Figure S1. CLUSTAL O(1.2.1) multiple sequence alignment

Figure S1. A CLUSTAL O(1.2.1) multiple sequence alignment AT5G48657.1 --------------------------------MNVANPNEYPDMNPNAAQNRNMSRPDQQ 28 AT5G48657.2 MANRPHVPKFGDWTEDAPFTVVFEKASKSKKNMNVANPNEYPDMNPNAAQNRNMSRPDQQ 60 **************************** AT5G48657.1 PPNHNVRPRHERFNSRDETEFRPSPAHNERNNRVRSVPPTPETYNHQTYGGGGRSMGNPT 88 AT5G48657.2 PPNHNVRPRHERFNSRDETEFRPSPAHNERNNRVRSVPPTPETYNHQTYGGGGRSMGNPT 120 ************************************************************ AT5G48657.1 EINRRQSRDHVPARPIRNLRGQSSERVATIPPFPGTGSNMENQSYTLIFDKVKEDRNQAR 148 AT5G48657.2 EINRRQSRDHVPARPIRNLRGQSSERVATIPPFPGTGSNMENQSYTLIFDKVKEDRNQAR 180 ************************************************************ AT5G48657.1 SYNGTDHSTPTRPIIDQHHQPLPSSPKVDNQNIISLSFINPLFLQLTLLCVGLLFSSMEP 208 AT5G48657.2 SYNGTDHSTPTRPIIDQHHQPLPSSPKGCCFPPWS------------------------R 216 *************************** * AT5G48657.1 KGKLMKMIISLFYIILFFFICFCLIFFNNIFVILMEM 245 AT5G48657.2 KGS---------------------------------- 219 **. B CLUSTAL O(1.2.1) multiple sequence alignment NOI1 GKPLPKFGEWDV 12 NOI2 DRPLPKFGEWDV 12 NOI3 GRALPKFGEWDV 12 NOI4 GRPLPKFGEWDV 12 NOI5 ARPLPKFGEWDA 12 NOI6 WLSVPQFGDWDQ 12 NOI7 WLSVPQFGDWDQ 12 NOI8 SGQIPRFGEWEE 12 NOI9 RGHVPAFGSWDW 12 NOI11 RPHVPKFGDWNN 12 NOI12 GRPLPKFGEWDV 12 NOI13 ARPLPKFGEWDV 12 NOI14 WTPVPQFGGWDQ 12 RIN4-RCS1 RSNVPKFGNWEA 12 RIN4-RCS2 VTVVPKFGDWDE 12 :* ** *: Figure S1. Alignments of both NOI10 gene models and of the predicted AvrRpt2 cleavage sites of all the NOI proteins. (A,B) Multiple sequence alignments were performed with CLUSTAL O(1.2.1) (http://www.ebi.ac.uk/tools/msa/clustalo/; Sievers et al., 2011). (A) Sequence alignment of both NOI10 (AT5G48657) splice variants, the conserved AvrRpt2 cleavage site is highlighted in grey. (B) Sequence alignment of AvrRpt2 cleavage sites of RIN4/NOI protein family members. Note: the representative gene model of NOI10 (AT5G48657.1) does not contain a cleavage site. 1

Figure S2. A 2

B ATG - start codon GCG - exon gta - intron TAA - stop codon cta UTR NOI1 At5g63270: tagggatttacgcgggaaagaagaagaggtgtaataatgttgtttgatttggttaatgc Insertion (SAIL_1173_C11) tgttgtcaacgacgacgacgacgtgtttccctcttatgtttctccttcctctctctatattacaactg catttcccttctctctcctcccacctccctctctctctctcaatagatctttgatatttcattttgct ttgatctcaacaacaatggcggtaagaacaaacaacgtcttcatcaagtctctctcctttcatcatca ccacgatgatcagtttcagaaaaagtctctatagctttcttgatgtttgatgaatctctgtttctttg gtgcaggaaaacaaagggaaaccattgccaaagtttggggaatgggatgtgaacaatccggcatcagc CGAAGGATTCACAGTCATTTTCAGCAAAGCTAGTGACGAGAAGAAGACCAAGAAAGCATCCGGCGCTG GCCCTAATAGTCTGGTTTCACCTCAGAGAAACCAAAACTCCGATCAAAACAATAACCATAGTTCCCAA AACCCTAAAGCTAAGgttgttttctttttctcattacattcaacttcttagctctttatccaagaaca tagatttgctcacctgatttatgctttctgcgcagaacaaatggttttgcttccgttaaatcctgcca tcttctgaagaatccgaggggtagggaagctgaagcagttttgcagagaggtctcaaacaaagtgaaa caaaagggttaacgcttaaagacccatatatctcagctttcttcttcttctctcttctgctgatatta tacttgttcagtatactttttgttcctttctttttgtttgccaacattttgcgcacaatggatataat ttgtatcttttgactctccatctaagaactatggaaataagattttgagtaattgtccttcgcagaca taatgctattagctattattcatatcgacatggctttctgtacatcgttccaatagct NOI4 At5g55850.2: agtgattcacgtcacttgggtctaataactccgtttatataaaccgttacagacatcgcacaccgaga gagagataataaagagagaagcaaaaaagaatttttatagaaagaaaggaaaaatcagaaaaggagag aaaaattaaaaaagtctctcttttctctttttcttcttcgtctcccatattctccgatcaaaaatata tttctctctcactttccttggttttctcggcggcttcttcttctcctatttctccctcaaatcgtttt ctccagttttcttgggataaatcttcctcctttctgcagacgccatgtcggtgcgtttaaatctcctc tctttattatatacatatgcttgagaacagttttgaattcgatttgtttggttttgagaattttagtt TCGTTTTGGTGAGTTAGATGATTATATGTTTCCTTTTGATTGGAGGgtttttgttgtttcttttatat ccgctgaaaatcttagattaaatagaaaaacagatgtgaaattgtgattctgatgagctattgtcttg ttgttgaatctgataatcctgattctggatctcgggttttattcaagttttgatctttttctgaattt gattgattaagcttagatttttgattcagggtttctgattttggatttgagatgaaataaaggtttga taaagagaagttgttttgttgtctttgtgttagcttatctttttttccccagattgttgttctttgat tttattgatagtgaaagaggcatgaatattgtgtggcattgtttatggacaagcaaaagttctttttg ccttgtggatttgtttgcttatttagagacacaattgaaatccactagtttcaaacttttggttttat tattttctcacctaatcaaatggattataaagttgtttggaactttaaaatttctcttatggtttggt tttggtaatccttttgacttgtgctctatattgtttgctacaaaatggaattataattagtttgggca aattgtacaaaccattttttcctcgttattgatgatagaaaaggttccattccttaagtcctcattct tatttacttggttgattctttttttcggtttatccttgcctcttctcttttgtatgtgctcccactta agacttgcttagcttatgaattcatctacttggcaagattgaacttatttcttgatttcaccatttgg tttttatatgaacaacaggacaaaggtcgtcccttgccaaaatttggtgaatgggatgtgaatgatcc AGCATCAGCAGAAGGTTTTACAGTGATATTCAACAAAGCTAGGGATGAGAAAAAGACCGGTGGCAAAC CGGGATCACCCGGTAAATCCAGTGAGGGTCATGTTAAATCTGGAGGAGGAGATCCTAGTAAACCTCAG CCTGTAAgtttcctcttcttcatctttatctccttgatccattgtcacactaaatggggttttatctt tggtttcagaaaaaatggctctgctgcatgcaagctccagctgtggactcttgacagacacaaagatg GATTTGCTTGCTGCTAAAAAAAAAGTCACAGTGCTTACACATCTAAAAGCAATGGTTCTTTTTATGTT TTATTGTCGTCTTTCTTGA Insertion (SALK_042784C) 3

