDQGENDPSMWINDPIGTPGSNEP----------------------------------MYC3 195 GQIYGLKTMVCIATQNGVVELGSSEVISQSSDLMHKVNNLFNFNNGGGNNGVEASSWGFN LNPDQGENDPALWISEPTNTGI--ES--------------------------------P-MYC4 210 GQIYGLQTMVCVATENGVVELGSSEIIHQSSDLVDKVDTFFNFNNGGGEF----GSWAFN LNPDQGENDPGLWISEPNGVDSGLVA--------------------------------A-

JAM1 291 SWEGYNNNKGPTFGYT-PQRDDVKVLENVN--MVVDNNNYKT--------------------------------------QIE----------------FAGS-----------------bHLH013 312 RLDAYPNNGN-RFMFSNPGTNNNTLLSPT----WVQPENYTR-PINVKEVPSTDEF-----KFLPLQQSSQRLLPPAQM-QID----------------FSAA-----------------bHLH003 262 ------------------KTEDDTGFSL---------ESYEV----------------------------------------Q----------------AIGG-----------------MYC2 294 ------GNGAPSS-SSQLFSKS-IQFENGSSSTITENPNLDPTPSPVHSQTQNPKFNNTFSRELNFSTSSSTLVKPRSGEILNFGDEGKRSSGNPDPSSYSGQTQFENKRKR-SMVLNEDMYC3 280 ---ARVNNGNNSNSNSKSDSHQISKLEKNDI---SSVENQNRQ------------SSCLVEKDLTFQGGL-----LKSNET----------------LSFCGNESSKKRTSVSKGSNNDEMYC4 293 ---PVMNNGGN-DSTSNSDSQPISKLCNGSS---VENPNPKVL------------KSCEM---VNFKNGI-----ENG------------------------QEEDSSNKKRSPVSNNEE

JAM1 337 -------SVAASSNPSTNTQQEKSESCTEKRPVSLLAGAGIVSVVDEKRPRKRGRKPANGREEPLNHVEAERQRREKLNQRFYALRSVVPNISKMDKASLLGDAISYIKELQEKVKIMEDbHLH013 387 -------SSRASENNSD------------GEGGGEWADAVGADESGNNRPRKRGRRPANGRAEALNHVEAERQRREKLNQRFYALRSVVPNISKMDKASLLGDAVSYINELHAKLKVMEAbHLH003 282 ------------SNQV---------------YGYEQGKDETLYLTDEQKPRKRGRKPANGREEALNHVEAERQRREKLNQRFYALRAVVPNISKMDKASLLADAITYITDMQKKIRVYETMYC2 405 KVLSFGD---KTAGESDHSDL--------EASVVKEV-------AVEKRPKKRGRKPANGREEPLNHVEAERQRREKLNQRFYALRAVVPNVSKMDKASLLGDAIAYINELKSKVVKTESMYC3 361 GMLSFSTVVRSAANDSDHSDL--------EASVVKE---AIVVEPPEKKPRKRGRKPANGREEPLNHVEAERQRREKLNQRFYSLRAVVPNVSKMDKASLLGDAISYINELKSKLQQAESMYC4 362 GMLSFTSVLPCD---SNHSDL--------EASVAKEAESNRVVVEPEKKPRKRGRKPANGREEPLNHVEAERQRREKLNQRFYSLRAVVPNVSKMDKASLLGDAISYISELKSKLQKAES

JAM1 450 ERVGTDKSLS----------------------ESNTITVEESPEVDIQAMNEEVVVRVISPLDSHPASRIIQAMRNSNVSLMEAKLSLAEDTMFHTFVIKSNNGS-D-PLTKEKLIAAFYbHLH013 488 ERERLGYS--------------------------SNPPISLDSDINVQTSGEDVTVRINCPLESHPASRIFHAFEESKVEVINSNLEVSQDTVLHTFVVKSE------ELTKEKLISALSbHLH003 375 EKQIMKRRES----------------------N-----QITPAEVDYQQRHDDAVVRLSCPLETHPVSKVIQTLRENEVMPHDSNVAITEEGVVHTFTLRPQGGC-TAEQLKDKLLASLMYC2 507 EKLQIKNQLEEVKLELAGRKASASG---GDMSSSCSSIKPVGMEIEVKIIGWDAMIRVESSKRNHPAARLMSALMDLELEVNHASMSVVNDLMIQQATVKMGFRIYTQEQLRASLISKIMYC3 470 DKEEIQKKLDGMSKEGNNGKGCGSRAKERKSSNQDSTASSIEMEIDVKIIGWDVMIRVQCGKKDHPGARFMEALKELDLEVNHASLSVVNDLMIQQATVKMGSQFFNHDQLKVALMTKVGMYC4 471 DKEELQKQIDVMNKEAGNAKSS---VKDRKCLNQ-ESSVLIEMEVDVKIIGWDAMIRIQCSKRNHPGAKFMEALKELDLEVNHASLSVVNDLMIQQATVKMGNQFFTQDQLKVALTEKVG

JAM1 546 PETSSTQPPLPSSSSQVSGDIbHLH013 576 REQTNSVQSRTSSGR------bHLH003 467 Q--------------------MYC2 624 ---------------------MYC3 590 ENY------------------MYC4 587 ECP------------------

Supplemental Figure 8. Alignment of amino acid sequences of JAM1, MYC2 and closely related bHLHs.The amino acid sequences of JAM1, MYC2 and their closely related bHLHs were aligned using the Clustal Omega program (http://www.ebi.ac.uk/Tools/msa/clustalo/) (Sievers et al., 2011). Identical and similar amino acids are shaded black and gray, respectively, using the BoxShade server 3.21 (www.ch.embnet.org/software/BOX_form.html). Red box indicates the acidic region corresponding to activation domain in MYC2 (Fernandez-Calvo et al., 2011). Blue underlined region indicates the bHLHdomain (Heim et al., 2003).

