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Origin and Genetics of Kranz Anatomy
and C4 Anatomical Specialization
Transferring C4 into C3 crops
• Modeling predicts that it could eventually boost
yields by 50% (Sage and Zhu (2011) JXB )
• Engineering specific architecture and cell-cell
interactions is considered the major challenge
Thomas L. Slewinski
Emerging Leader in Science
• Despite 30+ years of research - No known genes
that function in Kranz anatomy have been described
Monsanto Company
1/11/2014
C4 Photosynthesis
C3
Basics of Kranz anatomy
- Reduction in Photorespiration
- Exclude O2 from Rubisco
- Saturates Rubisco with CO2 so it can
achieve maximal catalytic efficiency
- C4 plants use water and nitrogen more
efficiently
C4
-Increases the Radiation Use Efficiency
- More productive per unit area!
Single cell layer
that surrounds
the vascular
core
BS cells
preferentially
accumulate
starch
BS cells are
suberized
Increased vein density (only 2 M
cells between veins)
Reduced M size
V-BS-M-M-BS-V
Langdale (2011) Plant Cell 11 3879-3892
Basics of Kranz anatomy
Formation of minor veins – PIN-YFP expression
Dimorphic
Chloroplasts
Specialized
Plasmodesmata at
the M-BS
interface
BS
M
Low PS II in
BS
chloroplasts
BS cells form clonal patches
along the length of the vein
BS cells form before the internal
Vascular core differentiates
Slewinski et al (2012) PCP
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Evolution of C4 in plants
Esau - Plant Anatomy
• Kranz-type C4 evolved at least 70 times
independently in both dicots and monocots
Sage et al (2011) JXB
• One of the most remarkable examples of
convergent evolution
• Evolves “Fully Formed” – little evidence for slow
evolutionary progress towards Kranz-type C4
(Langdale (2011) Plant Cell 11 3879-3892)
Kranz-type C4 is derived from a conserved
tissue found in angiosperms
The endodermis is primed with C4
metabolism
• Hypothesis – Kranz anatomy and C4 photosynthesis are derived from the
projection of endodermal identity onto the bundle sheath within the
photosynthetic leaf tissue. This results in a synergistic interaction between
photosynthetic cells and the endodermis
•
Would explain how C4 always arises “fully formed”
•
Starch preferentially accumulates in the BS – because it is adapted from the
“Starch Sheath” in the stem and petiole
Is it possible that BS plastids are a “photosynthetic amyloplasts” ?
•
Suberin synthesis is also characteristic of the endodermis
•
In roots, stems, and petioles the endodermis is an Auxin conducting tissue
(PIN expression)
– May explain why there is a shift in vein density?
Hypothesis for the evolution of Kranz-type C4
Endodermal program projects into the
leaf blade - giving rise to Kranz anatomy
and preconditioning the metabolism for
C4
Kranz-type C4 is a synergistic interaction between
photosynthetic cells and the endodermis
Leaf identity projects the photosynthetic
program into the endodermis
Photosynthetic
cell
(mesophyll)
Vascular tissue
C3 Bundle sheath
Endodermis/
Starch sheath
Endodermis/
Starch sheath
Slewinski TL. (2013) Front Plant Sci. 4:212.
Endodermis
(bundle sheath)
Vascular
core
The endodermal cells project a cortex-like identity onto the
mesopyll/photosynthetic cells (CO2 metabolic shuffling)
Projecting the endodermal program into C3 leaves may be one way to engineer C4
Or at least precondition the leaf for the C4 program
Slewinski TL. (2013) Front Plant Sci. 4:212.
