Science - 27.03.2020

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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