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Supplemental Figure 9. Subcellular localization of JAM1-GFP.Bright field image (left panel) and GFP fluorescence (by fluorescence microscopy; right panel) in root tips of Pro35S:JAM1-GFP and Pro35S:GFP plants. Bars indicate 50 µm.

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Supplemental Figure 10. Enhanced repression activity of JAM1-SRDX.(A) Schematic representation of the constructs used for transient expression assays. The reporter construct consists of the CaMV35S promoter, five repeats of the GAL4 binding sequence (5xGAL4BS), HSP terminator (HSPter) and firefly luciferase (LUC) coding sequence. Effector constructs express JAM1 or JAM1-SRDX fused to GAL4 DNA binding domain (GALDB) under the control of CaMV35S promoter.(B) Transient expression assays of GAL4-JAM1 and GAL4-JAM1-SRDX. Constructs for the assay are shown in Supplemental Figure 10A. The value for GAL4 was set to 1 and relative values are shown. Error bars indicate SD of results from three replicates.

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Supplemental Figure 11. jam1 transposon insertion line and effects of ProJAM1:JAM1-SRDX on Ler.(A) Schematic diagram of the structure of JAM1 and the position of the transposon insertion in jam1(CS170389). Numbers indicate nucleotide positions of insertion from the translational initiation site. Box represents coding region. Met and Stop indicate translational start and termination codons, respectively. Gray bar indicates region used for amplification in quantitative RT-PCR shown in Figure 4A.(B) Seven-week-old inflorescences of Ler (left) and ProJAM1:JAM1-SRDX on Ler (right) plants.(C) Root lengths of ProJAM1:JAM1-SRDX on Ler plants grown on MS medium containing 0 and 50 µM MeJA. Black and gray bars indicate Ler and ProJAM1:JAM1-SRDX, respectively. Values and error bars indicate average of three independent lines (n > 7) and SD. Asterisk indicates significant difference from Ler (Welch’s t test, P < 0.05).(D) Root lengths of 14-day-old seedlings grown on MS medium supplemented with 0, 5 and 50 mM MeJA. Black and gray bars indicate Ler and jam1, respectively. The values of Ler and jam1 were not significantly different in any of the MeJA concentrations examined (n > 12, Welch’s t test, P > 0.05).(E) Inflorescences of seven-week-old wild type (Ler, left) and jam1 plant (right)

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Supplemental Figure 12. RT-PCR analysis of JAZ10 in jam1.Relative expression of JAZ10.1 (A) and JAZ10.4 (B) in seven-day-old seedlings of Ler (black bars) and jam1(gray bars) grown in liquid MS medium treated with 50 µM MeJA. The value for Ler at 0 hour was set to 1. The horizontal axis indicates hours after JA treatment. Error bars represent SD of results from three biological replicates. Asterisk indicates significant difference from Ler (Welch’s t test, P < 0.05).

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Supplemental Figure 13. Complementation analysis of jin1-8 by MYC2 and JAM1.Transgenic plants of jin1-8 ProMYC2:MYC2 and jin1-8 ProMYC2:JAM1 selected on MS medium containing hygromycin were grown for an additional 10 days on MS medium containing 50 µM MeJA. Col-0 and jin1-8 (-) plants were grown on MS medium and then transferred to 50 µM MeJA containing MS medium. Error bars indicate SD (n > 12). Asterisks indicate significant difference from Col-0 (Welch’s t test, P < 0.01)

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Supplemental Figure 14. Activation activity of JAM1-VP16.(A) Schematic representation of the constructs used for transient expression assays. The reporter construct consists of the CaMV35S promoter, five repeats of the GAL4 binding sequence (5xGAL4BS), HSP terminator (HSPter) and firefly luciferase (LUC) coding sequence. Effector constructs express JAM1 or JAM1-VP16 fused to GAL4 DNA binding domain (GAL4DB) under the control of CaMV35S promoter.(B) Transient expression assays of GAL4-JAM1 and GAL4-JAM1-VP16. Constructs for the assay are shown in Supplemental Figure 14A. The value for GAL4 was set to 1 and relative values are shown. Error bars indicate SD of results from four replicates.

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Supplemental Figure 15. Determination of genotype of coi1-30 JAM1-VP16 plants.(A) Schematic diagram of the structure of COI1 gene and JAM1-VP16 transgene and the position of T-DNA tag in coi1-30. Arrows indicate primer positions and direction used for genomic PCR in (B). Boxes represent coding sequence. Met and Stop indicate translational start and termination codons, respectively.(B) Genomic PCR analysis of Col-0, coi1-30 and coi1-30 JAM1-VP16 plants. coi1-30 JAM1-VP16 lines exhibited amplification of JAM1-VP16 transgene (d+e) and T-DNA (b+c) but not COI1 genome (a+b), indicating that the lines are homozygous coi1-30 plants.

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Supplemental Figure 16. Expression of JAM1 in coi1-30 Pro35S:JAM1-VP16 and coi1-30 Pro35S:JAM1 plants.Relative expression of JAM1 in coi1-30 Pro35S:JAM1-VP16 and coi1-30 Pro35S:JAM1 plants. Value of Col-0 was set to 1 and relative values are shown. Error bars indicate SD of results from three replicates.