aattacccacacacacaaaaaaaaaaggttcctaagatgatttggagctcacccacctttagtatcac gattatgattccttcttctttgaagttgtttcttcttcttaaaatgctgtaaaatcgtgtctccattt tattggaaaaaaaaaaggaagaaaaacgaaaaaaaacacagttactgtgtctatatgtaatgattctc ttggggtctttttgttttttgattttatggtgtctttttccttgattttttaaattatattatcttgg gctcctttttaatattttcctccttagtaaatcaaacaagtttcattaatgaaaacgaacaagaactt NOI5 At3g48450: atattacaatttcacctctctttttcccagatcatcatcatcctctctctttctctagattttggata tttcctcgtatcggattcgaaatcctctgtttctaattccaacaatggcaacggtaagaacaatatgc atctcttatatctacgaattctgattcatcttacatttcttgatatgatttcaaaacttccctccttt catacaattttccaaatctcttatcttgaaaaaaagtctcaaactttggctagaaatctgcatttttt ttaatgatgaacaattctgttttttgttttgtttgtttggtggtgtgtagggaaatagagcgagacca TTGCCAAAATTTGGGGAATGGGATGCGACTAATCCTGGCTCGGCTGAAGGATTCACTGTCATTTTCAA TAAAGCTCGCGATGACAAGAAGACGATGAAAACCGCTGTCGCAGGACCTGAGAGTATTGTTTCACCTC CAAGAAATGAAGAACCTCCTAAAAACAACAACAACCATCATCACAACCATCACACCCGTCATTCCCAA ACCCCAAGATCAAAGgtacttttttgccacagatactcaaagggtatatattaggtatttctctgctt gacttgttatgtttttgttcttttgttgtgcagaagaaatggctttgttttcgttaactctggtattg Insertion (SALK_118446) gctctgttctgagaaaacccgacgacaagcaagcagaatcgtggagcgagtaatctccaacagattat agcgaccaaaccagtgatcatctaagatgttgttgcaaacttctcttcttcttattgctcgttgagtt tctttctcttttgttttggacacatttgggtatatgaatatacgtttttcttgatgattcttagtttc ataagtatatataaag NOI7 At5g09960: tagtgattgctgattgggtataattcgagctgtttttttgctgtaataatttcacaattctctttttt tttcgcttttaaataattttgtctctccatcttcttctcttttgctagtctctcatatcagctaagaa aagaaattcagaacaaaaaaataacacaaagctctgtgtttctgtctatctgttgaatcaaatcatat GGAAGACGATCGAAAAGAGgtataaccatctgttttattatctcttgttagtttccccataaagtttc tttaggaaactttgattttactaagaaatcgagatctgcttgtttggattttttcgagaattgaacat tggttttgtagagctaatcaagctccaagcactactgatactgtttatattgaagattttgattaaaa atcagcactaatcttggtaatcggaaacaaaaaaaattgagttagtagtatattctaaagtttcatcc tttttcacatctcgataagctgaaattttgttatcacgaggtatattttctactagtacttggtaagt tcgaaactttattggattgataaagaaaagatcttactttaccctgattttatcgatcattgattaaa ccaaagcttgtaaattgtcccataaagtcaagaacttgactttgtttcttagagagttcgtcctcttt cttcttcacttgcttgctcatgatgcatcgttttgtaaatagctctgtttttttttcctttaagtata tgcttcattatgatttatgaaacctttttaagtttggtaatgtttgtttcgaaatgtctttgtttttg cagaagaacactccgtggctatcagtgccacagtttggtgattgggaccaaaaaggaggaggaacaat GCCTGATTACTCTATGGATTTCACTAAGATTAGAGAGATGAGGAAACAAAACAAGAGAGACCCTTCTC GAGCCAGTTTAGGCAATGAGGAAGAGCTCATTAAGCCACCCGAGTCAGCAACATCAACTGCTGAGCTT ACCACGGTCCAAAGTGAAAACCAACGAGAGTTCTCTCCCAGCCACCATCATCAACCACATTCTCCTTC Tgtaagattcttcaccatt Insertion (SALK_007305C) atttggtttatatcatattattccttaattatatttgcattactatcattaagacatttagaggcata ttactatattagcacatggttttgtaatcttgcaatctatatacagtattaaaattttgaatagttgt atttgagctggtagtgattatcactgtggaatagatgagaatttggtgtggttgttttttttgacaga CGAGGAGAAGTATGTTCAGCTGCTTCAACTGCTGCGTTAAAGCTTGAagatttcttcttgagcaaagt agcagttttattattgacttgtgatttgaatgtggaaatgtgttaatgtcatgacactttaatatatg ttccaatccatttttctttttctttgggaaccatataagctctattacaaggaaaagattatggggga aattaagtatacacaaaaacataatcgtctctattagtacactct NOI8 At5g18310.2: 4

