03 Miller Soil Physics meeting.pdf

New measurement methods in soil physics: future frontiers
How soil physical conditions affect root
development and nitrate uptake
Tony Miller, Nick Chapman, Keith Lindsey, Richard Whalley

Plant biologist’s root methods
• Hydroponically grown plants
• Agar Petri dish
• Sand culture
• Soil

ROOT PROLIFERATION AND TRANSPORTERS
> Root architecture > Nutrient distribution
> Transporter activity > Soil strength
MASS FLOW AND DIFFUSION
> Water uptake > Hydraulic conductivity
> Net ion uptake > Water potential
> Plant demand > Ion mobility
> Diffusion gradient
Root nutrient acquisition
Plant traits and soil physical properties
LOW
HIGH
KEY
Water movement
Passive water uptake
Active water uptake
Nutrient ion uptake
Nutrient ion efflux
Substrate particle
(Chapman et al., TiPS, 2012 in press)

Root nutrient acquisition
Soil physical properties
(Chapman et al. 2012, TiPS, submitted)

Arabidopsis root nutrient acquisition in sand rhizotrons
Experimental approach
Novel sand rhizotron system
aspirator
h, cm
rhizotron
funnel
(Chapman et al. 2011, PC&E)

Arabidopsis root nutrient acquisition
Physical characterisation of the root growth environment
Water release characteristic
sand
graduated
burette
L, cm
h, cm
Q, cm/s
water
sand
funnel
Saturated hydraulic conductivity
h, cm

Water content (cm 3 cm -3 )
5.00
4.50
4.00
3.50
3.00
2.50
2.00
1.50
1.00
0.50
0.00
(Chapman et al. 2011, PC&E)
Arabidopsis root nutrient acquisition
Physical characterisation of the root growth environment
k (m/d-1)
3.50
3.00
2.50
Redhill T (measured)
Redhill T (modelled)
425 um (modelled)
2.00
1 10 100 1000
1.50
Matric potential (-hPa)
1.00
0.50
0.00
1.00E-01
Redhill T 425 µm
Relative hydraulic conductivity
1.00E-03
1.00E-05
1.00E-07
1.00E-09
1.00E-11
Matric potential (-hPa)
1 10 100 1000
Redhill T
425 um

Sand
Arabidopsis root nutrient acquisition
Physical characterisation of the root growth environment
Water potential
(MPa)
Matric potential
(kPa)
Measured K sat
(m.d -1 )
Calculated K unsat
(m.d -1 )
Redhill T -0.11 -1.5 - 1.35
Redhill T -0.33 -3.0 1.81 0.79
Redhill T -0.40 -4.5 - 0.18
425 μm -0.33 -3.0 3.23 0.17

Primary root length (PRL)
Lateral root density (LRD)
Arabidopsis root nutrient acquisition
Root proliferation
PRL (mm)
LRD
30.0
25.0
20.0
15.0
10.0
5.0
0.0
0.20
0.16
0.12
0.08
0.04
0.00
**
30.0
25.0
20.0
15.0
10.0
5.0
0.0
0.20
0.16
0.12
0.08
0.04
0.00
*
*
30.0
25.0
20.0
15.0
10.0
5.0
0.0
0.20
0.16
0.12
0.08
0.04
0.00
**
**
(Chapman et al. 2011, PC&E)

Arabidopsis root nutrient acquisition
Relating root proliferation to physical properties of the sand
PRL response conserved response to water potential and hydraulic
conductivity
Lateral root growth strongest relationship with hydraulic conductivity
Percentage of roots with laterals
80
75
70
65
60
55
50
K unsat (m.d -1 )
r 2 = 0.93
45
0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6
30.0
25.0
20.0
15.0
10.0
5.0
0.0
PRL (mm)
*
30.0
25.0
20.0
15.0
10.0
5.0
0.0
PRL (mm)
**
(Chapman et al. 2011, PC&E)

Root proliferation
Arabidopsis root nutrient acquisition
Root proliferation conclusions
Primary root length increases with decreasing water supply and this is
independent of nitrate concentration at the root surface
Lateral root number, length and density increase with increasing nitrate
concentration but decrease with decreasing water supply
Root proliferation and soil physical characteristics
Primary root length increases with decreasing hydraulic conductivity and
increasingly negative water potential
Lateral root number, length and density increase with nitrate
concentration but also increased hydraulic conductivity and less negative
water potential

