Science - USA (2021-12-24)

Roszaket al.,Science 374 , eaba5531 (2021) 24 December 2021 3of9

Row Z−Score

ROPGEF2

ROPGEF3

ROP9

-2 02

ROPGEF2 ROPGEF3

-1 0 1 2

PSE

MSE

procambium

PSE

MSE

procambium

1 st

2 nd

pROPGEF3::

Cit-ROPGEF3

pROPGEF2::

Cit-ROPGEF2 pROPGEF5::Cit-ROPGEF5

Parental line spk1

15

20

25

30

35

0’ 27’ 57’ 88’

AA' BB'

A A’B B’

0’ 33’ 60’ 90’

pROPGEF3::

Cit-ROPGEF3

pROPGEF2::

Cit-ROPGEF2

pROPGEF5::

Cit-ROPGEF5

f’ f’ f’’

f’’

*

*

*

*

ROP9-CA (n=18)

Col-0 (n=18)

0 50 100 150 200

0

5

10

15

20

[μm] distance from the QC

count

Parental

line

spk1

Non-induced Induced (24h) pRPS5A::XVE>>Cit-GEF5

***

p=6.623E-05

7/10 10/10 3/5 2/5

B

A C D E

F G H

LM

I J K

Fig. 2. PEARs control asymmetric divisions by promoting ROP signaling in
the phloem pole.(A) Schematic indicating position of the two periclinal divisions
in the phloem cell lineage. (B) Expression of ROPGEF2 and ROPGEF3 at the
time of phloem lineage bifurcation. (C) Peak expression of ROPGEF2, ROPGEF3,
and ROP9 in the early phloem cells as detected in the pseudotime-ordered single-
cell protophloem sieve element transcriptome data. (D) Expression pattern of
phloem-enriched ROPGEFs. ROPGEF3, and ROPGEF5 share similar expression
domains, which are enriched in protophloem sieve element and adjacent vascular
cell files; ROPGEF2 is expressed in protophloem sieve element but also in other
outer procambial cells and pericycle (fig. S4D). Scale bars, 25mm. (E) Expression

of ROPGEF2, ROPGEF3, and ROPGEF5 in thepearsextuple mutant background.

Scale bars, 25mm. (F) Protein localization ofpROPGEF5::Cit-ROPGEF5during

anticlinal (f′) and periclinal (f′′) cell division. Gaps in ROPGEF5 signal are indicated

with an asterisk. Scale bars, 25mm. (G) Depletion of Cit-ROPGEF5 membrane

signal at the cortical division zone (CDZ) during cell division. CDZ is marked by

accumulating cortical microtubules (mCherry-TUA5) forming a pre-prophase

band (white arrowheads). Scale bars, 25mm. (H) Time course analysis of the

dynamic pattern of active ROP signaling in the dividing phloem cells. Depletion of

pPEAR1::mScarlet-I-MIDD1DNsignal at the CDZ in the anticlinally (upper row)

and periclinally (lower row) dividing cells (yellow arrowheads). Quantification of

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