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1Supplementary materials for “Motility 2regulation and oxygen signaling of PAS 3domain-containing proteins in Xanthomonas 4campestris pv. campestris” 5by Daqing Maoa,b, Jun Taoc,d, Chunxia Lic,e, Chao Luoa, Linlin Zhenga, Chaozu 6Hea,f

7 8aSchool of Life Sciences, Tsinghua University, Beijing 100084, China 9bSchool of Life Sciences and Biological Pharmaceutics, Shenyang 10Pharmaceutical University, Shenyang, 110016, China 11cState Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese 12Academy of Sciences, Beijing 100101, China 13dCAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of 14Microbiology, Chinese Academy of Sciences, Beijing 100101, China 15eGraduate School of Chinese Academy of Sciences, Beijing 100039, China 16fHainan Key Laboratory for Sustainable Utilization of Tropical Bioresource, 17Hainan University, Haikou, Hainan 570228, China 18 19Corresponding author: 20Daqing Mao 21Wenhua Road 16#, Shenhe District, Shenyang, 110016, China 22Phone: +86-10-62446110 23Email : [email protected] 24Chaozu He 25Shuangqing Road 30#, Haidian District, Beijing 100084, China 26Phone: +86-10-64807395 27Email : [email protected] 28 29 30 31 32 33 34 35

1 1 36Table S1. Strains and plasmids used in this study Vector and Strain Description Origin or reference Vectors: pK18mobSacB Suicide vector for insertional mutagenesis, Kanr (Schäfer et al. 1994) pGEM T-easy T-vector for cloning PCR products Promega Co., USA pRK2013 Helper plasmid for mobilization of non-self-transmissible plasmids, Kanr (Figurski and Helinski 1979) Bacterial strains: E. coli DH10B F– mcrA Δ(mrr-hsdRMS-mcrBC) φ80dlacZ ΔM15 ΔlacX74 deoR recA1 endA1 araD139 Δ (ara, (Grant et al. 1990) leu)7697 galU galK λ– rpsL nupG E. coli S17-1 hsdR hsdM+ recA pro RP4-2 (Tc::Mu) (Km::Tn5), Spr (Simon et al. 1983) Xcc 8004 Wild type, RifR (Qian et al. 2005) In-frame deletion Mutants: DLT0728 Full length in-frame deletion mutant of XC_0728 This study DLT0818 Full length in-frame deletion mutant of XC_0818 This study DLT1036 Full length in-frame deletion mutant of XC_1036 This study DLT1756 Full length in-frame deletion mutant of XC_1756 This study DLT2324 Full length in-frame deletion mutant of XC_2324 This study DLT2579 Full length in-frame deletion mutant of XC_2579 This study DLT4313 Full length in-frame deletion mutant of XC_4313 This study Insertional mutant: DLT0637 pK18mobSacB derivative containing a 593bp fragment of XC_0637 This study DLT1965 pK18mobSacB derivative containing a 515bp fragment of XC_1965 This study S0 pK18mobSacB derivative containing a 677bp of intergenic region between XC_1972 and XC_1973 (Qian et al. 2008) 37All deletion mutants were in Xcc 8004 background with use of suicide construct (KanR) of pK18mobSacB derivative.

2 2 38Table S2. Primers used to amplify fragments for in-frame or insertional deletion of targeted 39genes in the genome of Xcc 8004

