Root growth, cell processes and responses to environment

(1)

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Root growth, cell processes and responses to environment

Marie-Béatrice Bogeat-Triboulot

To cite this version:

Marie-Béatrice Bogeat-Triboulot. Root growth, cell processes and responses to environment. Bio- physique de l’interaction racine-sol, GDR PHYP (Physique des Plantes), Nov 2019, Clermont-Ferrand, France. pp.1-36. �hal-03139637�

(2)

Root growth, cell processes and responses to environment

Marie-Béatrice BOGEAT-TRIBOULOT UMR SILVA

INRA - Nancy - France

(3)

Root growth = expansion of material

• scale + growth intensity

• quantified by the Elemental Expansion Rate (EER) orstrain rate

mm mm^-1h^-1

2 0.00

0.10 0.20 0.30 0.40

0 2 4 6 8

ElementalExpansion Rate (h-1 )

Distance to root tip (mm) surface under the curve :

proportional to the root growth rate

(4)

100 µm

14.1 µm 127 µm

Pinus pinaster root apex

Cell length : 15 µm  320 µm : x 20!

Cell elongation

(5)

The expanding cell

cell wall tonoplast plasmalemma

cell wall extensibility

water

hydrostatique pressure of the cell sap on the cell wall

Π

_i

P

_i

Ψ

_o

solutes

• Cell wall deformation and synthesis

• Water and solutes inflow

(6)

Water flow into cell

• Le potentiel hydrique de la cellule : ^Ψ

_i

= P

_i

- Π

_i

– P = pression de turgescence : force hydrostatique – Π = pression osmotique (due aux solutés dissous)

σ= coefficient de réflectivité Si la membrane plasmique est

semi-perméable : σ ≈1

(7)

Cell wall relaxation reduces turgor pressure

6

• Excised pea growing stem segment

• connected to a water source

• not connected to a water source, without auxin

• not connected to a water source, with auxin (increases cell wall relaxation)

(8)

Cell wall relaxation : the basis of cell expansion

augmentation du volume cellulaire

RELACHEMENT DES PAROIS

entrée d ’eau et de solutés

baisse de P et donc de Ψ

_i

∆Ψ

_o,i

augmente rétablissement de P

arrêt de croissance

retour ∆Ψ à 0

(Ψ

_i

= P - Π

_i

)

J

_v

≈ Lp • ( ∆ P - σ ∆Π )

(9)

Growth rate depends linearly on turgor pressure

Growth Rate = φ (P - Y)

φ Y

Pritchard, 1994

La croissance est linéairement proportionnelle à la pression de turgescence

(le moteur de la croissance), au delà d’un seuil minimum

(10)

 Turgor pressure P

 Wall tension σ

𝜎𝜎 _{𝑧𝑧𝑧𝑧} P

𝜎𝜎 _{𝜃𝜃𝜃𝜃}

t R

𝜎𝜎

_{𝑧𝑧𝑧𝑧}

= 𝑅𝑅

2𝑡𝑡 𝑃𝑃 𝜎𝜎

_{𝜃𝜃𝜃𝜃}

= 𝑅𝑅

𝑡𝑡 𝑃𝑃 = 2𝜎𝜎

_{𝑧𝑧𝑧𝑧}

Wall tension and anisotropic growth

10 µm

𝜎𝜎

_{𝑧𝑧𝑧𝑧}

P

• Cell wall rheogical properties are different longitudinally and axially

• role of cellulose microfibrils orientation

(11)

Cell expansion parameters

• Force motrice du grandissement cellulaire :

– Pression de turgescence

• Contrôle du grandissement cellulaire :

– extensibilité des parois ( ø , Y )

– conductivité hydraulique volumique cellulaire (-> Lp) – Disponibilité en solutés / en eau

paroi cellulaire

tonoplaste plasmalemme

extensibilité des parois eau

Pression hydrostatique du suc vacuolaire sur les parois

P_i

Π

_i

Ψ

_o

solutés

Ψ

_i

_{= P}

_i

_- Π

_i

(12)

A biophysical model of cell expansion : Lockhart (1965)

Hydraulics

1/V (dV/dt) = L [σ (Π

_i

- Π

_o

) - P ]

Rheology of cell walls

1/V (dV/dt) = φ (P - Y)

) Y L (

L dt

dV

V ⋅

ⁱ^,^o

−

= +

⋅ σ ∆Π

φ φ 1

If L >> φ : Lφ/(L+φ) ≈ φ

Cell expansion is limited by the cell wall mechanical properties If L << φ : Lφ/(L+φ) ≈ L

Cell expansion is limited by the membrane hydraulic conductance

(13)

) Y L (

L dt

dV

V ⋅

ⁱ^,^o

−

= +

⋅ σ ∆Π

φ φ 1

Lockhart (1965)