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Supplemental Figure 17. RT-PCR analysis of JA-inducible genes after 1 h of JA treatment in Col-0, ProMYC2:MYC2-SRDX and ProJAM1:JAM1-SRDX plants.Relative expression levels of JA-inducible genes in 11-day-old seedlings of Col-0, ProMYC2:MYC2-SRDX(MYC2) and ProJAM1:JAM1-SRDX (JAM1) plants 50 µM MeJA-treated (gray bars) or mock-treated (black bars) for one hour. Values of mock-treated Col-0 were set to 1. Error bars represent SD of results from at least three biological replicates. Asterisks indicate significant difference from Col-0 (Welch’s t test, P < 0.05).

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Supplemental Figure 18. JAM1 and MYC2 did not interact in yeast and plants.(A) Yeast two-hybrid analysis of JAM1 and MYC2. GAL4-JAM1 or -JAZ3 and LexA-MYC2 were used as bait and prey, respectively. Empty vectors for the bait or prey plasmids were used as negative control. Interactions between MYC2 and JAZ3 are displayed as the positive control.(B) Constructs used for transient expression assays. Reporter construct consists of CaMV35S promoter with five GAL4 binding sequence (5xGAL4BS), NOS terminator (NOSter) and firefly luciferase (LUC) coding sequence. Effector1 construct expresses JAM1 or JAZ3 fused to GAL4 DNA binding domain (GAL4DB) under the control of CaMV35S promoter. Effector2 construct expresses MYC2-VP16 or MYC2-SRDX under the control of CaMV35S promoter.(C) Transient expression assay for analyses of the interaction of JAM1 and MYC2 in aos leaves. Constructs for the assay are shown in Supplemental Figure 18B. Reporter construct and effector1 construct with or without effector2 construct were co-bombarded into rosette leaves of aos plants and LUC activity was assayed. Value of GAL4-JAM1 without effector2 was set to 1 and relative values are shown. Error bars indicate SD of results from at least three replicates. Asterisk indicates significant difference from GAL4-JAZ3 without effector2 (Welch’s t test, P < 0.01).(D) Interaction of JAM1 and MYC2 in aos leaves was examined after JA treatment. Particle bombardment was performed as described in (C), and then leaves were incubated with (white bars) or without (black bars) 50 µM MeJA for 12 hours. Error bars indicate SD of results from at least three replicates. Asterisk indicates significant difference from GAL4-JAM1 without effector2 (Welch’s t test, P < 0.05)

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Supplemental Figure 19. Electrophoretic mobility shift assay of MYC2 and JAM1.(A) Sequences of oligo DNA probes used for EMSA. cis-elements are underlined.(B) and (C) Binding of MYC2 (B) and JAM1 (C) to IRDye-labeled oligo DNA probes containing three repeats of cis-elements. Non-labeled probes were added in 1000-fold excess for competition analyses. Three repeats of the mutated G box oligo DNA (3xmGbox) was used as mutated competitor.

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His-JAM1 - + + + - + + + - + + +competitor - - + - - - + - - - + -mutated

competitor- - - + - - - + - - - +

G box TG box MBS G box TG box MBSB C

19


-418 TTCACGTTAAAATAGCGCACGTGCCTCGCCACGCGTCTCACCACAAAAAGAAACGCCACACACTGCACGACGATGCACGT-338 GCCTTTTTAACCGACTATATCCGGTGACGTAACCGGGACAACCTGTAGCTGGTTTTTGCGCCAATAGGATATGAGAAAAA-258 AGAATCTTTTTTTTCTTGTTATAGTTATAAATTAATGGTACGATTCACATGTCGCATGCGATTTGTTAATAAAACTAGAA-178 ATTGCGGTGAATTAACTTCATTCCACGTTTTTTCATTTTCTCCCTCAAAAGTCTCTGTTTTTTTTCCTTTTTCCGGCGAA-98 GCTCTATTTAGCTTGATTCCGGCGTTTAACACGCGTTTTAATCGAAACAGACATTTGAGATCGAATTAATTTTGTAGCGG-18 GCTGTGTCTTTATTATAGATG

Supplemental Figure 20. The binding affinity of JAM1 and MYC2 to the G-box and MYC2 target site.(A) Constructs used for transient expression assays. Reporter constructs consist of firefly luciferase (LUC) coding sequence and TATA-box of CaMV35S promoter with or without five repeats of the G-box (5xG-box) or MBS (5xMBS). The effector constructs express VP16, MYC2-VP16 or JAM1-VP16 under the control of CaMV35S promoter.(B) and (C) Analysis of binding affinity of JAM1 and MYC2 to G-box (B) or MBS (C) by transient expression assays. Black and gray bars indicate reporter constructs with or without the repeated cis-regulatory sequences. Values of VP16 of each reporter construct were set to 1 and relative values are shown. Error bars indicate SD of results from four replicates. Asterisks indicate significant difference from the value of VP16 of each reporter construct (Welch’s t test, P < 0.05). This experiment was repeated twice with the same results.(D) DNA sequence of 418 bp upstream region of JAZ3 used for reporter constructs. Translational start site (red font) and putative MYC2-binding sites (underlined) are highlighted.(E) Schematic representation of the constructs used for transient expression assays. Reporter construct consists of JAZ3 promoter, HSP terminator (HSPter) and firefly luciferase (LUC) coding sequence. The effector constructs express VP16, MYC2-VP16 or JAM1-VP16 under the control of CaMV35S promoter.(F) Binding of JAM1 and MYC2 on putative MYC2-binding sequences in JAZ3 promoter by transient expression assays. Black, gray, and white bars indicate values for no promoter (-), JAZ3 promoter (ProJAZ3) and mutated JAZ3 promoter (mProJAZ3) as shown in Figure S20E. Values of VP16 of each reporter construct were set to 1 and relative values are shown. Error bars indicate SD of results from three replicates. Asterisks indicate significant difference from the value of VP16 of each reporter construct (Welch’s t test, P < 0.05). This experiment was repeated twice with the same results.

