Supplemental Materials

Arabidopsis nonhost resistance PSS1 gene encodes a novel glycine rich plasma membrane protein that enhances disease resistance in transgenic soybean plants.

Bing Wang1, Rishi Sumit1, Binod B. Sahu, Micheline N. Ngaki, Subodh K. Srivastava, Yang Yang, Sivakumar Swaminathan, Madan K. Bhattacharyya*

Department of Agronomy, Iowa State University, Ames 50011, U.S.A

1, B.W. and R.S. contributed equally.

*Correspondence to: mbhattac@iastate.edu

Supplemental Figure S1. Complementation analysis of the pss1 mutant with the CaMV 35S promoter fused PSS1 cDNA. Five pss1 RILs (Sumit et al. 2012), including pss1-43, pss1-230, pss1-463, pss1-S2#5 and pss1-S2#7, were individually transformed with the construct carrying WT Col-0 PSS1 cDNA driven by the CaMV 35S promoter. Leaves of Basta resistant F3 progenies were inoculated with P. sojae. Lines ending with C or C#5 and C#7 were transformed and complemented with PSS1.

Supplemental Figure S2. Structure comparison of PSS1 and its mutant pss1 protein. Protein structure was predicated using I-TASSER program and then structure alignment was conducted by TM-align program (TM-score = 0.26). Both programs were obtained from http://zhanglab.ccmb.med.umich.edu/. The protein in green color is PSS1. The protein in red color is the mutant pss1 protein, whose Gly residue at 119

th was substituted with Asp.

Supplemental Figure S3. Alignment of PSS1 homologs. Sequences of 10 PSS1 homologs and PSS1 clustered in a subclade were aligned using MUSCLE program. The red asterisk indicates the 119th Gly mutated to Asp in PSS1 gene in the pss1 mutant. The identified conserved glycine rich and transmembrane motifs, and nuclear localization signal, are denoted with black lines.

Supplemental Figure S4. Coexpression gene analysis based on microarray dataset.A, Co-expressed genes that show expression patterns similar to that of PSS1. Co-expression gene network associated with PSS1 (At3g59640) was retrieved from ATTED-II (http://atted.jp/) (Obayashi et al., 2007). PSS1 is highlighted in yellow. The thickness of lines connecting the circles corresponds to the relative strength of co-expression. The dark orange lines indicate conserved co-expression between Arabidopsis and at least one of the three mammalian species (human, mouse and rat). Nodes marked with yellow, red and green dots indicate genes related to the spliceosome (ath03040), autophagy (ath04140) pathway and vesicular transport process, respectively. Interactions among the proteins encoded by the genes of the network are unknown based on available TAIR (Rhee et al., 2003) and Intact (Kerrien et al., 2007) protein-protein interaction databases. B, Go ontology enrichment of Pss1 coexpression genes.

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N-terminal GFP-PSS1 PM-mCherry Merge

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Supplemental Figure S5. Subcellular localization of N- or C- terminal GFP tagged PSS1 A, Diffusive green fluorescence from N- terminal GFP fusion PSS1 cannot merge with red fluorescence of plasma marker PM-mCherry. B, Hechtian strands (arrows) are only present in salt plasmolyzed cells carrying C-terminal fusion PSS1-GFP. Bar = 50 µm. The localizations were conducted in Nicotiana benthamiana.