Science - 27.03.2020

(Axel Boer) #1

SCIENCE 27 MARCH 2020•VOL 367 ISSUE 6485^1483


Fig. 2.SlPhyt2expression is induced by drought
stress.(A)SlPhyt2expression is induced in leaves of
drought-stressed (color-shaded bars) WT plants
(gray) and overexpressing plants (magenta) but not
in knockdown plants (blue); bars show mean expres-
sion levels ± SD as ln fold change relative to the
watered WT control (open bars). (B) Phytaspase
activity in cell wall extracts of leaves from drought-
stressed plants is reduced in knockdown and
increased in overexpressing plants as compared to
wild type. Means (A) and medians (B) with no letter in
common are significantly different (ttest). (C)
Histochemical staining ofSlPhyt2pro::GUS activity in
the proximal pedicel of developing tomato flowers;
numbers indicate developmental stages according to
( 30 ). Arrowheads mark the abscission zone. Early
abscission was typically observed between flower
stages 18 and 20. (D) GUS staining in leaves and
inflorescences of control and drought-stressed plants.
Scale bars in (C) and (D) represent 5 mm.

OE

6 12

17

19 20 F1 F3

C


D

KD1 -KD2 -KD3 -WT -OE1 -OE2 -OE3 -

0

10

20

30

40

SlPhyt2 activity (RFU/min)

A B


SlPhyt2

expression (ln fold change)

-2 - - -

0

2

4

B

D

A

C
C C

A

A
A

C

B
C
C



control drought

control drought

KD WT KD OEWT

Fig. 3. Abscission is regulated bySlPhyt2 and
PSK in an auxin- and ethylene-independent
manner.(A) Induction ofSlPhyt2expression
in the proximal (prox) compared to the distal
(dist) pedicel after flower removal analyzed
by quantitative polymerase chain reaction
(qPCR) before (open bars, control) and
14 hours after flower removal (gray bars)
normalized to abscission zone (AZ) control
(mean ± SD;Pvalues mark significant
differences to the abscission zone control;
ttest). (B) qPCR analysis ofSlPhyt2expression
in abscission zone + 5 mm of the flanking
pedicel compared toTAPG4and phytohormone
response markers. Gene expression in
knockdown (blue) and overexpressing plants
(magenta) is shown relative to wild type
normalized to 1 (dashed line; mean ± SD).
Red asterisks mark significant differences
between transgenic plants and wild type (ttest).
Pvalues are shown for significant differences
between knockdown and overexpressing
plants (ttest). (CtoE) Pedicel abscission
assayed over time forSlPhyt2knockdown, over-
expressing, and WT plants in a detached-flower
bioassay (error bars indicate 95% confidence
interval;n= number of inflorescences analyzed;
drop curves marked by the same letter are
not significantly different ata=0.05).(C)
Control in H 2 O; (D) 1mMPSK;and(E)5mM PSK.
(FandG) Bioassay for abscission in knockdown,
overexpressing, and WT inflorescence explants
showing the time (day) until >50% of pedicels
had abscised. (F) 1-MCP–treated inflorescences
compared to controls in ambient air. (G)
50 mM indole-3-acetic acid (IAA)-treated
inflorescences compared to solvent-treated
controls; Mann Whitney test based on ranks.

SlPhyt2

expression (fold change)

A


ERF4

Gene expression (fold change)

B

SlPhyt2TAPG4

IAA3 TPRP ERT10 PK7

0

5

10

**

** *

*

*

ns
**

ns ns

ns

KD
OE

ns

p=0.0015
p<0.0001
p=0.0023

p=0.0318

p=0.0038

prox AZ distprox AZ dist

0

2

4

6

G

> 50 % Pedicel drop (days)

0

5

10

15

KD WT WTKDOE OE
control IAA

p<0.0001

p<0.0001

p<0.0001

p<0.0001

C


012345678910

0

20

40

60

80

100

Pedicel drop (%)

WT n = 66, A
KD n = 149, B
OE n = 39, C

H 2 O

D


Time (days)

012345678910

0

20

40

60

80

100

Time (days)

WT n = 6, A
KD n = 18, B
OE n = 18, B

Pedicel drop (%)

1 μM PSK

Time (days)

012345678910

0

20

40

60

80

100

WT n = 16, A
KD n = 51, A
OE n = 21, A

Pedicel drop (%)

E


5 μM PSK

F

0

2

4

6

8

10

KD WT KDOE WT OE
control 1-MCP

> 50 % Flower drop (days)

p<0.0001
p<0.0001

p<0.0001
p<0.0001

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