5 gactatttagacttttagaggtaaggacaatatcttctttgcttacataccccactcttgttataaaa gacagtaacctttcttgagatcactctttcttatttgtgtgttagtgcattgagagagagagagagag atagaaagagaaagaaatcgtcatggctcatgtaagtgaaacttctctaatcctctatccttgtttct tcctgtcgtctttaacatatagatcgtcaaaac Insertion (SALK_101426) ctgcctaataatgtcttatcctcacataacacctttttttttatctccatctctctgttacttagaac cctccattcatatatacactaggtgccaaatttggtcacgtgtttggcccttcacgttcttgactctt agctagatccttgtttcttgtggattcttgaatcttaaatcttgaatccctgagttttgttggttctt gattttgattctcgattctttagtcattgattattcgatctttgttggtttttgattagagatttaat tagcatatttatgtactaataattgaataaatgggaacaggggagatatgatacttacaagaagaaaa GTGGACAGATTCCTCGGTTTGGAGAATGGGAAGAAGCAAATGAGATGCCAATAACACAATACTTTGAG AATCCAAGACAAGCTGGTCTGATTCGTCACCAATACACAACAACTTCTTCTGCTTCTTCCACTACCAC TTCATCTTCATCTTCTTCTTCTTCTTCTGCAGAAGCCTTAAAGCTTGCTTCTCACCATCCCCGTCCAC GTCACCTCCACGCTCAAAGACAGgttcattcatctaatcactacttatttttcttaattatataacat ttatttaactcgataaacgtaattaagtatctaaagaagaacgcgttgtttgtttgattaatgacttg gcgtagacggcggggacgaaggagaaaagaggaccacaaaggcgtgtgcgtgacgtcagtgcacagtc GGACAAGTATTACATTGACGTCAACGGTGTTAAGCAGTTCAAAAACGACGTTGCTCTGACTTGTAAGC CACCTAAGCCCGTTGATGAAGATCTCTACAAGATTCCTCCTGAGTTTATCCATTCTTCAACAAGGgta agagctccttctcacacacaacttattttactgcattttataacaaagttttatatatttttaatcaa cttttcaattttatatcactttttctttaactttgtttgttatttaaaaagagttttaaaataaattt ggtgttgcagaagagaaggcctagctttttagcttgtttggttccatgcgcatgaatcgaaaaggaag agcgaagaagaagaagaaaaagtaggatcatgaatgaataatgtaatttgttttttgtttttgtaatt tcatcctaatatacagatatgatatatacgtggtttgacagaaaaagagagcatacgagtggtttttg aaaaagagtaacaaagtaatatcgtagtaaactagagtaatatatccttttgtcttgttacatatcta ctttataagattttctctcatttttttttcacgt NOI9 At5g48500: tatattattttcacttaccctacactatatttttgaaaacaccactctatgctcagagacacacagtc cctcactactctttctctatctcctacaagtagactttaagacaacttttgtctcagtgttgagtaat gtaaaatacccatctggttatttattacgcatagatctctctcttctctctttctctatttagattac ttaaagcagagaggtccgaacatcatcaatcaaacccattcacaactactacgacaatggactacgta agctccttcctctttcttcttcttcttcgtcatcttcttgtgggttatacaaagtttttaacttttta tgttttctcaatcaggaatacggaggaggagagtactgtcggagaggacac Insertion (SALK_150777C) GTGCCAGCTTTTGGAAGCTGGGATTGGAACGACGCCGTTCCATTCACTCAGTGTTTCGAGACAGCAAC AACTCAACAACCAGCTTTTCTTCACTATGCTTCTTACCCTCAAGATCGTGATCTTTACCTCGCCGGTG ATCTCTACGACAACCACCACCTTGTAGCTCCGGCCGTTATCATTGTCCCTCGTCGCCGGgtactttct tgattcatgtattcaacacgacgccttgagtttcttgatttagtcttatgttcgtggaaaatgtttat ttctaggctaaagtgggtcaagaaccagcggagaagaagagaaatgtgtcgaagcagcaacacaacta TAAGACGGAGGCGCGTGAGTTTAACGCGCCGGTAAGCTGTCCAACACCGGTTGTGAAGCGGAGGATGA AGGCACCAAAGCCTGTGGATGAAGACTTGTACAAAGTTTCCCCACAACTTCTCTCAGTCAAATCCAAA AGGgtaattaaactttaatttaattaacttgttattaaattagtcttatctttgttagtctgcttcga ttaatgattttgattatggaaaatgttaataattgagatttgcagaaaagaggaggaggtgggtttgg GTGCATTTCAAGGTGTTTTTTGCCAACACGGGTGCTTTGAagaaacaagcgtgagagacttgagtcag aagaacactttgtttttttttcttttctttttctaatcctattatatatatcaaaagactatgcttgt gagagtatgtatttatgaatgcgtaggaaaaaaaaagatgaatgctaaaagatagtagaccaatgtag cttttgcgaaaagcttagccatattatattaactaaactataaatgagacttggttttgcaaattatg NOI10 At5g48657: ttttttcttttgggtgtggttttagtcttctttttattttgattttctaaagcttattatcctacaag tgtagtatttattatacccacaatctctgagttttattccgtgagaatggagagtaattgtcaaatat ttgtagttctaaatatggatcctatatcatcctgtttaggacatatagtgttttgttttagaatcggc ggtatcatgagattttcttttgtttgacatttatgcgtaggaagatcacctaattctttttggtcatc

ttctgaacatcaatataattcatatgtaatgaaattatacttagattctacagatgaggtttctgttc aatcaaaaccaaaaaaaaagagataaggtttatgtttcttgcaccttaagattcagctctaaacattt agccaaaaacaaagtttcaactgtaaatctctggcatcaactccaaaataccgtatatatgagatttc gattttctaatatttatgtctcctactttcaatttaaatgcaagcctatgtttctcaaaatctttttt ttttggttcttgtggttatagaatcgtccacacgtgcccaaattcggagactggaccgaagatgctcc attcacggtcgtgttcgaaaaagcaagcaagagcaagaaaaatatgaacgtggccaacccgaatgaat ATCCAGATATGAATCCAAACGCTGCACAAAATCGAAATATGTCTAGGCCTGACCAACAACCACCAAAC CATAATGTTAGACCAAGACATGAAAGATTCAATAGCAGAGACGAAACCGAATTCAGACCGTCTCCTGC ACACAATGAAAGAAACAACAGAGTTAGATCAGTTCCTCCCACACCAGAAACATACAACCATCAAACAT ATGGTGGAGGTGGCAGATCA Insertion (SALK_080643C) ATGGGAAATCCAACAGAAATAAATAGACGGCAATCACGTGATCATGTCCCGGCGCGGCCAATACGCAA TCTTAGAGGGCAAAGTAGTGAAAGGgtatgcataactttaccattttatagtacaaaatatgtgtcac aactcataaatatatatcgctaagtaagtgtatttttctcaattaggtggcgactattcctccatttc CTGGAACGGGTTCTAACATGGAAAATCAGAGTTACACACTTATCTTTGACAAAGTGAAAGAAGATAGG AACCAAGCGAGATCTTACAATGGAACCGATCATAGCACCCCGACTCGACCCATCATCGACCAACATCA TCAACCATTACCTTCTAGTCCCAAGGTAGATAACCAAAATATCATTTCCCTTTCTTTCATAAATCCTC TTTTTTTACAATTGACGTTGTTATGTGTAGGGTTGCTGTTTTCCTCCATGGAGCCGAAAGGGAAGCTA ATGAAGATGATCATTTCCCTTTTTTACATAATCCTCTTTTTTTTTATTTGCTTCTGCTTGATTTTTTT TAATAATATCTTTGTTATTCTTATGGAGATGTAAggagtagtttttgctaaaacattatgcaacttct gttactctatttgcgtttttttgaacaatactcagattcatttataggggtgaatccttttattcacc tcctattataataaac NOI11 At3g07195: agagaagaaacggagacattcaaac Insertion (SALK_028339) tttacagagagtaacacgaatcgtcaacgcctttatatattacacacacattttatgtgtatctctgt gtttcagaatcagacctaagccattgttgaagcctttcctcatttctccatttagatcaagaaatatg GCAgtaagttcgaaaactcttttctatctaacaaaattgtgttatatccccctctcgtttgttatcat cttaaatcttgttatcaaaccatgttttatgacgaaaccatcagagtgttggaattttagcttgttga tccgaatttgtctgttcatttcccaatgattgtttaaattgtttgtttgatcatattagaacagagat ctgagtctttgtttcatatcagttgactttcgtatatattcttcttctgaattcaatctgttccagta tgtgttgagaagctaaagatagttctcataggtctgattagggttcatatacatatcgctttcttgca tctcaagtttgatattgaagtcattaaagctcaaatcacttttgaccattttctcaaaaacatttttt attgttgaatgattgattgcagaaccgtccacacgttccgaagtttggggactggaacaaccaagacc AACCATTCACGGTCGTCTTTGACAACGCAAGGACGAACAAACGACAAGACTTGTATGAATCTATAGAG AAACCAGAGACCAAACCTCAAGAGCTAGCTCCTCCTCCTCCTCAGCCTGCTCGTAGAATTCAAAAACC GGAAGCACCAAAACCGGTTAAGCAAGACACGCCGAGAGCACCTCCACCAACCGAAAAAAACAGAGTCA AAGCTCCACCGGCAGATCAACTCTACGGAGGCGGGGGCAGTGGAGGCGGAGGCAGGAGTGGAAGTGGA AGCGCAGGTGGTCTTTATGGAGGTTATGGAGGCGGATCAGTCGGGAACCAACGGCAGCCACCAGCTCC TCGTCCTACGCAGCCTAGACAAAATCTCAGAGGCGGCAACAACGGAAGGgtatgaaagacacaatata taaatgaacttggatctttgtttgcgatttgcattaacatgttattatgtgattgaagagtcacatta taatctaagatatatatatgatatgaatgaaaaagtttgcttatagtcattatatgatctgtagggag GGACAACGATACCGCCATTCCCAGGATCAGTTGGTGCAGGGGGGGACATGAGTTACACACAAATTTTC GAGAAAGTCAAAGAAGAGAGAAACGAAGGTGTCATCAGACCCAACGGAGGAACAGCCGGCAACACTCC TTCCCGCCCCATCAACAGTCAACATGATCAATCTACCAACCAAACCTCTTCCAAGgtagtttatttat taatatacaaatgtttaattacaaacacaactactttgtccatcgttagaatttgtgttttgatgtga attgggatgtgcagggctgctgcttctcatggtgccgcaggggaagtaaatattgaagacaatgtaca tatttatccctatttcgatatatctttttcttttgtaacaaaaaggagtgaagtttttgctgacaacg ttggagcttctacaatttttttttaaattattttaatttgtttctcaagtatggatggtgtagtttat ttgtgttgtgtatttgaattttgtaatgaaaaattgtttatttctgctaccgacaaaac NOI13 At4g35655: 6