Targeted genes a Sequence of the primer pairs (5’-3’) Product (bp) In-frame deletion: U: GTCAAGCTTGCGCTGCACTTATGATATGCC 289 U: ACTGGATCCGCTATGCGACACCCACTCCG XC_0728 D: ACTGGATCCGAAGCGCCCAACCAAGAGT 259 D: AGCTCTAGAGAGCACTTCCAGCCCATCC D: GTCAAGCTTCGCCTGAATCTGGAAGACAA 315 D: ACTGGATCCGCGTTGCGAGGGATACATC XC_0818 U: ACTGGATCCCGCGTCATCCTGGTCACC 296 U: AGCTCTAGACAGCGTCGTCTTCATCATCG D: GTCAAGCTTACGGAAATTAAGCTGCTGTCATT 360 D: ACTGGATCCGCATACTGGAGTCGGTGAATACG XC_1036 U: ACTGGATCCGCTGAACCATCATGTCCTTGC 373 U: AGCTCTAGAGCCCTTGTACCCTTCTGCCTT U: GTCAAGCTTTGGCGAATACCAGTGGATAGTG 254 U: ACTGGATCCCTGATCTGTGCCCCACGTCT XC_1756 D: ACTGGATCCACGCTCCAGCTCGTTATGCC 268 D: AGCTCTAGACGCACGTGAATGCACTAGCC D: GTCAAGCTTTGAAGCAAACCGCCAAGAC 310 D: TTAGGTACCTATTCCAGCCTGAGCCACC XC_2324 U: TATGGTACCCGATTGCAGATCGCAGATC 267 U: AGCTCTAGAAGGAAGTGGTGCCCTTGG D: GTCAAGCTTATCAGCGACCTGCGACAA 304 D: ACTGGATCCTGGCACCGTGCGGAAAT XC_2579 U: ACTGGATCCCGGTGTTGTTCGTGACCG 326 U: AGCTCTAGATGTTGTACCCGAGCTTGCC U: GTCAAGCTTGCGTCTTCAGCGTTTCCTTG 284 U: TATGAATTCGCCGACAAGTCGTTGTACGC XC_4313 D: TATGAATTCGACCACCCGCAGCAACG 211 D: AGCTCTAGAGGATGCAGCCGCACATG Insertional deletion: AGTTCTAGATCCCGATCCCCGCCAAC XC_0637 506 GTCAAGCTTTCTGCCGCTGTGCCTGC AGTTCTAGATGTTGAGTTGCTGCCAGAGC XC_1965 460 GTCAAGCTTGTGGCTGCGTTCGTCTGTG S0 (intergenic AGCAAACCACAGGCAGGTG 677 region) CAAAGACGGCTGACGGTTG 40a: The codes for genes are according to that of the Xcc 8004 genome. U: upstream fragment; 41D: downstream fragment. 42 43

3 3 44Table S3. 16 known PAS proteins Uniprot Name Function Literature P16113 PYP in Halorhodospira Blue light (Kort et al. 1996; Baca et al. 1994; halophila Kyndt et al. 2007; Getzoff et al. 2003) P93025a Phototropin-2, PAS1 Blue light (Kagawa et al. 2009; Doi et al. 2004; P93025b Phototropin-2, PAS2 Blue light Kaiserli et al. 2009; Harper et al. 2003) O48963a Phototropin-1, PAS1 Blue light (Christie et al. 2007; Harper et al. O48963b Phototropin-1, PAS2 Blue light 2003; Christie et al. 1999) O07014 RsbP in Bacillus subtilis Red light (Brody et al. 2009) Q31NI4 SL2 in Synechococcus Blue light (Cao et al. 2010) elongatus Q8YC53 HK(Brucella melitensis) Blue light (Swartz et al. 2007) Q2NCA3 HK1 (Erythrobacter litoralis) Blue light (Swartz et al. 2007) Q5PXG9 White collar 1(WC1) in Blue light (Cheng et al. 2002; Ballario et al. Trichoderma reesei 1998; He and Liu 2005) Q5PXG8 White collar 2(WC2) in Blue light (Moriwaki et al. 2008; Cheng et al. Trichoderma reesei 2002; He and Liu 2005) P30663 NifL Oxygen/Redox (Key et al. 2007; Söderbäck et al. 1998; Hill et al. 1996) P23222 FixL Oxygen/Redox (Gong et al. 2000; Gong et al. 1998) Q02280 EAG in Sophophora Voltage (Wang et al. 2002; Warmke et al. 1991) P76129 DosP in Escherichia coli Oxygen/Redox (Liebl et al. 2003; Park et al. 2004; Tuckerman et al. 2009; Kurokawa et al. 2004; Delgado-Nixon et al. 2000) O34627 YtvA Blue light (Möglich and Moffat 2007) 45 46 47

