Journal Pre-proof

Regulation of vascular cambium activity

Huanzhong Wang

PII: S0168-9452(19)31495-5
DOI: https://doi.org/10.1016/j.plantsci.2019.110322
Reference: PSL 110322

To appear in: Plant Science

Received Date: 16 August 2019

Revised Date: 25 September 2019
Accepted Date: 24 October 2019

Please cite this article as: Wang H, Regulation of vascular cambium activity, Plant Science
(2019), doi: https://doi.org/10.1016/j.plantsci.2019.110322

This is a PDF file of an article that has undergone enhancements after acceptance, such as
the addition of a cover page and metadata, and formatting for readability, but it is not yet the
definitive version of record. This version will undergo additional copyediting, typesetting and
review before it is published in its final form, but we are providing this version to give early
visibility of the article. Please note that, during the production process, errors may be
discovered which could affect the content, and all legal disclaimers that apply to the journal
pertain.

© 2019 Published by Elsevier.

Table of Contents
1. Introduction ................................................................................................................................ 1
2. Formation of stem vascular cambium ...................................................................................... 2
3. Long-distance hormonal signaling regulates cambium activity ............................................ 3
3.1 Auxin ..........................................................................................................................3

3.2 Cytokinin and gibberellin ...........................................................................................4

3.3 Other plant hormones .................................................................................................4

of
4. Short-range peptide signaling regulates cambium activity .........................................5

5. Interaction between long- and short-range signaling ..................................................6

ro
6. Peptide Signals from outer-layers affect cambial activity ...................................................... 7
7. Future perspectives ........................................................................................................8

-p
re
lP
na
ur
Jo

0
Regulation of vascular cambium activity

Huanzhong Wang

Department of Plant Science and Landscape Architecture, University of Connecticut, 1376 Storrs

Rd, Storrs, CT 06269

Abstract

Vascular cambium contributes to lateral growth in dicotyledonous plants and

of
gymnosperms. Physiological, genetics and molecular studies indicate that cambial activity is

ro
regulated by a combination of long-distance hormonal signals and short-range peptide signaling

pathways. Communication from endodermis and phloem tissues also affects cambial stem cell

-p
proliferation. Interactions between these signaling pathways provide flexibility for vascular

development. In this mini-review, we discuss the new findings in long- and short-range signaling
re
pathways in regulating vascular cambium proliferation and provide future perspectives in the
lP

cambium research. Deep imaging and mathematical modeling will help further dissecting the

functional mechanisms of cambial activity control.

na

1. Introduction

Vascular cambium is responsible for the secondary growth in plant stem, hypocotyl, and
ur

root tissues. During secondary growth, cambial stem cells proliferate and produce daughter cells,

which can then differentiate into secondary phloem and secondary xylem. Phloem and xylem
Jo

form a complex transport system that supplies nutrients and water to all plant organs [1, 2]. In

xylem tissues, tracheary elements and xylary fibers develop secondary cell walls, providing

mechanical strength for upward growth. Vascular cambium is considered as an evolutionary

innovation enabling plants to access better light conditions by growing tall and strong.

1
 Second growth consumes large amounts of energy and resources and has to be coordinated

with primary growth. Therefore, cambial activity should respond to developmental cues from the

primary meristems and the ever-changing environment. Plant hormones produced from apical

meristems, such as auxin and cytokinin, may serve as long-distance signals to regulate the

cambium activity. Endodermis and cortex surrounding the cambium also influence cambium

proliferation. In tree species, cambial activity has dormant and active cycles in response to

seasonal changes. Therefore, cambial activity is under the regulation of a plethora of

of
developmental and environmental signals. The main focus of this mini-review is to discuss the

ro
regulation of vascular cambial activity during secondary growth in stem. For pre-procambium

initiation, vascular cell specification and differentiation, and cambium regeneration, readers can

refer to recent review papers [1, 3-6].