• établi pour une cellule et pour un régime stationnaire

• conceptuel mais ne reflète pas tout, ne prend pas en compte:

– régime transitoire

– échelle spatiale : cellule / tissu (approximation pour un tissu)

(14)

Intensité du ‘gradient de potentiel induit par la croissance’

• _{P et} Π ± constant le long de la zone de croissance et au delà

• Ψ

_i

∼ Ψ

_o

 GIWP-gradient très faible : - Lp non limitante

- croissance contrôlée par propriétés mécaniques

Triboulot et al, 1995

water

(15)

ZC d’une feuille de fétuque

 Π ~ constant dans ZC et ZM

 P passe de ~ 5 bars dans ZC à ~ 9 bars dans la zone mature

Martre et al, 1999

 GIWP-gradient voisin de 0.3 MPa : Lp joue un rôle dans la limitation de la croissance

Intensité du ‘gradient de potentiel induit par la croissance’

ZC ZM

: P

∆ : Π

: REGR

(16)

Spatial characterisation of growth : time lapse photography and kinematics

15

0h

5h 4h 3h 2h 1h

2 mm

Infra-red illumination : monitoring of natural marks - no thigmomorphogenesis response

- renewing of marks -> long-time monitoring

F. Bizet

particule image velocimetry

Kineroot - Basuet al., 2007 Kymorod - Bastien et al, 2016

T0

T+1

V_max EER_max

GZ length

(17)

Infra-red illumination - natural marks

- renewing of marks -> long-time monitoring

Long term monitoring highlights temporal oscillations

16

MaximalEER (h-1)Root growth rate (mm/h) F. Bizet

(18)

Beemster et al, 2002 F. Bizet

Root growth = cell production + cell elongation

17

• quantitatively , most of growth is due to cell elongation

• meristem provides cells to the elongation zone

 cell proliferation rate affects root growth rate

(19)

Controls of transition from cell proliferation to cell expansion

18 Perilli et al. (2012)

Tsukagochi et al. (2012)

De Vos et al. (2014)

• cell-autonomous regulatory rules are insufficient to simulate smoothed developmental zones

• spatial cues solve the problem

timer/counter spatial cue

Regulation of cell status

(dividing/ expanding)  meristem size  cell proliferation rate  root growth rate

(20)

Root apical meristem characterisation

19

• RAM lengthdetermined from luminance profile (Bizet et al, 2015)

Relative luminance (%)

Distance from root tip (mm)

RAM length 75%

Velocity_RAM Cell length_RAM

• Cell production rate =

V_RAM

Cell length

IR spot length

RAM length(fromcelllength)

RMSE = 0,073

(21)

Variability of root growth components in poplar

20

• Inter-species variability of root growth rate is mostly explained by cell production rate (Gazquez and Beemster, 2017)

• Growth rate highly variable among roots within a root system, among species.

• origin?

Root growthrate (mm.h-1 )

Poplar genotypes PC1 (41%) x PC2(14%)

Youssef et al, 2018 J Exp Bot

(22)

Variability of root growth components in poplar

21

ORER EZ N_div Diam

P EER_max

Diam N_div

P

EZ ORER

EER_max

ORER P

N_div Diam

EER_max EZ

Youssef et al, 2018 J Exp Bot

Species and cultivars 24h of chemical treatments 1h of chemical treatments

• All parameters correlated

• Steady state growth rate : more driven by P than EER max

• Short term response : mainly due to the high responsiveness of EER

(23)

Root growth is highly responsive to environment

22

• Sensitive to water availability, nutrient availability, temperature, soil mechanical impedance, …

Rootgrowthrate (mm h-1)

maize Sharp et al. (1988)

Soil water potential (MPa)

Distance to root tip (mm)

EER (h-1 )

Skirycz & Inze (2010) Pritchard et al. (1991)

wheat

steady state versus transition phase

Root growthrate (mm h-1)

EER (h-1 )

Distance to root tip (mm)

Silk (1992) maize

29 °C 25 °C 19 °C 16 °C

(24)

Dynamics of root growth in response to osmotic stress

23

• Root growth is sensitive to water deficit

• time course of cell turgor recovery (motor of cell expansion) differs from that of growth : there are biological responses in addition to mechanical limitation

French et al (1994) Stress

What’s happen during the transition phase ?