aagcaagattttgaactctttttgcatcaaaattaggtaaaaacactaatcaaattacttgttaaaaa agg Insertion (SAIL_147_D02) cataataaatgacttgaagaaaatgtgatacaaatttagatttgttgaactcattgaccaggaggata ttatataaggtctctgattactaaacaagcatgtagagaaccaaagataacagtagcaccaaataaga aaacagagcagtcatggataagaagcattttgctataattagggttttacagggtatgacgcgcaaag accaaagattttgaacctcagaatgtttaagaagaaatgcgtaaagagaaatatcttgtctccgtgtt tcatacgagtacattccgtaggaacatttcataacaaaacttctctgtttctgctttattgaaatctt ccaactataaaatacaccctgagtccatcatcttttgatcaattagaacgatctcagaatagaagtca ttgaaaaagaagaacaacaataaaaggaagaaaataatttgtttttgagaaattttgctctgatttgt tgatcctgaagattgaaatggcatcgaatagtgacgcaaggcctttgccaaaattcggagaatgggat GTGAATGATCCGGCGACGGCAGAGGGATTCACCGTTATATTTAGTAAAGCCGGCGAGGATAAAAAAAC AGGTCGGAGCTCTAGCAAGGCACCTTCACAGAGAAAACAAGACGGCGTTAAACCAACCAAGAAATGGC TCTGTTTTACTTTTTCTTGAtttcctgtaaatttcttaaaacgtttgagaatggcattgagaaactct taccttagaaccttttctcatttgcttagcatttctctttataagatgttgcagagacgagaaagtag atagtttcttgtatatcttgtttgatgtttaatgtttaatgttatgtttcgtttgaagcttaagacta NOI14 At5g19473: aaacaatttgtactataaaggaaaaagtggaattttacaagagagatcgttctcctcctcgtcatgaa caccttcatcaatgtttaagaacatctttcttctaaaccaagcaatatctatttatggataaaagaag GGAGgcatgctctctaatttttttataaaatcttgtttcttttcttttatccaactattactaaatca tcgatctttatgatgtctgagttacaatcgagacatgttgagctttcatcggttaatgcgcgttttat agaatggggaaaattaaccgatgcaatctcaacatcgatctcattgttcttttttttttttccggaaa tatttggttccctaagcaaattatatgtttatctaacgagttttattcttggatacagaatgctgcat GGACTCCAGTTCCACAGTTCGGAGGGTGGGACCAAAAAGGACCAAATGATGCTACAAACTACTCGGTT GTCTTTTCTAAAGCTCGCGCAAACAGGAAGCAGAACAAAGCTGGTGTAAGACATTCTAGTTTGGGAAG CGAACAAGAACTCATGGTAAGTGTTCGTCGCAACCACCAACAACTTCACCATCGCCATGAAACTCAAG ACGATGACCCTGTCATGgtaattaaattatctctctctacttactcttccatggattctttggttcat attcaccttgtgatgatccttttccccttagtcaacgcgcatggatcatcacaaggtggtataacaat tcagga Insertion (SALK_110602) attgattttctctttgcgtcttttgtttgttttgtgtttttgcatgcgcctaaaaagaatatttcatt ccatttctttcttaactcagagacataattgaaccatgaatgttattcaacttaattatgagttaata catgcatcatagtatcaggtctcatttatgaatatgtaatgatagtgtatctttaaatttgatgttat agtgtattggcttgcagtttgtttccatggttttgaaatgatgtgttttgttcgaacaatgaatttat agttcgtttccattttcccattttctactttctttttatttgttaggaaaactttgtttttttttttg cattttttccataacatttcaataagcaaccatattttaacattatattcccacctaagctaaatttc attacttctattgttagataaaaatataattaaaccaactaaaagtctgaatcatattataatctttg cattatgttttaactatatgttgggagttacataaatttgttttgcttggagcctccaaactttgaag aggcaaacatgaacgaaatgtacactgcttttctagagcgaatatgaaaactccatctctagtttgta taacagtcgaaatcaccttgttatacgaagcattatgcaatatgtgaaatgtattaagaggaagcatg ttgtcaccgtcataagtctctctaacattactaatttaattaccatgcagaagaagaagaggatcttg ACCTACATTAATTGTTGTATAAGGCCAAACTAGggctttgttcttctacaaagtcggaatctttctct cttcagcgatatagcaaatatgtctcaagtgtgtaaagacagtagtgtaacgtttctgtaatcttctt tatcacacacaaaaaaaaaaaacgttctgttctatgtgcatcatctattaagatacccctggttgtgt tgtgttgtgatacattttgacttcttcctacatc Figure S2. Positions of T-DNA insertions in the NOI-genes. (A) Schematic representation of the T-DNA insertions with respect to the exon/intron genomic structures of the NOI genes. For exact location, see sequence information in (B). (generated with Exon-Intron Graphic Maker v4 (http://wormweb.org/exonintron). (B) Sequencing of genomic regions flanking T-DNA insertions in NOI-genes. Genomic regions covering the NOI genes were exported from the TAIR SeqViewer tool (https://seqviewer.arabidopsis.org/). T-DNA insertions are indicated, based on results obtained by sequencing PCR products of the respective regions. PCR reactions were performed using 7

the primers LBa1 (SALK lines) or LB3 (SAIL lines) plus respective RP primer (Table S2). PCR products were separated by gel electrophoresis and isolated bands were sequenced with the primers LBb1.3 (SALK lines) or LB2 (SAIL lines). Figure S3. Figure S3. Verification of knock out / knock down of transcript accumulation in noi-t-dna insertion lines. Leaf material of Col-0 and noi T-DNA insertion lines was used for quantitative RT-PCR experiments with NOI gene-specific primers (Table S2). 8