-p
re
2. Formation of stem vascular cambium

All the above-ground tissues are ultimately derived from the shoot apical meristem (SAM).
lP

In Arabidopsis shoot, the rib zone of the shoot apical meristem is responsible for the initiation of

all stem tissues, including the stem vascular system [7, 8]. However, the rib zone is deep
na

underneath the shoot apex and surrounded by several layers of other cell types. It is impossible to

directly observe the rib zone without first clearing the outside tissues. Besides, vascular bundles
ur

are indistinguishable from ground tissues at the shoot apical region; and become visible in stem

cross-sections at about 50 micrometers below the shoot apices [9]. A suitable marker is needed
Jo

but still lacking to facilitate studying vascular initiation. Mutants with vascular initiation defects

have not been discovered due to the inaccessibility of vascular tissue. These technical difficulties

have impeded genetics studies of vascular initiation in stem tissues.

2
 After vascular initiation, discrete vascular bundles form along the periphery of the stem

(Fig. 1A). The vascular bundles are organized in a collateral pattern, with phloem facing outside

and xylem inside the stem. The vascular procambium/cambium is located between the phloem

and xylem cells. During secondary growth, interfascicular cambium is transdifferentiated from

parenchyma cells, starting from the bundle toward the middle of the interfascicular region.

Eventually, cambial cells form a closed ring structure [3, 4]. We still know very little about the

de novo formation of cambial cells in the interfascicular regions, let alone the regulation of this

of
process. Once formed, the cambium cells can proliferate and continuously produce secondary

ro
xylem and secondary phloem, enabling lateral thickening of the stem.

3. Long-distance hormonal signaling regulates cambium activity

-p
Secondary growth in stem and root tissues has to be coordinated with primary growth at the
re
shoot-root axis. Phytohormones play critical roles in communication among different organs and

may function as signal molecules between primary and secondary meristems (Fig 1B).
lP

3.1 Auxin

Auxin is produced from shoot apex and transport basipetally toward the root [10]. Removal
na

of shoot apex halts cambium activity and secondary growth, whereas exogenous application of

synthetic auxin re-activate cambial cell proliferation [3, 11]. Indeed, auxin distributes in a radial
ur

concentration gradient, showing the highest concentration at the cambium zone in tree stem

sections [12, 13]. The auxin distribution pattern, rather than absolute auxin concentration, may
Jo

define the spatial localization of vascular stem cells. This hypothesis is supported by the

observation of a decreasing auxin concentration in cell types moving away from the cambium

region [12, 13]. Auxin distribution depends on plasma membrane-localized auxin efflux carriers

in the PIN-FORMED (PIN) family [14, 15]. In addition, tonoplast localized auxin influx

3
transporters affect auxin retention in cambial cells [16]. A recent paper highlighted the

importance of auxin in defining the stem cell organizer function of the xylem identity cells in the

cambium [17]. These studies have confirmed the importance of auxin to cambium activity, but

the functional mechanism is still not clear due to complex interactions with other plant

hormones.

3.2 Cytokinin and gibberellin

Cytokinin is another essential plant hormone in regulating cambium activity. In the primary

of
root, either disruption of the expression of cytokinin receptor or depletion of cytokinin content

ro
by overexpressing cytokinin oxidase/dehydrogenase (CKX) genes prohibits periclinal cell

division of pro-cambium cells and reduce the size of the vasculature [18-20]. A mutually

-p
inhibitory feedback loop between auxin and cytokinin involves multiple layers of transcriptional
re
regulation [19]. The feedback loop may help to define the boundaries between the cambium and

differentiating xylem. The histidine kinases CYTOKININ-INDEPENDENT1 and

lP

ARABIDOPSIS HISTIDINE KINASE2 and 3 play positive roles in regulating vascular

cambium proliferation in Arabidopsis stem [21]. In poplar trees, overexpression of the cytokinin
na

biosynthesis gene ISOPENTENYLTRANSFERASE (IPT) enhances cambial cell division and

biomass production [22]. Gibberellin also plays a vital role in cambium activity. Overexpression
ur

of Gibberellin 20-oxidase (GA20ox), a gibberellin biosynthesis gene, enhances cambial

proliferation and secondary growth [23]. In contrast, mutants with defects in GA biosynthesis
Jo

have reduced cambium activity [24].