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Formation of minor veins – PIN-YFP expression
Gene candidates based on endodermis formation in
the roots
Endodermis
Stele
Cortex
Stele
Endodermis
JKD, MGP
(IDD-likes)
SCR
SHR
Development of the minor
vein mirrors root formation
Cortex
JKD
(ID-likes)
SHR
-- SCARECROW (SCR)
is a primary regulator
of endodermis
development and
identity
Ogsawara et al 2011 PMB
Helariutta et al
(2000)
Helariutta et al (2000)
Slewinski et al (2012) Plant Cell Phys ; Slewinski et al submitted
ZmScarecrow Expression
zmscr mutants produce extra BS cell files
in leaves
zmscr
Li et al (2010) Nature Genetics
IKI stained
m
mutant alleles
m2
ZmScarecrow
GRMZM2G131516
m1
Exon 1
Exon 2
Slewinski et al (2012) Plant Cell Phys
Slewinski et al (2012) Plant Cell Phys
zmscr mutants have defective minor veins
Wild type
IKI stained
Veins in the scr mutant leaves frequently collide
IKI stained
zmscr
Wild type
zmscr
zmscr
zmscr
BS cells merge into a continuous structure that displaces mesophyll cells
Slewinski et al (2012) Plant Cell Phys
Slewinski et al (2012) Plant Cell Phys
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ZmSCR regulates BS formation and identity
IKI stained
Wild type
Disruption of ZmSCR results in undifferentiated BS cells
zmscr
zmscr
zmscr
Undifferentiated plastids
In BS
zmscr
zmscr
Altered Plasmodesmata
Slewinski et al (2012) Plant Cell Phys
Slewinski et al (2012) Plant Cell Phys
zmshortroot1 mutant
Gene candidates based on endodermis formation in
the roots
Endodermis
Stele
Mutant allele of ZmSHR1
Cortex
Stele
Endodermis
JKD, MGP
(IDD-likes)
SCR
SHR
Cortex
JKD
ZmSHR1 expression
(ID-likes)
Li et al, Nature Genetics, 2010
WT
SHR
IKI stained
zmshr1
WT
zmshr1
Ogsawara et al 2011 PMB
-- What role does SHORTROOT (SHR)
play in Kranz anatomy and C4
physiology?
Slewinski et al. Submitted
Altered M cells in the zmshr1 mutant
Incomplete Kranz anatomy in the zmshr1 mutant
zmshr1
WT
zmshr1
WT
zmshr1
zmshr1
Slewinski et al. Submitted
Slewinski et al. Submitted
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BS without veins in the zmshr1 mutant
zmshr1
zmshr1
Formation of minor veins – PIN-YFP expression in developing maize leaf
zmshr1
Reprinted from Slewinski et al. (2012) Plant and Cell Physiology
zmshr1
Model of minor vein formation in developing WT leaves based on PIN-YFP expression
zmshr1
zmshr1
BS cells (yellow) form clonal patches along the
length of the vein
BS cells (yellow) form before the internal
Vascular core (blue) initiates and differentiates
In developing minor veins in leaves of the zmshr1 mutant – the vascular core fails to initiate leaving a track of cells that have the
default endodermis (external) identity. Distinctive cells arise from the same underlying developmental pathway as the BS
endodermis – just a modification of the SHR signaling pathway
Slewinski et al. Submitted
Arundinella hirta
“distinctive cells”
= vascular core cell (s)
= Bundle Sheath cell
Crookston and Moss (1973) Plant Physiology
= vascular founder cell
= Cell differentiation/division zone
Slewinski et al. Submitted
The Phyllode Theory and grass leaf development
Genetic regulation of Kranz anatomy
Reduction or Loss of leaf blade
Extrapolation of the petiole/lower
leaf blade in to a new
blade-like structure
SHR1
IDD ??
SCR
SCR-like?
IDD?
??
SHR1
IDD ??
Slewinski TL. (2013) Front Plant Sci. 4:212.
The Phyllode Theory and grass leaf
development
Early monocots
Morphological shift that led to the grasses also
preconditioned Kranz-type C4
Maize
Photosynthetic cells
Endodermis becomes photosynthetic BS - C4 metabolism
Selection for C4
Grasses/preconditioning event
Non-photosynthetic
parenchyma cells
Endodermis/
Starch sheath
Disruption in the
endodermal program
Grass leaves
??????
Rice
No selection for C4
Endodermis/Starch sheath incorporates into the photosynthetic tissue
- Possibly why all C4 grasses are Kranz-type?
Slewinski TL. (2013) Front Plant Sci. 4:212.
Slewinski TL. (2013)
Front Plant Sci. 4:212.
Endodermis becomes vascular BS – C3 metabolism
Mestome sheath may be a remnant of the pericycle tissue
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Acknowledgements
Turgeon Lab
Dr. Robert Turgeon
Dr. Cankui Zhang
Alyssa Anderson
Dr. Andre Jagendorf
Collaborators
NSF C4 Group
Dr. Thomas Brutnell
- Kevin Ahern
- Dr. Lin Wang
Richard Medville
-TEM
Dr. Adrienne Roeder
-Confocal
Funding
NSF Plant Genome
USDA-NIFA Fellowship
-Dr. Joe Colasanti
-Indeterminate genes
-Dr. Kimberly Gallagher
-Shortroot
Monsanto
Dr. Alice Barkan
- Mu Illumina project
Uniform Mu Project
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