Figure S4. A CLUSTAL O(1.2.1) multiple sequence alignment of AvrRpt2 homologs Pst 1 --------------------------------------------MKIAPVA--------- Ea 1 --------------------------------------------MKVSH----------- Ac 1 --------------------------------------------MRISSFSR-------- Rs 1 --------------------------------------------MKISHNGILGTHIPDT Mh 1 MKSRTYFSPNHSADR-FATRPNDDETATWVARLSGGQMQANDAHQQLSEPSLGGFDTDLD Sm 1 MDE-----TGVALRPYDASPENDHEVATWVARLSGGQMQANDAHQQLSEPSLGGFDTDLD Cf 1 ------------------------------------------------------------ Bp 1 ------------------------------------------------------------ Aa 1 ------------------------------------------------------------ Pst 8 ---INHSPLSREVPSHAAPT----QAKQTNLQSEAGDLDARK-----SSASSP---ETR- Ea 6 -------------LTSPAPVVIEHQPRQSEKVSRDGDVI--------------------- Ac 9 --AAGEN----EIPHTEEHQKKEGRTSQKMVSSEIFDQISKRDHTGVSSVAPRVNIGVAN Rs 17 SSQTGEA----SRHQEAQPNS-SGRPRPAGLA----GLQPKR-----NGLASRISVNKD- Mh 60 SDTIGRASMGESMHMPAQPSLGDGEPSSACLTPEAGVSQPPR-----LNVISRHS----- Sm 56 SGTIGRASMGESMHMPAQPSLGYGEFS--------------------------------- Cf 1 ------------------------------------------------------------ Bp 1 ------------------------------------------------------------ Aa 1 ------------------------------------------------------------ Pst 52 -ALLATKTVLGRHKIEVPAFGGWFKKKSSKHETGGSSANADSSSVASDSTEKPLFRLT-- Ea 32 ------K-PWSQLPAAAPSFGGCFGKSKKSRG--------YDSGSSSGSRSNAGFRLN-- Ac 63 FDRNASKMSANVEVGNLPAFGGFFKKKSTG--------QSSSSGRGEESSTLPMFRLR-- Rs 62 ALPKRTDQTTQSAITALPAFGGWFGKKSSK-----SKGASSSGADASASGELPMFRLR-- Mh 110 ---SAGQPGVISPRSSAPSFGGCFGSKAASSSHNEAKAASSSHNEASEATELPQFRVY-- Sm 83 ---------------SAPPFGGWFSKKSS-----GSKAASSSHNEASEANELPQFRLY-- Cf 1 ------------------------------------------------------------ Bp 1 ---------------------------------------------------MVDVRLNVG Aa 1 -------------------MGG-------------GHLQSAGSCSSSG------------ Pst Ea Ac Rs Mh Sm Cf Bp Aa Pst Ea Ac Rs Mh Sm Cf Bp Aa Pst Ea Ac Rs Mh Sm Cf Bp Aa 109 -----HV--PYVSQGNERMGCWYACARMVGHSVEAGP--RLGLPELYEGREGPAGLQDFS 75 -----HV--PYVSQQNERMGCWYACTRMLGHSISSGP--RLGLPELYDS-SGPQGLQQRE 113 -----HV--PYVSQEDDRTGCWYACCRMLGYSVASGP--RLGLPELFDPNHGHSHLQSFE 115 -----HV--PYVAQGSERTGCWYACARMLGHSVEAGP--RLGVPELFNPESGHSHLQHLH 165 -----HV--PYISQGSERMGCWYACARMLASSVEAGP--RLGLPELYSPQSGHDGLQDPT 121 -----HV--PYISQGSERSGCWYACARMVACSVETGP--RLGLPELYNPQSGHDGLQDLT 1 ------------------MGCWYACARMIGHSAEAGP--RLGLPQRYSSDAGHQALKEPA 10 LVDQSGVKMTKSGSKEVKNACWYAAACMVSYYFRPGP--RRGIPKLWQADRGLPEA---- 17 ---GSKLAVPYIPQN-GTRGCWEATMNMLRAYFGQPPLDKRQLAYLYDAEGVPKELNGQL Y191 D216 160 DVERFIHNEGLTRVDLPDNERFTHEELGALLYKHGPIIFGWKTPND--SWHMSVLTGVDK 125 DVLRLMRNENLAEVSLPESRQFSANELGNLLCRHGPIMFGWQTPAG--SWHMSVLTGIDK 164 HVERLLANEGLERVPLPSSQQFSLSELGALLYRHGPIIFGWQVPEG--YWHMSVMTGVDN 166 DVEQFIANEGLSKVPLPSSREFSHEQLGALLYRHGPIMFGWQTPQG--NWHMSVLTGVDH 216 HVEQFIVNEGLSKVDLPDSQEFSHEQLGDLLYRHGPIIFGWRTPEG--NSHMSVLTGVDK 172 HAEQFILNEGLSKLDLPDSQEFSHEELGELLYRHGPIIFGWQTPQG--NWHMSVLTGVDK 41 DIEQFISNEGLSRVNLPDSQAFSLEELVSLLEQHGPILFAWHPNQN--SGHMSVLIGVDK 64 RILELCAAEGLVPLDQPD--KITPEWLAAVLTDSGPIWAAGHFLDE--PGHVIVITGIQH 73 DYATRNRKAKLKNVPVPAGKSWDADKIKALLDQHGPLEARIESPHDELVAHAIVLTGIDE 218 ETSSITFHDPRQGPDLAMPLDYFNQRLAWQVPHAMLYR------ 183 PNDAIIFHDPQRGPDLTMPLDSFNQRLAWRVPHAMLYSEN---- 222 HTGSIIFHDPKKGPDITMPLAYFNQRLAWQVPHAMLYGGSS--- 224 HTGHVVFHDPQQGPDLTMPLSYFNQRLAWQVPHAMLHR------ 274 HTGRVVFHDPQKGPDLTMPLDYFNQRLAWQVPYAMLYRGVGAAP 230 HTGSVVFHDPQEGPDLTMPLDYFNQRLAWQVPHAMLYR------ 99 KTSSVVYHDPQKGPDRGMPLSHFNQHLAWEVPYAMMHR------ 120 --SVVHYNDPEGPPARTMLLGTLDSG-RWKPD-GLLVKDRSRY- 133 D-GDVICHDPELGPRQKVSIEMLNAAFDWTSHRTALTAYNG--- 9

B Percent Identity Matrix of AvrRpt2 homologs created by CLUSTAL O(1.2.1) Aa Bp Ea Ac Pst Rs Cf Mh Sm 1: Aa 100.00 23.08 33.12 27.95 27.33 27.95 23.70 27.44 26.09 2: Bp 23.08 100.00 26.90 27.89 29.86 29.17 32.54 26.85 29.86 3: Ea 33.12 26.90 100.00 53.64 56.02 54.46 54.07 50.45 54.82 4: Ac 27.95 27.89 53.64 100.00 52.26 55.65 53.68 50.60 58.57 5: Pst 27.33 29.86 56.02 52.26 100.00 55.79 61.03 52.80 62.74 6: Rs 27.95 29.17 54.46 55.65 55.79 100.00 61.76 61.81 68.92 7: Cf 23.70 32.54 54.07 53.68 61.03 61.76 100.00 64.71 61.76 8: Mh 27.44 26.85 50.45 50.60 52.80 61.81 64.71 100.00 83.08 9: Sm 26.09 29.86 54.82 58.57 62.74 68.92 61.76 83.08 100.00 Figure S4. Multiple sequence alignment and identity matrix of AvrRpt2 homologs. A,B. Multiple sequence alignments and identity matrix were done with CLUSTAL O(1.2.1) (http://www.ebi.ac.uk/tools/msa/clustalo/; Sievers et al., 2011). A. Boxshade representation of the alignment ([LVI]PxFGxW= consensus AvrRpt2 cleavage site based on RIN4/NOIs; C/H/D= catalytic triad; = Y191 and D216 [reported to be important for virulence of Pst AvrRpt2]). B. Percent Identity Matrix of AvrRpt2 homologs (Aa= Acidovorax avenae subsp. avenae; Ac= Acidovorax citrulli; Bp= Burkholderia pyrrocinia; Cf= Collimonas fungivorans; Ea= Erwinia amylovora; Mh= Mesorhizobium huakuii; Pst= Pseudomonas syringae pv. tomato; Rs= Ralstonia solanacearum; Sm= Sinorhizobium medicae). Figure S5. Figure S5. Detection of low expression of the Bp AvrRpt2 homolog (longer exposure of Fig. 6C). See figure legend to Fig. 6C (*= weak expression of the Bp homolog; Bp= Burkholderia pyrrocinia; Cf= Collimonas fungivorans; Ea= Erwinia amylovora). 10