A

B C

D

E

ReporterLUC

Effector

TATA HSPter

MYC2 VP16Pro35S NOSter

JAM1 VP16Pro35S NOSter

MYC2VP

JAM1VP

LUCTATA HSPter5 x G-box

LUCTATA HSPter5 x MBS

VP16Pro35S NOSterVP16

ReporterLUC

Effector

MYC2

- HSPter

VP16Pro35S NOSter

JAM1 VP16Pro35S NOSter

LUCProJAZ3 HSPterTG

LUCmProJAZ3 HSPter

MYC2VP

JAM1VP

G GTG MBS

mG

F

Pro35S NOSterVP16 VP16

0 2 4 6

VP16

JAM1VP

MYC2VP


*

*

0 5 10

VP16

JAM1VP

MYC2VP


*

0 20 40 60

VP16

JAM1VP

MYC2VP


*

*

20



MYC2SRDX JAM1SRDXb JA-inducedc

AT5G63450 CYP94B1 "cytochrome P450, family 94, subfamily B, polypeptide 1", cytochrome P450, family 94, subfamily B, polypeptide 1 (.1) 0.17 0.32 80.19AT3G22275 unknown protein 0.37 0.33 61.28AT2G30830 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein (.1) 0.04 0.45 34.75AT1G73325 Kunitz family trypsin and protease inhibitor protein (.1) 0.10 0.39 24.52AT2G38240 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein (.1) 0.40 0.08 19.00AT3G48520 CYP94B3 cytochrome P450, family 94, subfamily B, polypeptide 3 (.1) 0.06 0.18 18.64AT4G27140 SESA1, AT2S1 seed storage albumin 1 (.1) 0.12 0.25 18.62AT1G54020 GDSL-like Lipase/Acylhydrolase superfamily protein (.1.2.3) 0.25 0.65 17.50AT5G13220 JAS1, TIFY9, JAZ10, AT5G13220 TIFY DOMAIN PROTEIN 9, JASMONATE-ASSOCIATED 1, jasmonate-zim-domain protein 10 (.1.2.3.4) 0.57 0.37 16.45AT4G16740 ATTPS03 terpene synthase 03 (.1.2) 0.54 0.06 15.72AT5G24420 PGL5 6-phosphogluconolactonase 5 (.1) 0.25 0.33 15.40AT1G43160 RAP2.6, RAP2.06 related to AP2 6 (.1) 0.16 0.34 15.35AT5G19110 Eukaryotic aspartyl protease family protein (.1) 0.30 0.69 15.32AT5G59580 UGT76E1 UDP-glucosyl transferase 76E1 (.1) 0.36 0.07 14.43AT4G15440 CYP74B2, HPL1 hydroperoxide lyase 1 (.1) 0.20 0.34 14.26AT3G25180 CYP82G1 cytochrome P450, family 82, subfamily G, polypeptide 1 (.1.2) 0.33 0.19 13.77AT3G46660 UGT76E12 UDP-glucosyl transferase 76E12 (.1) 0.10 0.11 13.20AT1G44350 ILL6 IAA-leucine resistant (ILR)-like gene 6 (.1) 0.39 0.29 11.46AT1G12610 DDF1, DREB1F, FTL1 DWARF AND DELAYED FLOWERING 1, Integrase-type DNA-binding superfamily protein (.1) 0.38 0.68 11.25AT1G18710 ATMYB47 myb domain protein 47 (.1) 0.70 0.33 10.86AT5G17490 AtRGL3, RGL3 RGA-like protein 3 (.1) 0.64 0.65 10.42AT5G02170 Transmembrane amino acid transporter family protein (.1.2) 0.33 0.48 9.96AT1G65890 AAE12 acyl activating enzyme 12 (.1) 0.51 0.58 9.82AT1G51760 JR3, IAR3 JASMONIC ACID RESPONSIVE 3, IAA-ALANINE RESISTANT 3, peptidase M20/M25/M40 family protein (.1) 0.43 0.71 9.56AT5G65280 GCL1 GCR2-like 1 (.1) 0.37 0.29 9.32AT1G51780 ILL5 IAA-leucine resistant (ILR)-like gene 5 (.1) 0.40 0.63 9.28AT5G47240 ATNUDT8 nudix hydrolase homolog 8 (.1) 0.65 0.36 9.17AT5G05600 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein (.1) 0.53 0.45 9.13AT3G25573 unknown protein 0.14 0.33 8.94AT1G30135 TIFY5A, JAZ8 jasmonate-zim-domain protein 8 (.1) 0.15 0.47 8.94AT4G17470 alpha/beta-Hydrolases superfamily protein (.1.2.3) 0.21 0.13 8.70AT5G24780 ATVSP1, ATVSP, VSP1 vegetative storage protein 1 (.1.2) 0.57 0.55 8.51AT5G07010 ATST2A ARABIDOPSIS THALIANA SULFOTRANSFERASE 2A, sulfotransferase 2A (.1) 0.40 0.35 8.39AT5G38120 4CL8 AMP-dependent synthetase and ligase family protein (.1) 0.53 0.44 8.38AT3G12145 FLOR1, FLR1 FLOR1, Leucine-rich repeat (LRR) family protein (.1) 0.59 0.63 8.19AT3G25760 AOC1, ERD12 early-responsive to dehydration 12, allene oxide cyclase 1 (.1) 0.27 0.48 8.10AT3G51450 Calcium-dependent phosphotriesterase superfamily protein (.1) 0.37 0.65 8.06AT1G70700 TIFY7, JAZ9 JASMONATE-ZIM-DOMAIN PROTEIN 9, TIFY domain/Divergent CCT motif family protein (.1.2) 0.65 0.72 7.96AT4G13410 ATCSLA15 CELLULOSE SYNTHASE LIKE A15, Nucleotide-diphospho-sugar transferases superfamily protein (.1) 0.37 0.36 7.90AT2G40200 bHLH051 basic helix-loop-helix (bHLH) DNA-binding superfamily protein (.1) 0.40 0.37 7.90AT2G33380 AtCLO3, CLO-3, RD20 RESPONSIVE TO DESSICATION 20, Arabidopsis thaliana caleosin 3, Caleosin-related family protein (.1.2) 0.16 0.07 7.90

