Tapping into the mechanical link of expansins in leaf...
Transcript of Tapping into the mechanical link of expansins in leaf...
Hoe -Han Goh†, Nic Mullin‡, Alexis Peaucelle¤, Andreas Backhaus†, Jamie Hobbs‡ and Andrew Fleming†
†Dept. Animal and Plant Sciences, University Sheffield, UK, ‡Dept. Physics and Astronomy, University of Sheffield, UK, ¤INRA Versailles, Versailles, France
Introduction
Hypotheses
Aims:
References
The use of AFM for in vivo quantification of plant cell wall extensibility
Plant development relies largely on cell expansion. Cell wall looseningis a pre-requisite for cell expansion in which expansins, a conservedfamily of non-enzymatic proteins, play a major role1,2. Ectopicexpression of expansin at meristem and leaf flank has been used toinduce leaf initiation and aberrant leaf shape3,4. However, to whatextent this reflects an in vivo function for expansins remains unclear.Indeed, the role of cell wall mechanics in controlling morphogenesisremains debatable.
Atomic force microscopy can be used to characterise the
mechanical properties of cell wall during early leaf growth in
tobacco and Arabidopsis
• Differences in cell wall extensibility can be measured and mapped using
AFM with great resolution showing spatial heterogeneity of extensibility at
cellular and tissue levels.
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1. The cell wall protein expansins function in differential growth during leaf development
2.The differential regulation of expansin expression leads to organ-level coordination of leaf size and shape
alterations of plant cell wall properties modulate leaf growth
ModelPlant tissue can be modelled as growing network of viscoelastic springs (cell walls) with strain force generated by internal turgor pressure to explore general cellular behaviour during leaf development.
Cell wall is more extensible at leaf abaxial side (ab)
1. Functional analysis of expansins involve in leaf morphogenesis
2. In vivo quantification of expansin activity
3.Utilisation of a leaf growth model based on viscoelastic nature of plant cell wall
in testing various hypotheses regarding mechanical force and morphogenesis
• To determine the role of expansins in leaf morphogenesis
• To determine the role of plant cell wall in controlling leaf growth and form
Project outline:
1Cosgrove, D. J. 2000 Loosening of plant cell walls by expansins. Nature 407, 321-326.2Sampedro, J. & Cosgrove, D. J. 2005 The expansin superfamily. Genome Biology 6, 242-250.3Fleming, A. J., McQueen-Mason, S., Mandel, T. & Kuhlemeier, C. 1997 Induction of leaf primordia by the cell wall protein expansin. Science 276, 1415-1418.4Pien, S., Wyrzykowska, J., McQueen-Mason, S., Smart, C. & Fleming, A. 2001 Local expression of expansin induces the entire process of leaf development and modifies leaf shape. Proceedings of the National Academy of Sciences of the United States of America 98, 11812-118175Donnelly, P. M., Bonetta, D., Tsukaya, H., Dengler, R. E. & Dengler, N. G. 1999 Cell cycling and cell enlargement in developing leaves of Arabidopsis. Developmental Biology 215, 407-419.
Future plan:
Leaf staging for expansin expression study
• Identifying expansins involve in leaf development from expression studies
• Testing the function of expansin through inducible amiRNA silencing approach and
viscoelastic leaf growth model
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t (m
m)
Day after sowing
Petiole
Lamina
Width
Length
±1SE
5mm
5mm
Cantilever
deflection, s
Cantilever spring
constant, k
Force, F F = ks
Schematic diagram of the vertical tip movement during the approach and retract phases of a force spectroscopy measurement
Approach Retract
Jump to contact AdhesionRepulsive contact
Separation Separation
Mechanisms of atomic force microscopy (AFM)
50µm
100µm
200µm0.5mm
1mm
Tobacco primodia
Measurements on leaf number 6 of Arabidopsis thaliana Col-0 show that leaf development can be categorised into three phases: cell division predominant stage, cell expansion stage and mature stage.
Aniline blue staining
of callose which
formed during cell
division indicating
planes of cell division
*Occurrences of cell division are indicated by GUS staining
Scale bars = 100µm
Cantilever deflection
Quadrant photodiode
laser detector
Laser emission
Sample
Piezo
elementZ
Z
Type I
Type II
Electrical
current (V)
, Z
(hysteresis caused by viscous behaviour of the leaf surface)
Topography of mapped surface Force measurement
Height
Force
Height
Force
Day 12Day 14
Day 16