Figure S6. Figure S6. Western blot corresponding to experiment in Fig. 9A showing MAPK activation (αptepy), accumulation of MEKK1-YFP and HA-tagged CFP or AvrRpt2 variants. Note the three phosphorylated MAPK bands recognized by the α-ptepy antibodies are assumed to be MPK6, MPK3 and MPK4/MPK11 on the basis of the identical sizes to the flg22- activated MAPKs. 11

Table S1. AvrRpt2 homologs used in the study. organism Pseudomonas syringae pv. tomato JL1065 Mesorhizobium huakuii 7653R Erwinia amylovora ATCC 49946 Acidovorax avenae subsp. avenae ATCC 19860 Burkholderia pyrrocinia Lyc2 Collimonas fungivorans Acidovorax citrulli tw6 Sinorhizobium medicae WSM1369 Ralstonia solanacearum CMR15 GenBank-ID / NCBI 1 Reference Sequence CAA79815.2 AID34449.1 CBJ45097.1 ADX44172.1 KFL50402.1 WP_041743564.1 NZ_JXDJ01000021.1 NZ_AQUS01000051.1 NC_017559.1 (REGION: 3241390..3241941) 1 The National Center for Biotechnology Information (http://www.ncbi.nlm.nih.gov/) Table S2. List of primers and qpcr probes used in the study. name sequence purpose AvrRpt2_5 caccatgaaaattgctccagttgcc cloning of AvrRpt2 AvrRpt2_3 gcggtagagcattgcgtgtgg cloning of AvrRpt2 / qpcr (SYBRgreen) AvrRpt2_qPCR_5 cttttcacgatccccgacaggg qpcr (SYBRgreen) ACD11-like_5 gtacatcgccaaggttaagga qpcr (probe-based) ACD11-like_3 tctaggtcaagaatgttgtgtaacg qpcr (probe-based) ACD11-like-probe Universal ProbeLibrary probe #31 (Roche; cat. no. 04687647001) qpcr (probe-based) BAP1_5 tcgtcaatctttcgattttgg qpcr (SYBRgreen) BAP1_3 gcctccacaaaccagtaacc qpcr (SYBRgreen) CAD5_5 gggagttatcaacaatccattaca qpcr (probe-based) CAD5_3 tgaagctccccgttatcact qpcr (probe-based) CAD5-probe Universal ProbeLibrary probe #82 (Roche; cat. no. 04689054001) qpcr (probe-based) MPK3_5 tcaagcttcttcgtcatcttga qpcr (probe-based) MPK3_3 tgatttgatgaagatcagtatccatta qpcr (probe-based) MPK3-probe Universal ProbeLibrary probe #31 (Roche; cat. no. 04687647001) qpcr (probe-based) MPK4_5 ggcgcttatggaattgtctg qpcr (probe-based) MPK4_3 ttgtcaaaagcattaccaatcttc qpcr (probe-based) MPK4-probe Universal ProbeLibrary probe #55 (Roche; cat. no. 04688520001) qpcr (probe-based) MPK6_5 ataagcgatgagccagagtgta qpcr (probe-based) MPK6_3 cgcggtagattagttccttca qpcr (probe-based) MPK6-probe Universal ProbeLibrary probe #82 (Roche; cat. no. 04689054001) qpcr (probe-based) MPK11_5 gcttctggcatcgtctgtg qpcr (probe-based) MPK11_3 ctcaaagttctcttagcgtcgat qpcr (probe-based) MPK11-probe FAM-tggaattcggagacgggtgagg-BHQ1 qpcr (probe-based) NHL10_5 acgccggacagtctagga qpcr (probe-based) NHL10_3 ccctaagcctgaacttgatctc qpcr (probe-based) 12

NHL10-probe Universal ProbeLibrary probe #55 (Roche; cat. no. 04688520001) qpcr (probe-based) PCS1_5 acatatcaccgaggtgtatttaagc qpcr (probe-based) PCS1_3 gccaccaataggtgaaaagtg qpcr (probe-based) PCS1-probe Universal ProbeLibrary probe #95 (Roche; cat. no. 04692128001) qpcr (probe-based) WRKY33_5 gggaaacccaaatccaaga qpcr (probe-based) WRKY33_3 gtttcccttcgtaggttgtga qpcr (probe-based) WRKY33-probe FAM-tgtggagagagcatcacacgacatg-BHQ1 qpcr (probe-based) WRKY53_5 cggaagtccgagaagtgaag qpcr (probe-based) WRKY53_3 tctgaccactttggtaacatcttt qpcr (probe-based) WRKY53-probe Universal ProbeLibrary probe #25 (Roche; cat. no. 04686993001) qpcr (probe-based) ZAT12_5 ctttgggaggacacatgagg qpcr (probe-based) ZAT12_3 caaagcgcgtgtaaccaac qpcr (probe-based) ZAT12-probe Universal ProbeLibrary probe #66 (Roche; cat. no. 04688651001) qpcr (probe-based) LBb1.3 ATTTTGCCGATTTCGGAAC sequencing of SALK T-DNA insertion lines LBa1 TGGTTCACGTAGTGGGCCATCG sequencing of SALK T-DNA insertion lines LB2 gcttcctattatatcttcccaaattaccaataca sequencing of SAIL T-DNA insertion lines LB3 tagcatctgaatttcataaccaatctcgatacac sequencing of SAIL T-DNA insertion lines NOI1_LP ttgttttgggattgaatgattg genotyping of SAIL_1173_C11 NOI1_RP ttgttttgatcggagttttgg genotyping of SAIL_1173_C11 NOI3_LP ggcttgagtattctggggaag genotyping of SALK_133277 NOI3_RP gaagataaaaagacgggacgg genotyping of SALK_133277 NOI4_LP atcgttttgatcatcgcaaac genotyping of SALK_042784C NOI4_RP ttttcggtttatccttgcctc genotyping of SALK_042784C NOI5_LP gtgacggttggaattgacaac genotyping of SALK_118446 NOI5_RP ctaattccaacaatggcaacg genotyping of SALK_118446 NOI6_LP aacaacaaccgcaatgagttc genotyping of SALK_024766C NOI6_RP attttaattgtgatgcagccg genotyping of SALK_024766C NOi7_LP ttttcattcggatcactgtcc genotyping of SALK_007305C NOI7_RP agctaatcaagctccaagcac genotyping of SALK_007305C NOI8_LP tcaaggtgacctagtagggattg genotyping of SALK_101426 NOI8_RP aatcaaacaaacaacgcgttc genotyping of SALK_101426 NOI9_LP caaaatctaaagccaacaagaaaac genotyping of SALK_150777C NOI9_RP aacccacctcctcctcttttc genotyping of SALK_150777C NOI10_LP ggtgaataaaaggattcacccc genotyping of SALK_080643C NOI10_RP tggcatcaactccaaaatacc genotyping of SALK_080643C NOI11a_LP ccagctaaccttagagacgcc genotyping of SALK_028339 NOI11a_RP aacttgagatgcaagaaagcg genotyping of SALK_028339 NOI13_LP tggtttgtttcaggtgacacc genotyping of SAIL_147_D02 NOI13_RP caaaagatgatggactcaggg genotyping of SAIL_147_D02 NOI14_LP agaacaaagccctagtttggc genotyping of SALK_110602 NOI14_RP tttccggaaatatttggttcc genotyping of SALK_110602 NOI1_qPCR_5 CAGAGAAACCAAAACTCCGAT qpcr (SYBRgreen) NOI1_qPCR_3 TTAACGGAAGCAAAACCATTTG qpcr (SYBRgreen) NOI3_qPCR_5 GGACGGAGTTCGACGAA qpcr (SYBRgreen) NOI3_qPCR_3 TTAAGCAAAAGTGAAACAGAGCCA qpcr (SYBRgreen) NOI4_qPCR_5 GACAGACACAAAGATGGATTTG qpcr (SYBRgreen) NOI4_qPCR_3 TCAAGAAAGACGACAATAAAACATAAA qpcr (SYBRgreen) NOI5_qPCR_5 GAACCTCCTAAAAACAACAACAA qpcr (SYBRgreen) NOI5_qPCR_3 TTAACGAAAACAAAGCCATTTCTTC qpcr (SYBRgreen) NOI6_qPCR_5 CAATGAGTTCTCTCACCATC qpcr (SYBRgreen) 13