AT4G15210BAM5, AT-BETA-AMY, BMY1,RAM1, ATBETA-AMY

REDUCED BETA AMYLASE 1, ARABIDOPSIS THALIANA BETA-AMYLASE, beta-amylase 5 (.1.2) 0.11 0.04 7.80

AT5G41120 Esterase/lipase/thioesterase family protein (.1) 0.28 0.56 7.67AT3G17860 TIFY6B, JAI3, JAZ3 JASMONATE-INSENSITIVE 3, jasmonate-zim-domain protein 3 (.1.2.3) 0.67 0.37 7.58AT1G12240 ATBETAFRUCT4, VAC-INV VACUOLAR INVERTASE, Glycosyl hydrolases family 32 protein (.1) 0.41 0.61 7.55AT5G06870 PGIP2, ATPGIP2 ARABIDOPSIS POLYGALACTURONASE INHIBITING PROTEIN 2, polygalacturonase inhibiting protein 2 (.1) 0.53 0.70 7.46AT1G64200 VHA-E3 vacuolar H+-ATPase subunit E isoform 3, vacuolar H+-ATPase subunit E isoform 3 (.1) 0.25 0.60 7.42AT3G55970 ATJRG21 jasmonate-regulated gene 21 (.1) 0.11 0.11 7.40AT1G19640 JMT jasmonic acid carboxyl methyltransferase (.1) 0.19 0.11 7.16AT4G36830 HOS3-1 GNS1/SUR4 membrane protein family (.1) 0.39 0.66 6.57AT3G28220 TRAF-like family protein (.1) 0.09 0.49 6.50AT2G22770 NAI1, bHLH020 basic helix-loop-helix (bHLH) DNA-binding superfamily protein (.1) 0.68 0.59 6.41AT4G10390 Protein kinase superfamily protein (.1) 0.32 0.50 6.13AT1G10585 basic helix-loop-helix (bHLH) DNA-binding superfamily protein (.1) 0.16 0.27 5.98AT2G22760 bHLH019 basic helix-loop-helix (bHLH) DNA-binding superfamily protein (.1) 0.18 0.54 5.86AT2G22330 CYP79B3 "cytochrome P450, family 79, subfamily B, polypeptide 3", cytochrome P450, family 79, subfamily B, polypeptide 3 (.1) 0.52 0.71 5.63AT3G46650 UDP-Glycosyltransferase superfamily protein (.1) 0.31 0.71 5.61AT1G76790 IGMT5 indole glucosinolate O-methyltransferase 5, O-methyltransferase family protein (.1) 0.37 0.46 5.59AT4G19430 unknown protein 0.23 0.21 5.54AT1G20490 AMP-dependent synthetase and ligase family protein (.1) 0.41 0.49 5.53AT5G57550 XTR3, XTH25, EXGT-A5 xyloglucan endotransglycosylase 3, xyloglucan endotransglucosylase/hydrolase 25 (.1) 0.41 0.33 5.44AT2G43520 ATTI2 trypsin inhibitor protein 2 (.1) 0.41 0.15 5.42AT3G22740 HMT3 homocysteine S-methyltransferase 3 (.1) 0.47 0.68 5.25AT4G33370 DEA(D/H)-box RNA helicase family protein (.1) 0.70 0.74 5.24AT2G29450 ATGSTU1, AT103-1A, ATGSTU5 ARABIDOPSIS THALIANA GLUTATHIONE S-TRANSFERASE TAU 1, glutathione S-transferase tau 5 (.1) 0.59 0.67 5.19AT5G42650 CYP74A, AOS, DDE2 DELAYED DEHISCENCE 2, CYTOCHROME P450 74A, allene oxide synthase (.1) 0.66 0.46 5.14AT1G52030 MBP1.2, F-ATMBP, MBP2 myrosinase-binding protein 2 (.1.2) 0.49 0.53 5.11AT4G25410 bHLH126 basic helix-loop-helix (bHLH) DNA-binding superfamily protein (.1) 0.45 0.46 5.11AT3G16690 SWEET16, AtSWEET16 Nodulin MtN3 family protein (.1) 0.55 0.56 5.05AT1G73210 Protein of unknown function (DUF789) (.1), Protein of unknown function (DUF789) (.2) 0.66 0.20 4.97

