Supporting InformationCheng et al. 10.1073/pnas.1105685108

0

1

2

RPS

2/A

CT1

0

1

2

RPM

1/A

CT1

0

1

2

RPS

4/A

CT1

0

1

2

3

RPS

5/A

CT1

0

1

2

RPP

4/A

CT1

WT

cul1-70

1

2

RPP

2A/A

CT1

0

1

2

3

RPP

2B/A

CT1

A B

FE

D

G

C

WT

cul1-7

WT

cul1-7WT

cul1-7WT

cul1-7

WT

cul1-7WT

cul1-7

Fig. S1. qRT-PCR analysis of selected R gene transcript levels in WT and cul1-7. (A) RPP4 levels in the indicated genotypes. (B) RPS4 levels in the indicatedgenotypes. (C) RPP2A levels in the indicated genotypes. (D) RPP2B levels in the indicated genotypes. (E) RPS2 levels in the indicated genotypes. (F) RPM1 levelsin the indicated genotypes. (G) RPS5 levels in the indicated genotypes. Plants were grown on MS plates and RNA samples were prepared from 2-wk-oldseedlings.

WT

cpr3

0-1

(cpr1

-2)

cpr3

0-2

(cpr1

-3)cp

r1

(cpr1

-1) sn

c1

Fig. S2. Morphology of WT, cpr1 (later renamed to cpr1-1), cpr30-1 (renamed to cpr1-2), cpr30-2 (renamed to cpr1-3), and snc1.

Cheng et al. www.pnas.org/cgi/content/short/1105685108 1 of 6

0

1

2

RPP

4/A

CT1

c pr1 p

ad4cpr1WT

0

1

2

RPP

2A/A

CT1

c pr1 p

ad4cpr1WT

cpr1 p

ad4cpr1WT

0

1

2

RPP

2B/A

CT1

0

1

2

RPS

4/A

CT1

c pr1 p

ad4cpr1WT

0

1

2

RPS

5/A

CT1

c pr1 p

ad4cpr1WT

0

1

2

RPM

1/A

CT1

c pr1 p

ad4cpr1WT

0

1

2

RP

S2/

AC

T1

c pr1 p

ad4cpr1WT

G H I

D FC E

0

1

2

SNC

1/A

CT1

c pr1 p

ad4cpr1WT

150sn

c1-r1

WT

cpr1

cpr1

pad4

cpr3

0cp

r30 p

ad4

snc1

snc1

pad4

-SNC1

Rubisco

A B

Fig. S3. Analysis of the transcript levels of SNC1 and selected R genes and protein levels of SNC1 in WT, cpr1, and cpr1 pad4 double mutant. (A) qRT-PCRanalysis of SNC1 in WT, cpr1, and cpr1 pad4. (B) Western blot analysis of SNC1 levels in snc1-r1 (a loss-of-function deletion allele of SNC1), WT, cpr1, cpr1 pad4,cpr30, cpr30 pad4, snc1, and snc1 pad4. Rubisco levels were used as loading control. (C–I) qRT-PCR analysis of RPP4 (C), RPS4 (D), RPP2A (E), RPP2B (F), RPS2 (G),RPM1 (H), and RPS5 (I) transcript levels in WT, cpr1, and cpr1 pad4 double mutant.

Cheng et al. www.pnas.org/cgi/content/short/1105685108 2 of 6

Fig. S4. Sequence analysis of cpr1. (A) Gene structure of At4g12560. Gray boxes indicate untranslated regions, black boxes are exons, and black lines areeither intergenic region or introns. Uppercase letters are exons and lowercase letters are introns. Letters with underline are predicted splicing acceptor sites. (B)Protein alignment of wild-type CPR1 and cpr1-1. The asterisks at the end of the amino acid sequences indicate stop codons.

Cheng et al. www.pnas.org/cgi/content/short/1105685108 3 of 6

ME04283G00010 (Manihot esculenta)RC29206G00020 (Ricinus communis)PT16G01200 (Populus trichocarpa)

CP00075G01030 (Carica papaya)AL6G35820 (Arabidopsis lyrata)CPR1 AT4G12560

MT2G14520 (Medicago truncatula)ABO92991 (Solanum tuberosum)

BAJ11960 (Malus domestica )LJ6G007300 (Lotus japonicas)

BAJ24863 (Petunia axillaris)ABR18790 (Nicotiana alata)GM15G10860 (Glycine max)

EEE69828 (Oryza sativa)XP002267358 (Vitis vinifera)

ADD09566 (Trifolium repens)ACU01763 (Brachypodium distachyon)

0.5

Fig. S5. Molecular phylogenetic anaylsis by Maximum-Likelihood method. A BLAST was performed on National Center for Biotechnology Information andPLAZA (http://bioinformatics.psb.ugent.be/plaza/blast/index) using CPR1 whole protein sequence as query. Homologs were found in 17 species and sequenceswere retrieved of the homologs with the highest similarity with CPR1 in each species. The evolutionary history was inferred by using the Maximum-Likelihoodmethod based on the Jones et al. w/freq. model (1). The tree with the highest log likelihood (−7748.4484) is shown. A discrete γ-distribution was used to modelevolutionary rate differences among sites [five categories (+G, parameter = 4.6404)]. The tree is drawn to scale with branch lengths measured in the number ofsubstitutions per site. Evolutionary analyses were conducted in MEGA5 (2).