• cell proliferation rate

• cell expansion rate

• transcriptome rewiring in the division zone and the elongation zone

(25)

Poplar as a model species

24

• Clonal propagation

• Easy cultivation in hydroponics

• Fast adventitious rooting

• Plagiotropic roots

• Fast growing root

• Large growth zone

-> easy manipulation

C.Youssef

(26)

Dynamics of root growth in response to osmotic stress

25

osmotic stress

 strong growth reduction

 return to stable growth rate after 2 hours

-> 2 snapshots during the transition phase, after 0.5h and 3h of stress

 kinematics -> growth components

 transcriptome in the division zone and the elongation zone Populus nigra

PEG 4000 g/mol (Ψ= -0.35 MPa)

Royer et al, 2016 J Exp Bot

(27)

 root growth rate

Rapid impairment of cell division & cell expansion

26

*** ***

0.0 0.2 0.4 0.6 0.8 1.0 1.2

Root growth rate (mm h-1)

before 0.5 h 3 h

0 1 2 3 4 5 6

Growth zone length (mm)

*

*** *** ***

0.0 0.1 0.2 0.3

EERmax(h-1)

0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4

DZ length (mm)

** **

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0

Cell production rate (cell h- 1)

 cell production rate (cell division rate)

 growth zone length  EERmax

 division zone length

osmotic stress

(28)

1/2h 3h

CTLPEG

osmotic stress

Division

zone Elongation zone 4.5 mm 1.5mm

EZ-CTL-3h/4EZ-CTL-3h/1EZ-CTL-3h/3EZ-CTL-0.5h/1EZ-PEG-0.5h/1EZ-PEG-0.5h/2EZ-PEG-0.5h/3EZ-CTL-0.5h/4EZ-CTL-0.5h/2EZ-CTL-0.5h/3EZ-CTL-3h/2EZ-PEG-0.5h/4EZ-PEG-3h/3EZ-PEG-3h/4EZ-PEG-3h/2EZ-PEG-3h/1DZ-PEG-0.5h/4DZ-CTL-0.5h/4DZ-CTL-3h/4DZ-CTL-3h/3DZ-CTL-0.5h/3DZ-CTL-0.5h/1DZ-PEG-0.5h/1DZ-PEG-0.5h/2DZ-PEG-0.5h/3DZ-CTL-3h/2DZ-CTL-0.5h/2DZ-CTL-3h/1DZ-PEG-3h/4DZ-PEG-3h/3DZ-PEG-3h/2DZ-PEG-3h/1 Height0.00.1

32 Librairies & RNA sequencing

30 million paired-end reads 100 bp, HiSeq 2500

Dynamics of molecular controls

 Expression patterns are spatially structured distinct cell process ⇔ distinct genes & GO

Elongation zone Division zone

27

(29)

Division

zone Elongation zone 4.5 mm 1.5mm

Dynamics of molecular controls

1/2h 3h

CTLPEG

osmotic stress

28

Elongation zone Division zone

 Minor changes after 0.5h

 Strong remodelling after 3h

-> molecular response took time to fully deploy

3h PEG 3h PEG

0.5h PEG & CTL 0.5h PEG & CTL

EZ-CTL-3h/4EZ-CTL-3h/1EZ-CTL-3h/3EZ-CTL-0.5h/1EZ-PEG-0.5h/1EZ-PEG-0.5h/2EZ-PEG-0.5h/3EZ-CTL-0.5h/4EZ-CTL-0.5h/2EZ-CTL-0.5h/3EZ-CTL-3h/2EZ-PEG-0.5h/4EZ-PEG-3h/3EZ-PEG-3h/4EZ-PEG-3h/2EZ-PEG-3h/1DZ-PEG-0.5h/4DZ-CTL-0.5h/4DZ-CTL-3h/4DZ-CTL-3h/3DZ-CTL-0.5h/3DZ-CTL-0.5h/1DZ-PEG-0.5h/1DZ-PEG-0.5h/2DZ-PEG-0.5h/3DZ-CTL-3h/2DZ-CTL-0.5h/2DZ-CTL-3h/1DZ-PEG-3h/4DZ-PEG-3h/3DZ-PEG-3h/2DZ-PEG-3h/1 Height0.00.1

(30)

Interferences between growth effectors and hormonal pathways

29

GROWTH

 EERmax

 growth zone length

 cell production rate

 cell division rate

osmotic stress

 No regulation of core cell cycle genes - low regulation level?

- impact of expansive growth in DZ ?

- post-transcriptional regulation of cell cycle components ? (Skiryczet al., 2011)

TR ANSC RIP TO M E re m od el lin g & GO en ric hme nt s

^ Activation of regulatory processes with a strong emphasis on

genes related to hormone pathways (up to 20 % of DEGs)

Brassinosteroids (-) Jasmonates (+)

Auxin (+)Abscissic Acid (+)Ethylene (+)

Gibberellins (-) Cytokinins (+/-) Salicylic acid (+)

Modification of biosynthesis, catabolism, signaling cascade & transport

Control of hormonal status &

local hormone maxima

 Promotion of growth effectors suggesting the facilitation of cell expansion

• cell wall-strengthening XTHs(-)

• cell wall-loosening EXPs(EXPA1and EXPA8) (+)

• cell wall-loosening PECTINE LYASEs(+)

• Membrane hydraulic conductivity : aquaporins(+)

retroaction ?