3.3 Other plant hormones

The aforementioned plant hormones, i.e., auxin, cytokinin, and gibberellin, are all generally

considered as growth-promoting hormones. Other hormones, including ethylene, Jasmonic acid,

4
brassinosteroid and strigolactone (SL), also regulate cambium activity. Treatment of hybrid

aspen trees with ethylene or aminocyclopropane-1-carboxylate (ACC) promotes cambial division

and wood formation [25]. In transgenic trees, overexpression of ACC oxidase also increases

cambial activity and secondary growth [25]. In Arabidopsis, the ethylene overproducer1 (eto1)

plants demonstrate increased vascular size in hypocotyl and inflorescence stem [26]. Arabidopsis

plants treated with jasmonic acid enhance cambium proliferation in the interfascicular region

[27]. Brassinosteroid positively regulates procambial division and vascular bundle number

of
probably through reducing auxin transport in stem [9]. SL treatment stimulates cambial activity

ro
in Arabidopsis stem, while mutations of SL signaling or biosynthesis inhibit cambial activity

[28]. Interestingly, all of the aforementioned plant hormones, either function in promoting plant

-p
growth or response to environmental change, are positive regulators of cambium activity. It is
re
likely that some unidentified signals may function as negative regulators of cambial activity.

Nevertheless, these studies indicate that cambium can incorporate signals from both endogenous
lP

developmental programs and exogenous environmental cues.

4. Short-range peptide signaling regulates cambium activity

na

Both apical and lateral meristems are regulated by short-range peptide signals, which

involve a CLAVATA3 (CLV3)/EMBRYO SURROUNDING REGIONRELATED (CLE)

ur

peptide, a leucine-rich repeat receptor-like kinase (LRR-RLK) and a downstream WUSCHEL-

related homeobox (WOX) transcription factor [29, 30, 31 ]. In the vascular meristem, the CLE
Jo

peptide TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF)

promotes stem cell proliferation and inhibits the formation of tracheary elements [32]. The TDIF

peptide physically binds to its receptor TDR [33], also named Phloem intercalated with Xylem

(PXY) [34]. Mutation of TDR/PXY results in loss of vascular stem cells and enhanced xylem

5
differentiation [35, 36]. Downstream of the TDIF-PXY module, a WOX transcription factor,

WOX4 is responsible for promoting cambium proliferation [35, 37]. TDIF induces the

expression of WOX4 in the cambium, which is different from the function of CLE peptides in the

shoot or root apical meristem, where CLEs negatively regulate WOX genes [35, 37]. Xylem cell

differentiation is also regulated by TDIF-PXY module, but through a different pathway not

involving WOX4 (Fig 1B). The GLYCOGEN SYNTHASE KINASE 3 (GSK3) proteins interact

with PXY and repress the transcription factor BRI1-EMS SUPPRESSOR 1 (BES1), as a result

of
inhibiting xylem cell differentiation [38]. The TDIF signaling pathway has improved our

ro
understanding of cambial activity, but there are still many unanswered questions. For example,

how the maintenance of cambial stem cells is achieved with the feed-forward regulation of

-p
TDIF-PXY module on WOX4. Feedback mechanisms are important to peptide signaling in shoot
re
and root apical meristems, but which have not been identified in the vascular meristem. The

vascular patterning in pxy/tdr mutant is disrupted, but the functional mechanism is also still a
lP

mystery.

5. Interaction between long- and short-range signaling

na

WOX4 regulates cambial cell proliferation downstream of the TDIF-PXY module.

Meanwhile, WOX4 is also induced by auxin in the cambial cells. In Arabidopsis stem, the
ur

induction of WOX4expression is dependent on a functional PXY [39]. In a recent report, the

auxin response factor 5 (ARF5)/ MONOPTEROS (MP) directly binds to the WOX4 promoter
Jo

and attenuates WOX4 expression in cambial cells [40]. Cytokinin signaling also interacts with the

TDIF peptide signaling. A GSK3 family member BRASSINOSTEROID-INSENSITIVE 2-LIKE

1 (BIL1) inhibits cambium activity through phosphorylating ARF5/MP, as a result upregulating

the negative regulators of cytokinin signaling ARABIDOPSIS RESPONSE REGULATOR 7

6
(ARR7) and ARR15 [41]. The PXY/TDR can inhibit BIL1 activity, therefore attenuating the

effect of MP/ARF5 on ARR7 and ARR15 expression and increasing vascular cambial activity