AT1G16370 6-Oct, ATOCT6 ARABIDOPSIS THALIANA ORGANIC CATION/CARNITINE TRANSPORTER 6, organic cation/carnitine transporter 6 (.1) 0.64 0.49 4.97

AT2G47180 ATGOLS1 galactinol synthase 1 (.1) 0.42 0.74 4.95AT3G50280 HXXXD-type acyl-transferase family protein (.1) 0.39 0.63 4.89

AT5G55120 VTC5VITAMIN C DEFECTIVE 5, galactose-1-phosphate guanylyltransferase (GDP)s;GDP-D-glucose phosphorylases;quercetin4'-O-glucosyltransferases (.1)

0.58 0.70 4.78

AT4G27410 RD26, ANAC072NAC (No Apical Meristem) domain transcriptional regulator superfamily protein (.1), NAC (No Apical Meristem) domaintranscriptional regulator superfamily protein (.2), NAC (No Apical Meristem) domain transcriptional regulator superfamilyprotein (.3)

0.48 0.47 4.75

AT3G16470 JAL35, JR1 JASMONATE RESPONSIVE 1, jacalin-related lectin 35, Mannose-binding lectin superfamily protein (.1.2.3) 0.51 0.52 4.73AT4G27170 SESA4, AT2S4 seed storage albumin 4 (.1) 0.41 0.55 4.59AT1G69370 CM-3, CM3 chorismate mutase 3 (.1) 0.35 0.45 4.58AT5G62360 Plant invertase/pectin methylesterase inhibitor superfamily protein (.1) 0.50 0.21 4.58AT4G22620 SAUR-like auxin-responsive protein family (.1) 0.50 0.65 4.58

AT1G56650 ATMYB75, SIAA1, PAP1SUC-INDUCED ANTHOCYANIN ACCUMULATION 1, MYELOBLASTOSIS PROTEIN 75, MYB DOMAIN PROTEIN 75,production of anthocyanin pigment 1 (.1)

0.49 0.15 4.49

AT3G25770 AOC2, AOC3 allene oxide cyclase 2 (.1) 0.65 0.34 4.46AT1G16380 ATCHX1 CATION EXCHANGER 1, Cation/hydrogen exchanger family protein (.1) 0.72 0.47 4.28AT5G10625 unknown protein 0.43 0.66 4.04AT5G56490 D-arabinono-1,4-lactone oxidase family protein (.1) 0.38 0.17 4.03AT1G52890 ANAC019 NAC domain containing protein 19 (.1) 0.62 0.13 4.02AT4G24010 CSLG2, ATCSLG1 ARABIDOPSIS THALIANA CELLULOSE SYNTHASE LIKE G1, cellulose synthase like G1 (.1) 0.36 0.54 4.01AT4G23600 JR2, CORI3, TAT1 JASMONIC ACID RESPONSIVE 2, CORONATINE INDUCED 1, Tyrosine transaminase family protein (.1.2.3) 0.35 0.57 3.98AT1G17750 AtPEPR2 PEP1 receptor 2 (.1) 0.57 0.63 3.86AT1G09080 BIP3 binding protein 3, Heat shock protein 70 (Hsp 70) family protein (.1), Heat shock protein 70 (Hsp 70) family protein (.2) 0.20 0.66 3.79

AT2G37180 PIP2C, PIP2;3, RD28RESPONSIVE TO DESICCATION 28, PLASMA MEMBRANE INTRINSIC PROTEIN 2C, PLASMA MEMBRANEINTRINSIC PROTEIN 2;3, Aquaporin-like superfamily protein (.1)

0.47 0.53 3.72

Supplemental Table 1. List of early JA responsive genes down-regulated by both JAM1-SRDX and MYC2-SRDX.

Locus Common Name DescriptionExpression ratio

21


AT1G60260 BGLU5 beta glucosidase 5 (.1) 0.58 0.62 3.64AT1G58200 MSL3 MSCS-like 3 (.1.2) 0.46 0.71 3.63AT2G02498 unknown protein 0.56 0.54 3.59AT5G02940 Protein of unknown function (DUF1012) (.1) 0.36 0.48 3.54AT1G03940 HXXXD-type acyl-transferase family protein (.1) 0.37 0.27 3.49AT4G27560 UDP-Glycosyltransferase superfamily protein (.1) 0.51 0.66 3.47AT1G54010 GDSL-like Lipase/Acylhydrolase superfamily protein (.1) 0.58 0.56 3.45

AT1G29330 ATERD2, AERD2, ERD2ARABIDOPSIS THALIANA ENDOPLASMIC RETICULUM RETENTION DEFECTIVE 2, ARABIDOPSIS ENDOPLASMICRETICULUM RETENTION DEFECTIVE 2, ER lumen protein retaining receptor family protein (.1)

0.58 0.69 3.45

AT5G15240 Transmembrane amino acid transporter family protein (.1.2) 0.66 0.68 3.32AT3G46670 UGT76E11 UDP-glucosyl transferase 76E11 (.1) 0.56 0.45 3.20AT2G19780 Leucine-rich repeat (LRR) family protein (.1) 0.65 0.52 3.17

AT1G066402-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein (.1), 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein (.2), 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily protein

