Nature - 15.08.2019

Article reSeArcH

WT moca1

0 mM NaCl

60 mM^0620 [NaCl] (mM)^400

WT

moca1

0

10

K content (mg g

–1 DW)

5

0620 40 0

WT

moca1

[NaCl] (mM)

0

2

4

Na content (mg g

–1 DW)

6

WT

moca1

1

2

–NaCl +NaCl

[Ca2+]i (Cameleon)

0.2

[Ca

2+

] (Fi

535

/F

485

)

20 s

WT

moca1

d

080

[NaCl] (mM)

20 6040

WT

moca1

0

We

ight (mg FW

per seedling)

6

4

2

Na

+/H

+ exchange

(^6 %

F min

–1 mg

–1)

[NaCl] (mM)

06080

NS **** WT

***

50

100

Survival rate (%)

0

c

a

g h

b

f

e

NaCl

NS ***

moca1

40

30

20

0

10

WT moca1

–NaCl

+NaCl

***

NS

Fig. 2 | The moca1 mutant is defective in the SOS pathway and
hypersensitive to salt stress. a, b, Increases in [Ca^2 +]i induced by NaCl
in roots. YC3.6 emission images were taken every 3 s, and 200 mM NaCl
was added at the time indicated (a). Emission ratios are shown using a
pseudo-colour scale and quantified from experiments similar to these in
a (b; mean ± s.d.; n = 10 seedlings). Similar results were seen in more
than ten independent experiments. c, Plants were grown on half-strength
Murashige and Skoog (½ MS) medium containing 0.2 mM CaCl 2 with or
without 60 mM NaCl for 12 days. Similar results were seen in more than

ten independent experiments. d, e, Fresh weight (FW; d) and survival

rate (e) from experiments similar to those in c were quantified. Data are

from five independent experiments (mean ± s.d.; n = 12 pools (8–12

seedlings per pool); two-way ANOVA, P < 0.001; NS, not significant;

***P < 0.001). f, Na+/H+ exchange activity from plants treated with

water or 100 mM NaCl for 24 h (mean ± s.d.; n = 3; **P < 0.01).

g, h, The content of Na (g) a nd K (h) of plants from experiments similar to

those in c. Data are presented as mean ± s.e.m. (n = 6; two-way ANOVA,

P < 0.001).

abc

d e f

i

150

300

0

ROI (

μm)

036 72 108

Low High

150

300

0

moca1

01030 60 150

WT

Time after NaCl treatment (s)

Low [Ca2+]i High

NaCl

0 0.2 0.4

6 [Ca2+]i (F 535 /F 485 )

WT

mo

ca1

0.1 0.3

4

1

0

4210 16 2

3

2

WT

Temperature (°C)

moca1

0 200

[LiCl] (mM)

50 100 150

WT

1.2

0.8

0.4

1.6

0

moca1

0

1.2

0.8

0.4

1.6 WT

moca1

[H 2 O 2 ] (mM)

01510 5

2

1

0

0 200 600

WT

400

3

[CaCl 2 ] (mM)

moca1

0 200

[NaNO 3 ] (mM)

50 100 150

WT

1.2

0.8

0.4

1.6

0

moca1

0 200

[KCl] (mM)

50 100 150

WT

0

1.2

0.8

0.4

1.6

moca1

gh

[Ca

2+

] (i

μM)

[Ca

2+

] (i

μM)

[Ca

2+

] (i

μM)

[Ca

2+

] (i

μM)

[Ca

2+

] (i

μM)

[Ca

2+

] (i

μM)

[Ca2+]i

Time after NaCl treatment (s)

ROI (

μm)

Fig. 3 | The moca1 mutant abolishes [Ca^2 +]i spikes and waves induced
by monovalent cations. a−c, Increases in [Ca^2 +]i in seedlings plotted as a
function of applied [H 2 O 2 ] (a), temperature gradient (b), and [CaCl 2 ] (c).
Data are from three independent experiments similar to those in Fig. 1a–c
(mean ± s.d.; n = 32 seedlings; two-way ANOVA; a, P = 0.64; b, P = 0.06;
c, P = 0.11). d−f, Increases in [Ca^2 +]i plotted as a function of applied
[KCl] (d), [LiCl] (e), and [NaNO 3 ] (f). Data are from three independent
experiments (mean ± s.d.; n = 32 seedlings; two-way ANOVA, P < 0.001).
g−i, Wave-like propagation of [Ca^2 +]i through the root 1,000 μm from

the site at which root-tip was treated with 200 mM NaCl was analysed via

YC3.6 imaging. Images in regions of interest (ROIs) at indicated times

are shown (g; scale bar, 200 μm). The wave dynamics of [Ca^2 +]i were

examined shootward from roots similar to the ROIs in g by repeatedly

analysing images taken every 2 s, and data from five roots were averaged

and pseudo-colour coded (h). Similar results were seen in more than 20

independent experiments. Changes in peaks of [Ca^2 +]i waves in ROIs were

quantified from experiments similar to those in g (i; mean ± s.d.; n =  21

roots; Student’s t-test, P < 0.001).

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