[41]. WOX14 and WOX4 function redundantly in promoting cambial activity and are both

downstream of TDIF-PXY module [42]. WOX14 may also function upstream of GA

biosynthesis. Overexpression of WOX14 promotes the accumulation of active GA, whereas loss

of WOX14 function results in GA-deficient phenotypes that can be complemented by exogenous

GA application [43]. Ethylene response factors (ERFs) are required for cambial cell division.

of
The expression levels of ERF1, ERF109 and ERF18 are upregulated in pxy/tdr and wox4 mutant

ro
plants [26]. These ERFs promote procambial proliferation depending on a functional TDR [26].

These studies demonstrate complex interactions between long-distance hormonal signaling and

-p
short-range peptide signaling pathway. It seems that the local short-range TDIF signaling act as a
re
signaling hub to incorporate various hormonal signals. Further dissecting the interactions

between the long-distance and short-range signaling pathways should enhance our understanding
lP

of the regulation of cambial activity.

6. Peptide Signals from outer-layers affect the cambial activity

na

The ERECTA (ER) gene encodes a membrane-localized receptor kinase, and mutation of

ER results in a compact inflorescence with short internodes, short pedicels, and blunt fruits [44].
ur

ER-LIKE1 (ERL1) and ERL2 function redundantly with ER and together comprise the ER

family (ERf) [45]. Both ER and ERL1 are expressed in phloem cells in the stem [46]. Phloem
Jo

cells of the er erl1 mutant are often located adjacent to xylem cells without intervening

procambial cells [46]. This phenotype is similar to the pxy/tdr mutant, indicating that ERf

involved signaling pathway positively regulates the cambium activity. More server defects in

cambial cell development have been observed in the tdr er double mutant, suggesting that TDR

7
and ER act in parallel pathways in cambium maintenance [26, 46]. A recent study has shown that

PXY and ER cross- and inter-family transcriptional regulation occurs, but it differs between stem

and hypocotyl, indicating that PXY and ER signaling interact to coordinate development in

different organs[47]. In Arabidopsis, EPIDERMAL PATTERNING FACTOR1 (EPF1) and

EPF2 peptides are the founding members of the EPF-LIKE (EPFL) family and function as

ligands for ERf proteins [48-50]. EPFL4 and EPFL6 peptides are produced from the epidermis

and translocated to the phloem, where they bind the ER receptor and regulate cambial

of
proliferation and inflorescence stem growth [51, 52]. Transcription factor genes downstream of

ro
the EPFL4/6-ER module have not been identified to date. It is reasonable to hypothesize that a

downstream mobile component is needed to transmit the signal from the phloem, where the

-p
EPFL4/6-ERf module locates, into the cambial cells. Identification of such a mobile component
re
should help us to understand the functional mechanism of EPFL4/6-ERf signaling. In addition,

the signaling pathways initiated by the EPFL4/6-ER and TDIF-PXY modules both regulate the
lP

cambium activity. The interaction of these two signaling pathways should provide new insight

into the regulation of cambium activity.

na

7. Future perspectives

Physiological and genetics studies have shed light on the regulation of vascular cambium
ur

activity. However, compared with the apical meristems, very little is known about the vascular

stem cell maintenance and the control of cambial proliferation. Besides, pro-cambium and
Jo

cambium initiation in stem has not been investigated, while considerable progress has been made

in root and leaf vein systems [53, 54]. Of particular interest would be to investigate whether

mechanisms identified from apical meristems or leaf vein development can be applied to stem

vascular cambium. As mentioned earlier, one of the difficulties in studying vascular cambium is

8
the localization of this tissue that is embedded in layers of other cell types. Stem tissues can be

chemically cleared to obtain high-resolution whole-mount imaging (deep imaging) of the three-

dimensional stem tissue organization. Deep imaging coupled with gene expression assay help to

characterize the initiation and development of cambial cells [55, 56]. Mathematical modeling and

simulation combined with live imaging and clonal analysis are also powerful tools to investigate

the regulatory mechanisms [9, 57]. Further research should also focus on the interactions

between known regulatory pathways of vascular development.

of
Acknowledgement

ro
The author thanks Jonathan Mahoney for critical reading of the manuscript. This work is

supported by National Science Foundation (IOS-1453048), and in part, by USDA NIFA Hatch

project #CONS00925 to HW.