NOI6_qPCR_3 TTAAGCTTTAACGCAACAGTTGAA qpcr (SYBRgreen) NOI7_qPCR_5 CATCATCAACCACATTCTCCT qpcr (SYBRgreen) NOI7_qPCR_3 TCAAGCTTTAACGCAGCAGTT qpcr (SYBRgreen) NOI8_qPCR_5 ATGAAGATCTCTACAAGATTCCTC qpcr (SYBRgreen) NOI8_qPCR_3 TCATGCGCATGGAACCAAAC qpcr (SYBRgreen) NOI9_qPCR_5 CCCACAACTTCTCTCAGTCAA qpcr (SYBRgreen) NOI9_qPCR_3 TCAAAGCACCCGTGTTGGC qpcr (SYBRgreen) NOI10_qPCR_5 GCTAATGAAGATGATCATTTCCCT qpcr (SYBRgreen) NOI10_qPCR_3 TTACATCTCCATAAGAATAACAAAGATATTAT qpcr (SYBRgreen) NOI11_qPCR_5 CTTCCCGCCCCATCAACA qpcr (SYBRgreen) NOI11_qPCR_3 TCAATATTTACTTCCCCTGCGG qpcr (SYBRgreen) NOI13_qPCR_5 GGTCGGAGCTCTAGCAAG qpcr (SYBRgreen) NOI13_qPCR_3 TCAAGAAAAAGTAAAACAGAGCCATTT qpcr (SYBRgreen) NOI14_qPCR_5 CACCATCGCCATGAAACTCAA qpcr (SYBRgreen) NOI14_qPCR_3 CTAGTTTGGCCTTATACAACAATTAA qpcr (SYBRgreen) RIN4_FL_5 atggcacgttcgaatgtacc RT-PCR RIN4 full length RIN4_FL_3 tcattttcctccaaagccaaag RT-PCR RIN4 full length RIN4-RCS1/2_5 atgaactgggaagctgaggag cloning and RT-PCR of RIN4 ACP2 RIN4-RCS1/2_3 tcaaccgaatttaggcaccac cloning and RT-PCR of RIN4 ACP2 RIN4-RCS2-end_5 atggactgggacgagaacaac cloning and RT-PCR of RIN4 ACP3 RIN4-RCS2-end_3 tcattttcctccaaagccaaag cloning and RT-PCR of RIN4 ACP3 18S_5 gatggtaggatagtggccta RT-PCR 18S rrna 18S_3 tggttgagactaggacgata RT-PCR 18S rrna AvrRpt2_Y191C_5 aaaaaagaagactgtgtaagcacgggccgattatatttgg site-directed mutagenesis of AvrRpt2 (Y191C) AvrRpt2_Y191C_3 aaaaaagaagaccttacacaacagtgcacccaactcttcgtgtg site-directed mutagenesis of AvrRpt2 (Y191C) AvrRpt2_D216E_5 aaaaaagaagactcgagaaagagacgtcgtccattacttttcac site-directed mutagenesis of AvrRpt2 (D216E) AvrRpt2_D216E_3 aaaaaagaagactttctcgacaccagtgaggaccgacat site-directed mutagenesis of AvrRpt2 (D216E) MKK1_T218D S224D_5 aaaaaaggtctcttgctaatgatttcgtgggcacata site-directed mutagenesis of MKK1 (T218D S224D) MKK1_T218D S224D_3 aaaaaaggtctctagcaagactactatcgcttgtca site-directed mutagenesis of MKK1 (T218D S224D) MKK2_T229E T235E_5 aaaaaaggtctcagcaaacgaatttgtggggact site-directed mutagenesis of MKK2 (T229E T235E) MKK2_T229E T235E_3 aaaaaaggtctctttgctaaacctgcttcgtttgtc site-directed mutagenesis of MKK2 (T229E T235E) StNOX fwd tcaatgcataggtatgaaggaatc qpcr (probe-based) StNOX rev tctcttcctagctagagcatcaaat qpcr (probe-based) StNOX probe FAM-tgaaggatggtctgaggttgaatcgcg-BHQ1 qpcr (probe-based) BcCutA-fwd ataagcgccgagcatgtg qpcr (SYBRgreen) BcCutA-rev gggatgacggaaaatagacg qpcr (SYBRgreen) Labeling: cacc= 5 -extension for directional cloning into pentr /D-TOPO vector (Thermo Scientific) GAAGAC/GGTCTC= type IIs restriction enzyme recognition site (mutagenesis was performed as described (Eschen-Lippold et al., 2014; Palm-Forster et al., 2012) FAM= 6-carboxyfluorescein BHQ1= black hole quencher 1 14