0.48 0.44 3.09

AT1G02620 Ras-related small GTP-binding family protein (.1) 0.76 0.54 3.08AT1G72120 Major facilitator superfamily protein (.1) 0.64 0.52 3.07AT1G24380 unknown protein 0.78 0.70 3.07AT4G21910 MATE efflux family protein (.1.2.3.4) 0.49 0.55 3.06AT2G46370 FIN219, JAR1 JASMONATE RESISTANT 1, FAR-RED INSENSITIVE 219, Auxin-responsive GH3 family protein (.1.2.3.4) 0.54 0.55 3.05AT1G28050 COL15 B-box type zinc finger protein with CCT domain (.1) 0.62 0.46 3.02

a JA-treated MYC2-SRDX plants vs JA-treated control plants (ratio < 0.75 fold, FDR < 0.03)b JA-treated JAM1-SRDX plants vs JA-treated control plants (ratio < 0.75 fold, FDR < 0.03)c JA-treated control plants vs Mock-treated control plants (ratio > 3.0 fold, FDR < 0.03)

22


Purpose Gene primer sequence (5' to 3')JAZ3 GGGGACAAGTTTGTACAAAAAAGCAGGCTCCATGGAGAGAGATTTTCTCGG

GGGGACCACTTTGTACAAGAAAGCTGGGTCGGTTGCAGAGCTGAGAGAAGJAM1 GGGGACAAGTTTGTACAAAAAAGCAGGCTCCATGAATATGAGTGATTTAGGTTGGGATG

GGGGACCACTTTGTACAAGAAAGCTGGGTCTATATCACCAGAGACCTGTGAACTAGAAGMYC2 GGGGACAAGTTTGTACAAAAAAGCAGGCTCCATGACTGATTACCGGCTACA

GGGGACCACTTTGTACAAGAAAGCTGGGTCACCGATTTTTGAAATCAAACTTGCTCJAM1VP16 GGGGACAAGTTTGTACAAAAAAGCAGGCTCCATGAATATGAGTGATTTAGGTTGGGATG

GGGGACCACTTTGTACAAGAAAGCTGGGTCCTACCCACCGTACTCGTCAA5' end of JAM1 promoter AAAGCCACGTGGCGCATGATGTA3' end of JAM1 promoter ATCTTTCTTTATCTCCCAGATTACACC

JAM1 CTAACAACGGGTCGGATCCACACCAGAGACCTGTGAACTAGA

MYC2 CGCGAGTATGTCGGTGGTTATGCTCTGAGCTGTTCTTGCGTATA

VSP1 GTCAGAAGTCACTGTCGAGAATCACTTCGATCCGTTTGGCTTGAGTA

JAZ3.1 GTTCTACCAATGTAATGGCTCCAACACAATATGGGGATACGCTCGT

JAZ3.4 GTTCTACCAATGTAATGGCTCCAACAGGTGTATTCTAAAGAAGTGTTGCGTAC

JAZ10.1 AGCTCTTTGGCCAGAATCTAGAAGATGTTGATACTAATCTCTCCTTG

JAZ10.4 ATGGAGACAGATCTTTCAGATCTGTTTAGGCCGATGTCGGATAGT

DFR TGACAGGATGGATGTATTTCGACCGACCACCAATGTT

LDOX ACTGGAAAGATTCAAGGCTAGTACTCACTCGTTGCTTCTATG

PAP1 GAGGAAAGCCAAGAGGTACAAACCTATACACAAACGCA

MYB21 AAGTAGTGGAGGTTCGGGATCACCGTGGTTGGCGATATAGTTGA

TAT1 GACGGTGTCTTCAGAAACACCAATCAGGAATAGCCGCCTGGATA

VP16 CCAGGGCCGGGATTTACCCCTACCCACCGTACTCGTCAATTC

LOX3 CTCCCTGCCGATCTAATTCGGGCGTACGGATAGTCTTCGA

AOS GGTGGCGAGGTTGTTTGTGAGCGACGTACCAACCTCAATATCA

OPR3 ACGTGCTTCTCATGCAGTGTATCGGGAACCATCGGGCAACAAA

PP2AA3 GACCAAGTGAACCAGGTTATTGGTACTCTCCAGTGCCTGTCTTCA

COI1 (a) ATGGAGGATCCTGATATCAACOI1 (b) CCCATGTAAGAGAGACCAAGC

coi1t T-DNA LB (c) CTTTAATAGTGGACTCTTGTTCCJAM1-VP16 (d) AGGGATTCACAAGCTTTTCGJAM1-VP16 (e) TCGCCGTCTAAGTGGAGCTC

5' end of JAM1 promoter AAAGCCACGTGGCGCATGATGTA3' end of JAM1 CDS TATATCACCAGAGACCTGTGAACTAGAAGA

5' end of MYC2 promoter CCAAAATCTCTCTACCTTAACTACACG3' end of MYC2 CDS ACCGATTTTTGAAATCAAACTTGCTCTGAG

5' end of MYC2 promoter CCAAAATCTCTCTACCTTAACTACACG3' end of MYC2 CDS TTAACCGATTTTTGAAATCAAACTTGCTC

5' end of MYC2 promoter CCAAAATCTCTCTACCTTAACTACACG3' end of MYC2 promoter ACTCATATTCATTCCATAAACCGGTGACCGGT