-p
re
lP
na
ur
Jo

9
Reference:
[1] T. Greb, J.U. Lohmann, Plant Stem Cells, Current biology : CB, 26 (2016) R816-821.
[2] S. Miyashima, J. Sebastian, J.Y. Lee, Y. Helariutta, Stem cell function during plant vascular
development, The EMBO journal, 32 (2013) 178-193.
[3] U. Fischer, M. Kucukoglu, Y. Helariutta, R.P. Bhalerao, The Dynamics of Cambial Stem Cell Activity,
Annual review of plant biology, 70 (2019) 293-319.
[4] E. Pierre-Jerome, C. Drapek, P.N. Benfey, Regulation of Division and Differentiation of Plant Stem
Cells, Annual review of cell and developmental biology, 34 (2018) 289-310.
[5] R. Ruonala, D. Ko, Y. Helariutta, Genetic Networks in Plant Vascular Development, Annual review of
genetics, 51 (2017) 335-359.
[6] K.M. Furuta, E. Hellmann, Y. Helariutta, Molecular control of cell specification and cell differentiation
during procambial development, Annual review of plant biology, 65 (2014) 607-638.
[7] E. Aichinger, N. Kornet, T. Friedrich, T. Laux, Plant stem cell niches, Annual review of plant biology, 63

of
(2012) 615-636.
[8] Z.L. Nimchuk, Y. Zhou, P.T. Tarr, B.A. Peterson, E.M. Meyerowitz, Plant stem cell maintenance by
transcriptional cross-regulation of related receptor kinases, Development, 142 (2015) 1043-1049.

ro
[9] M. Ibanes, N. Fabregas, J. Chory, A.I. Cano-Delgado, Brassinosteroid signaling and auxin transport are
required to establish the periodic pattern of Arabidopsis shoot vascular bundles, Proceedings of the
National Academy of Sciences of the United States of America, 106 (2009) 13630-13635.

-p
[10] T. Berleth, J. Mattsson, C.S. Hardtke, Vascular continuity and auxin signals, Trends Plant Sci, 5 (2000)
387-393.
[11] J. Agusti, R. Lichtenberger, M. Schwarz, L. Nehlin, T. Greb, Characterization of transcriptome
remodeling during cambium formation identifies MOL1 and RUL1 as opposing regulators of secondary
re
growth, PLoS genetics, 7 (2011) e1001312.
[12] H. Tuominen, L. Puech, S. Fink, B. Sundberg, A radial concentration gradient of indole-3-acetic acid
is related to secondary xylem development in hybrid aspen, Plant physiology, 115 (1997) 577-585.
lP

[13] C. Uggla, T. Moritz, G. Sandberg, B. Sundberg, Auxin as a positional signal in pattern formation in
plants, Proceedings of the National Academy of Sciences of the United States of America, 93 (1996)
9282-9286.
[14] E. Benkova, M. Michniewicz, M. Sauer, T. Teichmann, D. Seifertova, G. Jurgens, J. Friml, Local, efflux-
na

dependent auxin gradients as a common module for plant organ formation, Cell, 115 (2003) 591-602.
[15] M. Sauer, J. Balla, C. Luschnig, J. Wisniewska, V. Reinohl, J. Friml, E. Benkova, Canalization of auxin
flow by Aux/IAA-ARF-dependent feedback regulation of PIN polarity, Genes & development, 20 (2006)
2902-2911.
[16] P. Ranocha, O. Dima, R. Nagy, J. Felten, C. Corratge-Faillie, O. Novak, K. Morreel, B. Lacombe, Y.
ur

Martinez, S. Pfrunder, X. Jin, J.P. Renou, J.B. Thibaud, K. Ljung, U. Fischer, E. Martinoia, W. Boerjan, D.
Goffner, Arabidopsis WAT1 is a vacuolar auxin transport facilitator required for auxin homoeostasis,
Nature communications, 4 (2013) 2625.
Jo