4 4 48Table S4. Secondary structure topologies of 9 PAS domains in Xcc

49 50 A β B β C α D α E α F α G β H β I β 51 * 20 * 40 * 60 * 80 * 100 * 120 * 52140 53 XC 0818 : ------LEHLPQ GVVVID AAL NLVAW NSRYV-EL FRFPAEL---LQVGRP IED---LFRFNARR GLLGPGP IEA------AIERR LNHLRSGK-PH MRESE KEDGT VLEIRGN PLPDGG FVTSYA DIT------: 108 54 XC_1965a: ------YFN QRWY-EY TGLN PD----TE ASAAWQQ-----AHSAAG LRMS----GQAWAL----ALN GE-----RA--Y EIECELR RHDGQ----- TRWHLAR-ALPVR DEHG RISEWI GT------: 84 55 XC_1965b: ------VYWN PQWY-EF TGQG EE----QA LDQGWWN----AMHPDD AARVR---HAFEQAL------D AR---G--A-- FIAEYRIR RHDGQ----- YRWCIDT-ASPHF GSDG RFLGH IG------: 84 56 XC_2579a: ------FYN DAYT-PI LG-PRQR---R A LGAP LRAL--- W AD-AWDAVRG--PIEAAF ------A---GHA S--R---FEEMPIGMN RHGS---PE QTWWTFS-FSPI YLDDGR VGGAFCVTNEV TDRK--- : 94 57 XC_2579b: ----FEQAPM FMAFLT GPQH- RVEFV NDGYS-KL IG-HRD-----VIGKPLA E A---LPDAAE QGYLQ--PLDEVY------R SGRA YTA--N---ALAYTMQA EPDGA---V ERRLLDLV-YQPIT GSDGA ISGVFIQGLD ITDR---- : 114 58 XC_2579c: - RQILDS VMDY AIIAT DLQG- RVTSW NEGAR-RI LGWA EA----DML GQTL ER---TF TLEDIA RKQL---LVEMAS-----AL ETG-----S--- GTDERWHV RSSGE----- RFWANGS-MMVLR EETGA AIGFVKVLR DR------: 111 59 XC_2579d: ------GVLSDD----LPAN PRWQQ---EL VHPDD RAQL-----AQASAT----AIA NT-----AS-- LDVEYRVN LPDQQ----- TGWTQMR-AVPLL DAHGR IIEWFG TA------: 77 60 XC_2324 : ------LDGSSS AILIC DLQSR LLYA NDGFE-RM FGYTEA---- EL VGQRPSH----VLTRALS DQTEV---ARIRER--AD ALQQ------T QADLLVY RKDGT----- PLWISLN-FNPIY DAQHN PSHYVAVLTD ITETK--- : 111 61 XC_1756a: -- LAALVSHSS EA IVAE TPDG EITH WNPAAE-AMF GYP AD----VAI GQDLTT----LLSPQP YAMPQ----IDQDE-----APP VAA------L SAAQTMR HRDGH----- QVFVLAS-KAPII EHDNA VSGHSHFFR DI------: 109 62 XC_1756b: --- RAILA HAGY AIIAT DSSG FVTM FNPAAE-KL LGYS ADD----VI GRRKAS---QFIEAD ELQERALR-LAEQ TGTPVAG TLEAVAALTS LGRS------D TSEWTYVS QDGR----- RLPVLLT-LSTLR DDNDQ VIGYLGV AV : 123 63 XC_0637a: ------LEAMPD GVVAC GTDG LLRE FNHAAR-QWH GTDPR----VLPPAQWAQHFD-LYAADGSTVL P----TDAIP--LVRAW RGEH------VRN AELVIRA TGQ----P PRSVLCN-ADPID GDHGV ALGAVCVMHD ITQLK--- : 116 64 XC_0637b: ------DV AIIVF DPLG HIEV FNPGAE-RLL GYT AA----EVM GQCPTQ----FHLPEE LARHRAT--L AVPA- PSYVALAAAAA G--DLL-EQ EVWTLL RKDGS----- QRRVRLS-FNVIQ DAQQG LTGYLAMAI DVTAE---- : 115 65 XC_0637c: - RFSGAFDTA PQ GMAIV SLEGR WSEI NSALC-TL LGYSR T----QLL GMTF QD----VT HPDD LDVDLA-LLGELLD -----GKRNS------YTLSKRYV SRQGA----- VLWTLLS-VSLVR DSDGA PVHFVSQVL DV------: 112 66 XC_0637d: ------DATPT LVSQF DAAQ RCLF ANEAHR-SW LGVD PN----SLV GCHITR----L LGDDLGEPVYA---ALAAA ------ARG------ER ATFDYVL QAGAS---- ARDVEITLVPATK GTQA QVQGFFLMAQD VTAH---- : 107 67 XC_4313 : ------ICTH DMDG RLISV N PAAA-RS LGRS VE----DMEG RALVE----LVRPER HAALHG----YLSR-----MVL DG-----Q---DS GVIELV AGDGS---- LRHWQYHN--VLD VEADG T--VVLAHAQD VTNQY--- : 100 68 XC_0728 : ------REP IVLL DARQ RIVL HNPAFA-EL YGLE ER------GNE LML----LDDVGDGAWRD PQIHQRLAD----VLLR DREL------WD FEHEQR GADGV----- LRTMLINARRMPLP DDGE EEVVLMTVSD ISL----- : 107 69 XC_1036 : ------PPKA STLLRVLSA LPF GIAWA SFPDG K--IQYC NAEFVR----LFG HDASHFRT------VDQFVDSL YLHE RQ------REMVRKRWL DIELH------: 76 70 72 Pink colour region indicates beta sheet, green colour region indicates alpha helices. 73 74 75 76 77 78 79 80 81

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