Localisation of
NRT (nitrate) and
AMT (ammonium)
uptake genes in
Arabidopsis
Shoot
NO3 -
NO 3 -
NO 3 - NO3 -
?
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
?
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NRT1.1 NRT1.2; NRT2.1; CLC; NRT1.5; NAXT1;
Phloem; Xylem; Vacuole; Cells; Soil;
Modified from Masclaux-Daubresse et al. 2010

Root nutrient acquisition
Root transporters
ROOT BULK
NO 3 -
NO 3 -
NO 3 -
NO 3 -
LOW
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
[NO 3 - ]
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
NO 3 -
HIGH

Arabidopsis root nutrient acquisition
Root transporters
Do soil physical changes influence root transporters?
AtNRT1.1 – dual-affinity nitrate uptake (and sensor)
AtNRT2.1 – high-affinity nitrate uptake
AtPIP2.2 – plasma membrane root aquaporin (drought response)
AtPIN1 – basipetal vascular auxin efflux (root elongation)
AtPIN2 – acropetal epidermal and basipetal cortical auxin efflux (lateral
root growth)

Using Arabidopsis to understand root nutrient acquisition
Gene expression
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
*
AtPIP2.2
*
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
0.100 AtPIP2.2
0.080
0.060
0.040
0.020
0.000
*
0.100
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
AtNRT2.1
*
AtPIP2.2
*

Using Arabidopsis to understand root nutrient acquisition
Gene expression
0.100
0.080
0.060
0.040
0.020
0.000
0.080
0.060
0.040
0.020
0.000
AtNRT2.1
*
0.100 AtPIP2.2
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
*
AtPIP2.2
*
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
0.100 AtPIP2.2
0.080
0.060
0.040
0.020
0.000
*
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
*
AtPIP2.2
*

Arabidopsis root nutrient acquisition
Root transporters tagged with GFP (green fluorescent protein)
16x 50x 256x

0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
*
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
*
Arabidopsis root nutrient acquisition
Root transporters
proAtNRT2.1:eGFP line was a gift to AJM from Kiba Takotoshi (Riken Institute, Japan)
-1.5 kPa -4.5kPa WT
425 µm WT

Arabidopsis root nutrient acquisition
Root transporters – using mutant plants lacking specific genes
35.0 PRL (mm)
30.0
25.0
20.0
15.0
10.0
5.0
0.0
7.0
6.0
5.0
4.0
3.0
2.0
1.0
0.0
*
*****
TLRL (mm) *****
****
2.5 LRN
2.0
1.5
1.0
0.5
0.0
0.14
0.12
0.10
0.08
0.06
0.04
0.02
0.00
*
*
*****
LRD *****
**
***

Root transporters
Arabidopsis root nutrient acquisition
Root transporter conclusions
AtNRT2.1 and AtPIP2.2 expression is coordinated across nitrate and water
treatments
atnrt2.1, atnar2.1 and atpip2.2 mutant lines lose primary root length
response to water supply
atnar2.1 and atpip2.2 (but not atnrt2.1) mutant lines lose lateral root
length response to water supply
Root transporters and soil physical characteristics
AtNRT2.1 and AtPIP2.2 expression strongly increases with an increase in
matric potential

Arabidopsis root nutrient acquisition
A ‘get-whilst-the-going-is-good’ conceptual model

Arabidopsis root nutrient acquisition
A ‘get-whilst-the-going-is-good’ conceptual model

Arabidopsis root nutrient acquisition
General conclusions
A ‘get-whilst-going-is-good’ strategy for nutrient acquisition is effective for a
weed species such as Arabidopsis
Understanding nutrient acquisition by weed species could help to exploit
areas with more patchy/poor nutrient supply
It is possible to experimentally combine advanced plant science tools AND
soil physical characteristics in the study of root nutrient acquisition
Can build a more complete picture of root nutrient acquisition by working
together

How soil physical conditions affect root development and
nitrate uptake
Acknowledgments
Richard
Whalley
Colin Webster
Chris Watts
Andy Gregory
Nick Chapman
Susan Smith
Yi Chen
Statistics Bio-imaging VCU
Steve Powers Allison van der Meene Nicky Seymour
Kirstie Halsey Lynda Castle
Keith Lindsey
Jen Topping