[17] O. Smetana, R. Makila, M. Lyu, A. Amiryousefi, F. Sanchez Rodriguez, M.F. Wu, A. Sole-Gil, M. Leal
Gavarron, R. Siligato, S. Miyashima, P. Roszak, T. Blomster, J.W. Reed, S. Broholm, A.P. Mahonen, High
levels of auxin signalling define the stem-cell organizer of the vascular cambium, Nature, 565 (2019)
485-489.
[18] K. Nieminen, J. Immanen, M. Laxell, L. Kauppinen, P. Tarkowski, K. Dolezal, S. Tahtiharju, A. Elo, M.
Decourteix, K. Ljung, R. Bhalerao, K. Keinonen, V.A. Albert, Y. Helariutta, Cytokinin signaling regulates
cambial development in poplar, Proceedings of the National Academy of Sciences of the United States of
America, 105 (2008) 20032-20037.

10
[19] A.P. Mahonen, A. Bishopp, M. Higuchi, K.M. Nieminen, K. Kinoshita, K. Tormakangas, Y. Ikeda, A.
Oka, T. Kakimoto, Y. Helariutta, Cytokinin signaling and its inhibitor AHP6 regulate cell fate during
vascular development, Science, 311 (2006) 94-98.
[20] T. Werner, V. Motyka, V. Laucou, R. Smets, H. Van Onckelen, T. Schmulling, Cytokinin-deficient
transgenic Arabidopsis plants show multiple developmental alterations indicating opposite functions of
cytokinins in the regulation of shoot and root meristem activity, The Plant cell, 15 (2003) 2532-2550.
[21] J. Hejatko, H. Ryu, G.T. Kim, R. Dobesova, S. Choi, S.M. Choi, P. Soucek, J. Horak, B. Pekarova, K.
Palme, B. Brzobohaty, I. Hwang, The histidine kinases CYTOKININ-INDEPENDENT1 and ARABIDOPSIS
HISTIDINE KINASE2 and 3 regulate vascular tissue development in Arabidopsis shoots, The Plant cell, 21
(2009) 2008-2021.
[22] M. Matsumoto-Kitano, T. Kusumoto, P. Tarkowski, K. Kinoshita-Tsujimura, K. Vaclavikova, K.
Miyawaki, T. Kakimoto, Cytokinins are central regulators of cambial activity, Proceedings of the National
Academy of Sciences of the United States of America, 105 (2008) 20027-20031.

of
[23] M. Mauriat, T. Moritz, Analyses of GA20ox- and GID1-over-expressing aspen suggest that
gibberellins play two distinct roles in wood formation, The Plant journal : for cell and molecular biology,
58 (2009) 989-1003.

ro
[24] L. Ragni, K. Nieminen, D. Pacheco-Villalobos, R. Sibout, C. Schwechheimer, C.S. Hardtke, Mobile
gibberellin directly stimulates Arabidopsis hypocotyl xylem expansion, The Plant cell, 23 (2011) 1322-
1336.

-p
[25] J. Love, S. Bjorklund, J. Vahala, M. Hertzberg, J. Kangasjarvi, B. Sundberg, Ethylene is an endogenous
stimulator of cell division in the cambial meristem of Populus, Proceedings of the National Academy of
Sciences of the United States of America, 106 (2009) 5984-5989.
[26] J.P. Etchells, C.M. Provost, S.R. Turner, Plant vascular cell division is maintained by an interaction
re
between PXY and ethylene signalling, PLoS genetics, 8 (2012) e1002997.
[27] E.M. Sehr, J. Agusti, R. Lehner, E.E. Farmer, M. Schwarz, T. Greb, Analysis of secondary growth in the
Arabidopsis shoot reveals a positive role of jasmonate signalling in cambium formation, The Plant
lP

journal : for cell and molecular biology, 63 (2010) 811-822.