Table S3. List of vectors and DNA constructs used for transient expression in this study. DNA construct structure reference pugw14-[gw] p35s::[gw] C-term. HA-tag Nakagawa et al., 2007 pugw15-[gw] p35s::n-term. HA-tag [GW] Nakagawa et al., 2007 pugw18-[gw] p35s::n-term. c-myc-tag [GW] Nakagawa et al., 2007 pugw15-cfp p35s::ha-cfp this study pugw14-avrrpt2 p35s::avrrpt2-ha this study phbt-avrrpt2 H208A-HA p35s::avrrpt2 H208A-HA Cui et al., 2013 phbt-avrrpt2 C122A-FLAG p35s::avrrpt2 C122A-FLAG Cui et al., 2013 pugw14-avrrpt2 Y191C p35s::avrrpt2 Y191C-HA this study pugw14-avrrpt2 D216E p35s::avrrpt2 D216E-HA this study pugw18-rin4 p35s::c-myc-rin4 this study pugw15-rin4 F9A p35s::ha-rin4 F9A this study pugw15-rin4 F151A p35s::ha-rin4 F151A this study pugw15-rin4 F9A F151A p35s::ha-rin4 F9A F151A this study pugw15-rin4 ACP2 p35s::ha-rin4 ACP2 this study pugw15-rin4 ACP3 p35s::ha-rin4 ACP3 this study pugw18-noi1 p35s::c-myc-noi1 this study pugw18-noi2 p35s::c-myc-noi2 this study pugw18-noi3 p35s::c-myc-noi3 this study pugw18-noi4 p35s::c-myc-noi4 this study pugw18-noi5 p35s::c-myc-noi5 this study pugw18-noi6 p35s::c-myc-noi6 this study pugw18-noi7 p35s::c-myc-noi7 this study pugw18-noi8 p35s::c-myc-noi8 this study pugw18-noi9 p35s::c-myc-noi9 this study pugw18-noi10 p35s::c-myc-noi10 this study pugw18-noi11 p35s::c-myc-noi11 this study pugw18-noi12 p35s::c-myc-noi12 this study pugw18-noi13 p35s::c-myc-noi13 this study pugw18-noi14 p35s::c-myc-noi14 this study pugw14-aa avrrpt2 p35s::aa avrrpt2-ha this study pugw14-ac avrrpt2 p35s::ac avrrpt2-ha this study pugw14-bp avrrpt2 p35s::bp avrrpt2-ha this study pugw14-cf avrrpt2 p35s::cf avrrpt2-ha this study pugw14-ea avrrpt2 p35s::ea avrrpt2-ha this study pugw14-mh avrrpt2 p35s::mh avrrpt2-ha this study pugw14-rs avrrpt2 p35s::rs avrrpt2-ha this study pugw14-sm avrrpt2 p35s::sm avrrpt2-ha this study phbt-axr2-ha p35s::axr2-ha Cui et al., 2013 phbt-axr3-ha p35s::axr3-ha Cui et al., 2013 pexsg-yfp-mekk1 p35s::mekk1-yfp this study pugw14-mkk1 p35s::mkk1-ha this study pugw14-mkk1 T218D S224D p35s::mkk1 T218D S224D-HA this study pugw14-mkk2 p35s::mkk2-ha this study pugw14-mkk2 T229E T235E p35s::mkk2 T229E T235E-HA this study prt100myc-pcmkk5 p35s::c-myc-pcmkk5 Lee et al., 2004 Abbreviations: [GW]= Gateway Cloning cassette; Aa= Acidovorax avenae subsp. avenae ATCC 19860; Ac= Acidovorax citrulli strain tw6; Bp= Burkholderia pyrrocinia Lyc2; Cf= Collimonas fungivorans; Ea= Erwinia amylovora ATCC 49946; Mh= Mesorhizobium huakuii 7653R; Rs= Ralstonia solanacearum CMR15; Sm= Sinorhizobium medicae WSM1369; Pc= Petroselinum crispum 15

Supplemental Experimental Procedures Plant lines and treatment The plant lines used were rps2 (rps2-101c; Bent et al., 1994; Mindrinos et al., 1994), rps2rin4 (Mackey et al., 2003), rpm1rps2 (Mindrinos et al., 1994), rpm1rps2rin4 (Belkhadir et al., 2004), DEX-T7-RIN4 and DEX-T7-RIN4 CCC>AAA (dexamethasone-inducible expression of T7- tagged RIN4 and RIN4 C203A/C204A/C205A in Col-0 background, respectively; Afzal et al., 2011), DEX-avrRpt2 (dexamethasone-inducible expression of AvrRpt2 in rps2-101c background; Mackey et al., 2003), 35S::axr2-1/DEX::avrRpt2/rps2 (over-expression of axr2-1 and dexamethasone-inducible expression of AvrRpt2 in the rps2 knock out background ; lines #20 and #23; Cui et al., 2013). To identify noi-gene knock out plants, the following T-DNA insertion lines were obtained from Nottingham Arabidopsis Stock Centre (Sessions et al., 2002; Alonso et al., 2003): noi1 - At5g63270 - SAIL_1173_C11; noi4 - At5g55850.2 - SALK_042784C; noi5 - At3g48450 - SALK_118446; noi7 - At5g09960 - SALK_007305C; noi8 - At5g18310.2 - SALK_101426; noi9 - At5g48500 - SALK_150777C; noi10 - At5g48657 - SALK_080643C; noi11 - At3g07195 - SALK_028339; noi13 - At4g35655 - SAIL_147_D02; noi14 - At5g19473 - SALK_110602. DEX-avrRpt2 plants were sprayed with 20 µm dexamethasone to induce AvrRpt2 expression. Five to six hours later, leaves were infiltrated with 1 µm flg22 to induce MAPK activation. Protoplast work For Western-blot analyses, 300 µl protoplast samples were each treated with 100 nm flg22 (or an equal volume of water) and harvested by centrifugation at the indicated time points. Media were discarded and cell pellets were snap-frozen in liquid nitrogen. To induce expression of RIN4 and RIN4 CCC>AAA in protoplasts derived from the respective DEX-inducible lines, 16

protoplasts were incubated after transfection in the presence of 10 µm dexamethasone overnight. Protein work Protein extraction from plant material was performed as described (Lee et al., 2004). In case of protoplasts, standard SDS loading dye was directly added to the pelleted samples. The antibodies used for immunoblotting were: α-ptepy (phospho-p44/42 MAPK [ERK1/2] [Thr202/Tyr204]; Cell Signaling Technology), α-mpk4 (Anti-AtMPK4; Sigma), α-ha (α-ha.11; Eurogentec), α-flag (FLAG Epitope Tag [DYKDDDDK] Antibody [FG4R]; Thermo Scientific), or α-rin4 (Mackey et al., 2002). Quantitative realtime PCR and semi-quantitative RT-PCR Genomic DNA was isolated with the GenElute Plant Genomic DNA Miniprep Kit (Sigma). RNA was isolated with standard Trizol reagent, treated with DNAseI and cdna synthesis was performed with the RevertAid kit (Thermo Scientific). For quantitative realtime PCR, amplicons from either genomic DNA samples (in case of Botrytis cinerea biomass quantification) or cdna samples (for expression analyses) as templates were analyzed with an Mx3005P qpcr system (Agilent) using the Maxima Probe qpcr and Maxima SYBR Green qpcr Master Mixes (Thermo Scientific). For the Botrytis cinerea experiments, an external control (plasmid clone of a potato StNOX gene) was spiked into the collected leaf material prior to genomic DNA extraction. For fungal biomass quantification, primers for Botrytis cinerea Cutinase A (accession number Z69264) gene were used. BcCutA data were normalized to corresponding StNOX data. Relative gene expression values of the genes-of-interest were calculated with the comparative C T method (Schmittgen and Livak, 2008) using the reference gene PP2A (AT1G13320; Czechowski et al. 2005). Primers and hydrolysis probes are listed in Table S2. 17

Semi-quantitative RT-PCR was performed using DreamTaq DNA Polymerase (Thermo Scientific) with the primers listed in Table S2. Callose detection Callose staining was performed with aniline blue (Adam & Somerville; 1996). Photographs were taken using a Nikon multizoom AZ100 microscope (Nikon Instruments) with the following UV filter combination: 370 36 nm excitation filter, 400 nm (LP) dichromatic mirror and 405 nm (LP) barrier filter. Reactive oxygen species measurement Production of reactive oxygen species was measured according to (Gomez-Gomez et al., 1999) using 3 mm leaf discs in a 96-well plate format measured in 2 min intervals (60 min total time) with a Luminoskan Ascent 2.1 device (Thermo Scientific). Phylogenetic analysis Phylogenetic analysis was performed with MEGA4 software (Tamura et al., 2007) using the Neighbor-Joining method (Saitou and Nei, 1987). The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches (Felsenstein, 1985). The trees are drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Poisson correction method (Zuckerkandl and Pauling, 1965) and are in the units of the number of amino acid substitutions per site. The optimal trees with the sum of branch lengths = 7.1187 (Fig. 3A) and 2.9233 (Fig. 6A) are shown. All positions containing gaps and missing data were eliminated from the dataset 18

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