Arabidopsis root nutrient acquisition
Root proliferation and ion-selective microelectrodes
RT-2mm
RT

Arabidopsis root nutrient acquisition
Root proliferation and ion-selective microelectrodes
-
[ NO3 ] mM
120
100
80
60
40
20
0
RT
RT-2mm
Average RT
Average RT-2mm

Using Arabidopsis to understand root nutrient acquisition
Explanatory multiple linear regression models
BRL
NRT1.1
HC KNO3 WP
MP
LRD
LRN
PRL TLRL TRL
NRT2.1 PIN1 PIN2 PIP2.2

Arabidopsis root nutrient acquisition
Experimental systems: are physical properties important?
Physical parameter Agar Sand Soil
Pore size Small Variable Variable
Pore distribution Regular Regular Irregular
Pore structure Limited Extensive Extensive
Hydraulic conductivity Low Decreases with increasing
particle size
Matric potential Increases with % Stronger with increasing
particle size
Decreases with increasing
particle size
Stronger with increasing
particle size
Strength Increases with % Decreases with saturation Decreases with saturation
Nutrient ion distribution Homogeneous Heterogeneous Heterogeneous

Results disparities
Arabidopsis root nutrient acquisition
Experimental systems: are physical properties important?
Differences between sand and agar for Arabidopsis root architecture
(Chapman et al. 2011) and root transporters (unpublished data)
Differences between soil and gels for barley root architecture (Hargreaves
et al. 2009 and Wojciechowski et al. 2009)
Differences between soil and sand for rice root architecture (Clark et al.
2002)
Differences between single stress systems and soil for mechanical effects
(Clark et al. 1998), water potential responses (Liang et al. 1997 and
Whalley et al. 1999), soil strength and water content (Whalley et al. 2008
and Whitmore et al. 2009)

Agar v sand
0.100
0.080
0.060
0.040
0.020
0.000
AtNRT1.1
3 KNO 3 concentrations
0.100 AtNRT1.1
0.080
0.060
0.040
0.020
0.000
*
Arabidopsis root nutrient acquisition
Root transporters
0.100
0.080
0.060
0.040
0.020
0.000
AtNRT2.1
*
0.100 AtNRT2.1
0.080
0.060
0.040
0.020
0.000
*
0.100
0.080
0.060
0.040
0.020
0.000
AtPIN1
*
0.100 AtPIN1
0.080
0.060
0.040
0.020
0.000
*
0.100
0.080
0.060
0.040
0.020
0.000
AtPIN2
0.10 AtPIN2
0.08
0.06
0.04
0.02
0.00
*
0.100
0.080
0.060
0.040
0.020
0.000
0.080
0.060
0.040
0.020
0.000
AtPIP2.2
0.100 AtPIP2.2
*

3 particle sizes
0.100
0.080
0.060
0.040
0.020
0.000
3 matric potentials
0.100
0.080
0.060
0.040
0.020
0.000
AtNRT1.1
*
AtNRT1.1
*
Arabidopsis root nutrient acquisition
Root transporters
0.100
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
*
AtNRT2.1
AtNRT2.1
*
0.100
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
AtPIN1
AtPIN1
0.100
0.080
0.060
0.040
* 0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
AtPIN2
*
AtPIN2
0.100
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
AtPIP2.2
AtPIP2.2
*

0.100 AtNRT1.1
0.080
0.060
0.040
0.020
0.000
0.100
0.080
0.060
0.040
0.020
0.000
*
AtNRT1.1
*
Arabidopsis root nutrient acquisition
Root transporters
425 µm WT
0.1 mM KNO 3 10.0 mM KNO 3 WT

0.080
0.060
0.040
0.020
0.000
Arabidopsis root nutrient acquisition
Root transporters
0.100 AtPIN2 -1.5 kPa -4.5kPa WT

Arabidopsis root nutrient acquisition
Root proliferation and ion-selective microelectrodes
Electrode output (mV)
- ] before and after (mM)
[NO 3
200
180
160
140
120
100
80
60
40
20
0
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32
20.0
10.0
0.0
RT RT-2mm
Time (mins)
30.0 [NO 3 - ] using before calibration (mM)
-
[NO3 ] using after calibration (mM)
Time (mins) vs Ref (mV)
RT RT-2mm
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32
Time (mins)
(Chapman et al. 2011, PC&E)