(31)

Dynamics of root growth in response to mechanical resistance

30

What is the growth response of a root encoutering an obstacle?

• force applied by the root

• growth rate

• curvature

Jin et al. (2013)

Croser et al. (1999)

• Root growth is sensitive to soil impedance

• Reduction of cell proliferation and cell expansion rates

(32)

3D kinematics

31

root

Root glass blade

• Adventitious root ( P. euramericana cv Soligo)

• Hydroponic culture

• Obstacle = home-made submersible sensor of force

force = k * deformation

• Time-lapse photography from two orthogonal directions -> 3D kinematics

Bizet et al, 2016 J Exp Bot

(33)

Dynamics of the response

32

Bending

critical force

 Several consecutive phases - « touch » response

- growth recovery and force build-up

- bending of the root & the critical force is reached - after bending : the root continues to grow with a

slower rate

(34)

Dynamics of cell expansion

33 3D reconstruction is necessary because

buckling does not occur in a 2D plan

 shortening of the growth zone

 tissue contraction at the junction between elongation zone and mature zone, where curvature was maximal

= zone of mechanical weakness

 after buckling : lower cell expansion rate but longer elongation zone EER (h^-1)

(35)

Is root bending a purely mechanical phenomena?

34

beam theory

• root dimensions

• Young’s elastic modulus (32 ± 5 MPa)

 Bending was predicted from root mechanical properties

 No biologically mediated accommodation to mechanical forces influenced bending during this short period of time. Bending was purely mechanical = BUCKLING

(36)

Bracing root increases the applied force

35

 Lateral bracing of the root increased the force applied by the growing root by preventing buckling

(37)

Thank you for your attention

Irène Hummel David Cohen Nathalie Aubry Cyril Buré François Bizet Mathilde Royer Chvan Youssef Thibault Gaillot

Simone Scalabrin Federica Cattonaro Vera Vendramin

Lionel Dupuy Glyn Bengough

36

Root growth, cell processes and responses to environment

Root growth, cell processes and responses to environment

Root growth = expansion of material

Cell length : 15 µm  320 µm : x 20!

Cell elongation

The expanding cell

cell wall tonoplast plasmalemma

cell wall extensibility

water

hydrostatique pressure of the cell sap on the cell wall

Π

P

Ψ

• Cell wall deformation and synthesis

• Water and solutes inflow

Water flow into cell

• Le potentiel hydrique de la cellule : Ψ

= P

- Π

• A l’équilibre

– ( ∆ P - σ ∆Π ) = 0  J

= 0

– P

≈ 0, P

= σ ∆Π

• Si déséquilibre de statut hydrique : flux directionnel – J

= Lp • ( ∆ P - σ ∆Π ) = Lp • (P

- σ ∆Π )

P

Π

Ψ

= P

- Π

≈ - Π

eau

Cell wall relaxation reduces turgor pressure

Cell wall relaxation : the basis of cell expansion

augmentation du volume cellulaire

RELACHEMENT DES PAROIS

entrée d ’eau et de solutés

baisse de P et donc de Ψ

∆Ψ

augmente rétablissement de P

arrêt de croissance

retour ∆Ψ à 0

(Ψ

= P - Π

)

J

≈ Lp • ( ∆ P - σ ∆Π )

Growth rate depends linearly on turgor pressure

Growth Rate = φ (P - Y)

φ Y

La croissance est linéairement proportionnelle à la pression de turgescence

(le moteur de la croissance), au delà d’un seuil minimum

 Turgor pressure P

 Wall tension σ

𝜎𝜎 𝑧𝑧𝑧𝑧 P

𝜎𝜎 𝜃𝜃𝜃𝜃

t R

𝜎𝜎

= 𝑅𝑅

2𝑡𝑡 𝑃𝑃 𝜎𝜎

= 𝑅𝑅

𝑡𝑡 𝑃𝑃 = 2𝜎𝜎

Wall tension and anisotropic growth

𝜎𝜎

P

Cell expansion parameters

• Force motrice du grandissement cellulaire :

– Pression de turgescence

• Contrôle du grandissement cellulaire :

– extensibilité des parois ( ø , Y )

– conductivité hydraulique volumique cellulaire (-> Lp) – Disponibilité en solutés / en eau

Π

Ψ

Ψ

= P

- Π

A biophysical model of cell expansion : Lockhart (1965)

• Le potentiel hydrique de la cellule : ^Ψ

𝜎𝜎 _{𝑧𝑧𝑧𝑧} P

𝜎𝜎 _{𝜃𝜃𝜃𝜃}

_{= P}

_- Π

• _{P et} Π ± constant le long de la zone de croissance et au delà