5' end of JAM1 CDS ACCGGTTTATGGAATGAATATGAGTGATTTAGG3' end of JAM1 CDS TTATATATCACCAGAGACCTGTGAAC5' end of JAM1 CDS GATGAATATGAGTGATTTAGGTTGGGATGA3' end of JAM1 CDS TATATCACCAGAGACCTGTGAACTAGAAGA5' end of JAM1 CDS GATGAATATGAGTGATTTAGGTTGGGATGA3' end of JAM1 CDS TTATATATCACCAGAGACCTGTGAAC

5' end of VP16 ATGGCCCCCCCGACCGATGTCAG3' end of VP16 CTACCCACCGTACTCGTCAATTC

AACACGTGAAAAACACGTGAAAAACACGTGAAAAACACGTGAAAAACACGTGAATCGAGTTCACGTGTTTTTCACGTGTTTTTCACGTGTTTTTCACGTGTTTTTCACGTGTTGAGCTAACACATGAAAAACACATGAAAAACACATGAAAAACACATGAAAAACACATGAATCGAGTTCATGTGTTTTTCATGTGTTTTTCATGTGTTTTTCATGTGTTTTTCATGTGTTGAGCT

3 x G-box-F, 5' labeled TTTCACGTGTTTCACGTGTTTCACGTGTTT3 x TG-box-F, 5' labeled TTTAACGTGTTTAACGTGTTTAACGTGTTT

3 x MBS-F, 5' labeled TTTCATGTGTTTCATGTGTTTCATGTGTTT3 x G-box-F, non-labeled TTTCACGTGTTTCACGTGTTTCACGTGTTT3 x G-box-R, non-labeled AAACACGTGAAACACGTGAAACACGTGAAA3 x TG-box-F, non-labeled TTTAACGTGTTTAACGTGTTTAACGTGTTT3 x TG-box-R, non-labeled AAACACGTTAAACACGTTAAACACGTTAAA

3 x MBS-F, non-labeled TTTCATGTGTTTCATGTGTTTCATGTGTTT3 x MBS-R, non-labeled AAACACATGAAACACATGAAACACATGAAA

3 x mutated G-box-F, non-labeled TTTAGGTCATTTAGGTCATTTAGGTCATTT3 x mutated G-box-R, non-labeled AAATGACCTAAATGACCTAAATGACCTAAA

CTCGAGTTCACGTTAAAATAGCGCACGGTCGACATCTATAATAAAGACACAGCCC

mutation for 1st TG box CTTCATTCTGACATTTTTCATTTTCTCCATGAAAAATGTCAGAATGAAGTTAATTC

mutation for 2nd MBS GTACGATTTGACCTTCGCATGCGATTTGCATGCGAAGGTCAAATCGTACCATTAAT

mutation for 3rd G box GACGATGTGACCTCCTTTTTAACCGTTAAAAAGGAGGTCACATCGTCGTGCAG

mutation for 4th and 5th Gbox and TG box CTCGAGTTTGACTAAAAATAGCGTGACCTCCTCGCCACGCGTGGAGCTCTTCATTAGGTATAGCTAGACTGGTGGGTCGACATATCTACTTATTTTTCTCTATTAGA

mutation for 1st G box GTTAATATCCTGACCTAGATATCAAATTTGATATCTAGGTCAGGATATTAAC

Xba_HSP_F AAACCCTCTAGAATATGAAGATGAAGATGAAATATBam_HSP_R AAATTTGGATCCTTATCTTTAATCATATTCCATAG

JAM1-F CCCATGGATATGAATATGAGTGATTTAGCJAM1-R CCCATGGTTATATATCACCAGAGACCTMYC2-F CCCATGGATATGACTGATTACCGGCTACMYC2-R CCCATGGTTAACCGATTTTTGAAATCAAAC

JAZ3 promoter (418 bp)

mutated JAZ3 promoter

TAT1 promoter (1 kbp)

mutated TAT1 promoter

HSP terminator for pGL4.1HSP vector

His-tagged protein expression

Supplemental Table 2. Primers used in this study.

EMSA

5 x MYC2BS, for reporter construct

5 x G-box, for reporter construct

Pro35S:VP16

Pro35S:JAM1

Pro35S:JAM1-GFP, Pro35S:JAM1-VP16

ProMYC2:JAM1

ProMYC2:MYC2

ProMYC2:MYC2-SRDX

ProJAM1:JAM1-SRDX

Genomic PCR

realtime PCR

ProJAM1:GUS

Entry clone

23


24

Supplemental References :

Fernández-Calvo, P., Chini, A., Fernández-Barbero, G., Chico, J.M., Gimenez-Ibanez, S., Geerinck, J., Eeckhout, D., Schweizer, F., Godoy, M., Franco-Zorrilla, J.M., Pauwels, L., Witters, E., Puga, M.I., Paz-Ares, J., Goossens, A., Reymond, P., De Jaeger, G., and Solano, R.(2011). The Arabidopsis bHLH transcription factors MYC3 and MYC4 are targets of JAZ repressors and act additively with MYC2 in the activation of jasmonate responses. Plant Cell 23: 701-705.

Heim, M.A., Jakoby, M., Werber, M., Martin, C., Weisshaar, B., and Bailey, P.C.(2003). The basic helix-loop-helix transcription factor family in plants: a genome-wide study of protein structure and functional diversity. Mol. Biol. Evol 20: 735-747.

Sievers, F., Wilm, A., Dineen, D., Gibson, T.J., Karplus, K., Li W., Lopez, R., McWilliam, H., Remmert, M., Söding, J., Thompson, J.D., and Higgins, D.G.(2011). Fast, scalable generation of high-quality protein multiple sequence alignments using clustal omega. Mol. Syst Biol. 7: 539-539.

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