1. Jones DT, Taylor WR, Thornton JM (1992) The rapid generation of mutation data matrices from protein sequences. Comput Appl Biosci 8:275e282.2. Tamura K, et al. (2011) MEGA5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol, in press.

0

10

20

30

40

CPR

1/A

CT1

WT

snc1

C

0

1

2

PR1/

AC

T1

WT

snc1

600

500

700

0

1

2

PR2/

AC

T1 60

80

100

WT

snc1

D

A

T1-5 T1-6

T1-5 T1-6 T1-5 T1-6

0

1

2

WT

snc1

T1-5T1-6

SNC

1/A

CT1

B

Fig. S6. qRT-PCR analysis of transcript levels of CPR1 (A), SNC1 (B), PR1 (C), and PR2 (D) in WT, snc1, and two independent CPR1 overexpression lines in snc1mutant background (T1-5 and T1-6).

Cheng et al. www.pnas.org/cgi/content/short/1105685108 4 of 6

0

2

4

6

8

10

T1-4T1-5W

T

CPR

1/A

CT1

WT eds1-2H.a. Cala2

HR

O

TN

35S::CPR1 in ColT1-4 T1-5

TN

fH

OHR

WT eds1-2H.a. Emwa1

TN

35S::CPR1 in ColT1-4 T1-5

TN

E F

0123456789

T1-4T1-5

eds1

-2WT

P.s.t. avrRps4Day 0Day 3

log(

cfu/

cm2 )

0

1

2

3

4

5

6

7P.s.t. avrPphB

T1-4T1-5

ndr1-1W

T

log(

cfu/

cm2 )

Day 0Day 3

C DA

012345678 P.s.t. avrRpm1

T1-4T1-5

ndr1-1W

T

log(

cfu/

cm2 )

Day 0Day 3

B

Fig. S7. Analysis of resistance mediated by RPM1, RPS5, RPS4, RPP2, and RPP4 in two independent transgenic lines (T1-4 and T1-5) overexpressing CPR1 in wild-type Col background. (A) qRT-PCR analysis of the transcript levels of CPR1 in WT, T1-4 and T1-5. (B–D) Growth of Pseudomonas syringae pv. tomato DC3000carrying avrRpm1 (B), avrPphB (C), and avrRps4 (D) in T1-4 and T1-5. WT, eds1-2, and ndr1-1 plants were used as controls. (E and F) Growth of Hyaloper-onospora arabidopsidis Cala2 (E) and H. arabidopsidis Emwa1 (F) on T1-4 and T1-5 plants. WT and eds1-2 plants were used as controls. fH, free hyphae; HR,hypersensitive response; O, oospores; and TN, trailing necrosis.

Cheng et al. www.pnas.org/cgi/content/short/1105685108 5 of 6

0

1

2

SNC

1/A

CT1

T1-4T1-5W

T

0

1

2

RPS

2/A

CT1

T1-4T1-5W

T0

1

2

T1-4T1-5W

T

RPM

1/A

CT1

B

G H

0

1

2

WT

T1-4T1-5

RPS

4/AC

T1

0

1

2

RPS

5/AC

T1

WT

T1-4T1-5

0

1

2

RPP

4/AC

T1

WT

T1-4T1-5

0

1

2

RPP

2A/A

CT1

WT

T1-4T1-5

0

1

2

RPP

2B/A

CT1

WT

T1-4T1-5

I

D

C

E F

0

2

4

6

8

10

CPR

1/A

CT1

T1-4T1-5W

T

A

Fig. S8. qRT-PCR analysis of transcript levels of CPR1 (A), SNC1 (B), RPP4 (C), RPS4 (D), RPP2A (E), RPP2B (F), RPS2 (G), RPM1 (H), and RPS5 (I) in WT and twoindependent CPR1 overexpression lines in wild-type Col background (T1-4 and T1-5).

130

170

kDaRPS4-H

A in cp

r1-3

RPS4-HA

cpr1-

3

Rubisco

170

13055

43

kDa

-SNC1

-HA-HA

-SNC1

-FLAG

Rubisco

RPS4-HA

T1-2 T1-3

pCPR1::CPR1-FLAG in RPS4-HA

A B

Fig. S9. RPS4-HA and SNC1 levels in two independent RPS4-HA lines (T1-2 and T1-3) transformed with CPR1-FLAG (A) and in cpr1-3 with RPS4-HA transgene (1)crossed into it (B). Total proteins were extracted from the indicated genotypes and probed with the specific antibodies to detect levels of CPR1-FLAG, RPS4-HA,and SNC1. Rubisco bands were used as loading controls.

1. Wirthmueller L, Zhang Y, Jones JDG, Parker JE (2007) Nuclear accumulation of the Arabidopsis immune receptor RPS4 is necessary for triggering EDS1-dependent defense. Curr Biol 17:2023e2029.

Other Supporting Information Files

Table S1 (XLS)

Cheng et al. www.pnas.org/cgi/content/short/1105685108 6 of 6