[28] J. Agusti, S. Herold, M. Schwarz, P. Sanchez, K. Ljung, E.A. Dun, P.B. Brewer, C.A. Beveridge, T.
Sieberer, E.M. Sehr, T. Greb, Strigolactone signaling is required for auxin-dependent stimulation of
secondary growth in plants, Proceedings of the National Academy of Sciences of the United States of
America, 108 (2011) 20242-20247.
na

[29] J.C. Fletcher, U. Brand, M.P. Running, R. Simon, E.M. Meyerowitz, Signaling of cell fate decisions by
CLAVATA3 in Arabidopsis shoot meristems, Science, 283 (1999) 1911-1914.
[30] K.F. Mayer, H. Schoof, A. Haecker, M. Lenhard, G. Jurgens, T. Laux, Role of WUSCHEL in regulating
stem cell fate in the Arabidopsis shoot meristem, Cell, 95 (1998) 805-815.
ur

[31] H. Schoof, M. Lenhard, A. Haecker, K.F.X. Mayer, G. Jurgens, T. Laux, The stem cell population of
Arabidopsis shoot meristems is maintained by a regulatory loop between the CLAVATA and WUSCHEL
genes, Cell, 100 (2000) 635-644.
Jo

[32] Y. Ito, I. Nakanomyo, H. Motose, K. Iwamoto, S. Sawa, N. Dohmae, H. Fukuda, Dodeca-CLE peptides
as suppressors of plant stem cell differentiation, Science, 313 (2006) 842-845.
[33] Y. Hirakawa, H. Shinohara, Y. Kondo, A. Inoue, I. Nakanomyo, M. Ogawa, S. Sawa, K. Ohashi-Ito, Y.
Matsubayashi, H. Fukuda, Non-cell-autonomous control of vascular stem cell fate by a CLE
peptide/receptor system, Proceedings of the National Academy of Sciences of the United States of
America, 105 (2008) 15208-15213.
[34] K. Fisher, S. Turner, PXY, a receptor-like kinase essential for maintaining polarity during plant
vascular-tissue development, Current biology : CB, 17 (2007) 1061-1066.
[35] Y. Hirakawa, Y. Kondo, H. Fukuda, TDIF peptide signaling regulates vascular stem cell proliferation
via the WOX4 homeobox gene in Arabidopsis, The Plant cell, 22 (2010) 2618-2629.

11
[36] J.P. Etchells, S.R. Turner, The PXY-CLE41 receptor ligand pair defines a multifunctional pathway that
controls the rate and orientation of vascular cell division, Development, 137 (2010) 767-774.
[37] J. Ji, J. Strable, R. Shimizu, D. Koenig, N. Sinha, M.J. Scanlon, WOX4 promotes procambial
development, Plant physiology, 152 (2010) 1346-1356.
[38] Y. Kondo, T. Ito, H. Nakagami, Y. Hirakawa, M. Saito, T. Tamaki, K. Shirasu, H. Fukuda, Plant GSK3
proteins regulate xylem cell differentiation downstream of TDIF-TDR signalling, Nature communications,
5 (2014) 3504.
[39] S. Suer, J. Agusti, P. Sanchez, M. Schwarz, T. Greb, WOX4 imparts auxin responsiveness to cambium
cells in Arabidopsis, The Plant cell, 23 (2011) 3247-3259.
[40] K. Brackmann, J. Qi, M. Gebert, V. Jouannet, T. Schlamp, K. Grunwald, E.S. Wallner, D.D. Novikova,
V.G. Levitsky, J. Agusti, P. Sanchez, J.U. Lohmann, T. Greb, Spatial specificity of auxin responses
coordinates wood formation, Nature communications, 9 (2018) 875.
[41] S. Han, H. Cho, J. Noh, J. Qi, H.J. Jung, H. Nam, S. Lee, D. Hwang, T. Greb, I. Hwang, BIL1-mediated

of
MP phosphorylation integrates PXY and cytokinin signalling in secondary growth, Nature plants, 4 (2018)
605-614.
[42] J.P. Etchells, C.M. Provost, L. Mishra, S.R. Turner, WOX4 and WOX14 act downstream of the PXY

ro
receptor kinase to regulate plant vascular proliferation independently of any role in vascular
organisation, Development, 140 (2013) 2224-2234.
[43] E. Denis, N. Kbiri, V. Mary, G. Claisse, E.S.N. Conde, M. Kreis, Y. Deveaux, WOX14 promotes

