Figure S1: Adult phenotype of nma-1 (Ler-0 back-ground) and nma-3 (Col-0 background) in comparison to corresponding wild-type plants. Scale bar = 5 cm

Ler-0 nma-3Col-0nma-1

Figure S1

[number of cells]30

20

10

25

15

5

nma-1n=15

Ler-0n=15

ssp-2n=15

Col-0n=15

* *

Figure S2

Figure S2: Number of cells in a central optical sectionof the proembryo 3 days after pollination.Significant differences as indicated by Mann Whitney U-test (p<0.001) are highlighted by an asterisk. Boxes represent lower and upper quartiles around median, whiskers indicate maximum and minimum values.The number of examined embryos is given below each genotype.

pollen tube length [µm]

nma-1n=189

Ler-0n=146

350

300

250

200

150

100

50

Figure S3:

Figure S3: Pollen tube length of in vitro germinated pollen 6h after inocculation.Boxes represent lower and upper quartiles around median, mean values are shown as triangles, whiskers indicate maximum and minimum values.The total number of analyzed pollen tubes is givenbelow the genotype.

suspensor length [µm]

100

WT x WT WT x nma nma x WT nma x nma

20

40

60

80

(n=107) (n=92) (n=94) (n=38)

Figure S4:

Figure S4: Measurements of suspensor length after reciprocal crosses of wild-type (Ler-0) and nma-1 mutant plants. Number of measured suspensors is given in brackets. Significant differences to wild-type cross (WT x WT) as indicated by Mann Whitney U-test (p<0.001) are highlighted by an asterisk. Boxes represent lower and upper quartiles around median. Mean values are shown as triangles.

* *

0

20

10

[recombinants]

13.7 13.8 14.414.314.214.114.013.9position [Mb]

Figure S5:

Figure S5: Fine mapping of nma-1. Number of recombinants in a 563 kb genomic interval around nma-1 on chromosome II are shown. Dashed lines indicate an intervall without further recombinations.

1kb nma-3(SALK_015991)

nma-2(SALK_126968)

nma-1(G>A)

A

signal peptidenma-1

(G279>D)

cleavage site

glycoside hydrolase, family 28

N C

100 aa

B

Figure S6:

Figure S6: Graphic representation of the NMA gene and protein.A, Gene model of the genomic NMA locus. Exons are depicted as white boxes, 3’UTR as black box, intergenic regions as line. The position of nma alleles are shown below the model. B, protein model of NMA. The predicted signal peptide is shown in black, the predicted polygalacturonase domain in grey. The position of the nma-1 substitution is shown on top of the model.

A CB

Figure S7:

Figure S7: Embryonic phenotype of T-DNA insertion alleles of nma.A, nma-2; B, nma-3; C, Col-0 wild-type. Scale bar = 20µm

20

120

100

80

60

40

[µm]Le

r-0

nma-

1

pNM

A::N

MA

pNM

A::Y

FP-N

MA

pSU

C5:

:YFP

-NM

A

pJ10

::YFP

-NM

A

pOB

P1:

:YFP

-NM

A

suspensor length

140

Figure S8:

Figure S8: Complementation assay based on suspensor length.The genotype of representative transgenic lines is given below chart. Boxes represent lower and upper quartiles around median. Whiskers depict maximum and minimum values.

normalized expression level

Figure S9:

RL SIPOINCL

Figure S9: quantitative RT-PCR data of NMA expression in various tissue types. RL, rosette leaves; CL, cauline leaves; IN, inflorescences; PO, pollen; SI, siliques; SH, seedling shoot including cotyledons and first leaves; RT, seedling root.Values are normalized to average expression levels in pollen. Mean values of three biological replicates with standard deviations are shown.

RTSH0

12

10

8

6

4

2

14

16

Figure S10:

Figure S10: Graphic representation of NMA expression data in publicly available microarray data.Screen shot of the Arabidopsis eFP browser (Winter et al. 2007).

Figure S11:

Figure S11: Expression of promoter::reporter gene fusions. A, pOBP1::n3xGFP; B, pJ10::n3xGFP. Scale bar = 10µm.

A B

Figure S12:

Figure S12: Average developmental stage of embryos 4 dap in pJ10::YFP-NMA, nma-1, and Ler-0. Embryos were classified in 6 developmental stages. Numerical values were asigned to each stage. Averages and standard deviations of at least three biological replicates are shown as bar graphs. Asterisks indicate significant differences in pair-wise comparison (t-test; p<0.05).

pJ10::YFP-NMA Ler-0nma-1

Developmental stage

n=500n=374 n=1102

mg

lg

tr

eh

lh

et

1

6

5

4

3

2

*

Table S1: Primer sequences

Primer name Primer sequence

160-3, R ctctcttcttcttcctcgcc

At2g33160P-FC cggatctgttgttttttctcag

At2g33160P-RC tctctcttctttggtttggt

At2g33160-RC aaggtagaggacgatgtgattc

LBb1.3 attttgccgatttcggaac

NMA dC-F aacaaactttgaatgagctgtaat

NMA dC-R tgcgatacggacttagcccatgtcttgatatttaaa

NMAi1 AP-F accaaaccaaagaagagagaggcgcgccaaaaattaattaaagcgtctagggttcttctgat

NMAi1 AP-R atcagaagaaccctagacgctttaattaatttttggcgcgcctctctcttctttggtttggt

NMAi1 ASP-F ccacgaaggaatatcccggcgcgccagtcgacttaattaaaatctttgacgtcagaaa

NMAi1 ASP-R tttctgacgtcaaagattttaattaagtcgactggcgcgccgggatattccttcgtgg

NMAi1-F ttggatatctctgatgtcagatgcggaccagggcatggta

NMAi1-R taccatgccctggtccgcatctgacatcagagatatccaa

qRT-F gcaaatctacaacgtcact

qRT-R gacactaataccatgccct

NRT-F3 gtcgtacactagctatgaa

NRT-R4 aatgacgatagggtttccaa

p35Sas aaaaaaggtaacccccgtgttctctccaaatg

p35Ss ttttttctcgagtcgacctgcaggcggccgca

pJ10as aaaaaaggttaccctttacaccgtctcttgctctc

pJ10s ttttttctcgaggtgccatgttgttctatattct

pOBP1as aaaaaaggttaccaatcggagaaaggttgaagctt

pOBP1s ttttttctcgagcatagatttggtgttgatgtgg

pSUC5as aaaaaaggttacctatgaaaagaaaaacgagcag

pSUC5s ttttttctcgagtgagtaagaacaggtagtcc

RT 160.2 F tttgtgtgagaacgttggtg

RT 160.2 R ggttttagtagcctcagta

RT-5F ttttgtgtgtgccttctctc

RT-5R aagtaatccaccgccagtaa

SSP1-I ttagagaccacacgagaaggc

SSP1-R taacatggcttggtctgatcc