-p
bioactive gibberellin synthesis and vascular cell differentiation in Arabidopsis, The Plant journal : for cell
and molecular biology, 90 (2017) 560-572.
[44] K.U. Torii, N. Mitsukawa, T. Oosumi, Y. Matsuura, R. Yokoyama, R.F. Whittier, Y. Komeda, The
Arabidopsis ERECTA gene encodes a putative receptor protein kinase with extracellular leucine-rich
re
repeats, The Plant cell, 8 (1996) 735-746.
[45] E.D. Shpak, C.T. Berthiaume, E.J. Hill, K.U. Torii, Synergistic interaction of three ERECTA-family
receptor-like kinases controls Arabidopsis organ growth and flower development by promoting cell
lP

proliferation, Development, 131 (2004) 1491-1501.

[46] N. Uchida, M. Tasaka, Regulation of plant vascular stem cells by endodermis-derived EPFL-family
peptide hormones and phloem-expressed ERECTA-family receptor kinases, Journal of experimental
botany, 64 (2013) 5335-5343.
[47] N. Wang, K.S. Bagdassarian, R.E. Doherty, J.T. Kroon, K.A. Connor, X.Y. Wang, W. Wang, I.H. Jermyn,
na

S.R. Turner, J.P. Etchells, Organ-specific genetic interactions between paralogues of the PXY and ER
receptor kinases enforce radial patterning in Arabidopsis vascular tissue, Development, 146 (2019).
[48] K. Hara, R. Kajita, K.U. Torii, D.C. Bergmann, T. Kakimoto, The secretory peptide gene EPF1 enforces
the stomatal one-cell-spacing rule, Genes & development, 21 (2007) 1720-1725.
ur

[49] L. Hunt, J.E. Gray, The signaling peptide EPF2 controls asymmetric cell divisions during stomatal
development, Current biology : CB, 19 (2009) 864-869.
[50] J.S. Lee, T. Kuroha, M. Hnilova, D. Khatayevich, M.M. Kanaoka, J.M. McAbee, M. Sarikaya, C.
Jo

Tamerler, K.U. Torii, Direct interaction of ligand-receptor pairs specifying stomatal patterning, Genes &
development, 26 (2012) 126-136.
[51] E.B. Abrash, K.A. Davies, D.C. Bergmann, Generation of signaling specificity in Arabidopsis by
spatially restricted buffering of ligand-receptor interactions, The Plant cell, 23 (2011) 2864-2879.
[52] N. Uchida, J.S. Lee, R.J. Horst, H.H. Lai, R. Kajita, T. Kakimoto, M. Tasaka, K.U. Torii, Regulation of
inflorescence architecture by intertissue layer ligand-receptor communication between endodermis and
phloem, Proceedings of the National Academy of Sciences of the United States of America, 109 (2012)
6337-6342.

12
[53] C.L. Wenzel, M. Schuetz, Q. Yu, J. Mattsson, Dynamics of MONOPTEROS and PIN-FORMED1
expression during leaf vein pattern formation in Arabidopsis thaliana, The Plant journal : for cell and
molecular biology, 49 (2007) 387-398.
[54] Z.H. Ye, Vascular tissue differentiation and pattern formation in plants, Annual review of plant
biology, 53 (2002) 183-202.
[55] V. Oralova, J.T. Rosa, M. Soenens, J.W. Bek, A. Willaert, P.E. Witten, A. Huysseune, Beyond the
whole-mount phenotype: high-resolution imaging in fluorescence-based applications on zebrafish,
Biology open, 8 (2019).
[56] E. Truernit, H. Bauby, B. Dubreucq, O. Grandjean, J. Runions, J. Barthelemy, J.C. Palauqui, High-
resolution whole-mount imaging of three-dimensional tissue organization and gene expression enables
the study of Phloem development and structure in Arabidopsis, The Plant cell, 20 (2008) 1494-1503.
[57] S. Bencivenga, A. Serrano-Mislata, M. Bush, S. Fox, R. Sablowski, Control of Oriented Tissue Growth
through Repression of Organ Boundary Genes Promotes Stem Morphogenesis, Developmental cell, 39

of
(2016) 198-208.

ro
-p
re
lP
na
ur
Jo

13
 Figure 1.

Jo
ur
na

14
lP
re
-p
ro
of