Quantitative analysis of lateral root morphogenesis in Arabidopsis thaliana Alexis Maizel Plant development is continuous Daniel von Wangenheim Arabidopsis thaliana A. Maizel l Montpellier 27-03-14 2 Roots: the hidden half Hodge A. The plastic plant: root responses to heterogeneous supplies of nutrients. New Phytologist. 2004. A. Maizel l Montpellier 27-03-14 3 Lateral root formation in Arabidopsis Lateral root emergence Development Differentiation zone Lateral root initiation and growth Initiation zone Basal meristem Lateral root pre-initiation Apical meristem A. Maizel l Montpellier 27-03-14 4 Plant embryonic development A. Maizel l Montpellier 27-03-14 5 Lateral root formation in Arabidopsis Lateral root emergence Differentiation zone Development Capture and quantify the dynamics of lateral root morphogenesis Lateral root initiation and growth Initiation zone Basal meristem Lateral root pre-initiation Apical meristem A. Maizel l Montpellier 27-03-14 6 Light-sheet fluorescence microscopy Illumination Detection from: J. Huisken, D. Y. Stainier, Development 136, 1963-75 (2009). A. Maizel l Montpellier 27-03-14 7 Digital Scanned Light Sheet Microscope camera (1392x1040) filterwheel Top Illumination lens Fr Imaging chamber on t Detection lens Top view stage x/z/y/psi Laser lines entry A. Maizel l Montpellier 27-03-14 Front view 8 Digital Scanned Light Sheet Microscope Illumination lens Detection lens side Front Front Side Capillary Carbon rod ½ MS medium 0.5% phytagel A. Maizel l Montpellier 27-03-14 9 Growth conditions during imaging Light source (off during scanning) Perfusion system provides fresh medium Roots are in the medium, leaves remain in the air Plant grows in an upright position A. Maizel l Montpellier 27-03-14 Daniel von Wangenheim 10 Growth conditions during imaging A. Maizel l Montpellier 27-03-14 Daniel von Wangenheim Dynamics of root development Growing root tip observed for: 23min MAP4-GFP Arabidopsis primary root tip recording time 15 h, 7 min interval H2B-RFP Lti6-GFP (Plasma mbrne) 10µm A. Maizel l Montpellier 27-03-14 12 Maizel et al. (2011) The Plant Journal, 68(2), 377–385. Morphodynamics of lateral root formation Light sheet microscopy A. Maizel l Montpellier 27-03-14 Daniel von Wangenheim 13 Pericycle swelling prior to the first division Pericycle Endodermis Pericycle Endodermis Daniel von Wangenheim 3.58 h 4.83 h 1.6 1.6 0h 3.58 h Normalized area 0h 1.4 1.4 1.2 1.2 11 0.8 0.8 0.6 0.6 00 side view 11 22 33 44 Time Time [h] [h] 55 radial view J. Vermeer et al., Science 2014. A. Maizel l Montpellier 27-03-14 18 The shape of LRP is highly stereotypical A. Maizel l Montpellier 27-03-14 Lucas M et al. (2013) Proc Natl Acad Sci USA. 25 Patterns of divisions side view front view Anticlinal Periclinal A. Maizel l Montpellier 27-03-14 Radial 28 Division patterns in cycles 1 & 2 Master cell file Anticlinal Anticlinal Periclinal Anticlinal A. Maizel l Montpellier 27-03-14 Daniel von Wangenheim 29 aurora kinase mutants have random planes of divisions wild type aur1-2, 2-2 Stage I Stage II Stage III Stage IV A. Maizel l Montpellier 27-03-14 Lucas M. et al. (2013) Proc Natl Acad Sci USA., 110(13), 5229–5234. 35 Growth dynamics of aurora mutant is affected Epidermis Cortex Endodermis A. Maizel l Montpellier 27-03-14 Lucas M et al. (2013) Proc Natl Acad Sci USA. 37 Importance of the overlying tissues 0 I II III IV V VI VII Emergence adapted from: Malamy, J. E., and Benfey, P. N. (1997). Development 124, 33–44. A. Maizel l Montpellier 27-03-14 38 Auxin signaling in endodermis is important for initiation Pericycle Endodermis pCASP1::shy2-2 shy2-2: dominant negative blocker of auxin signaling Col-0 CASP1::shy2-2 shy2-101 J. Vermeer et al., Science (2014). A. Maizel l Montpellier 27-03-14 Joop Vermeer (N. Geldner lab) 39 Overlaying tissue determine the shape of the primordium J0361>>axr3-GR (inducible inhibition of auxin response) -DEX +DEX -DEX +DEX IV V III A. Maizel l Montpellier 27-03-14 Lucas M. et al. (2013) Proc Natl Acad Sci USA., 110(13), 5229–5234. 40 Summary Morphogenesis of lateral root primordium is not underpinned by rigid initial organisation The lateral root primordium shape, cell division pattern and growth dynamics are emerging properties Canalization by surrounding tissue Oriented division Anisotropic growth A. Maizel l Montpellier 27-03-14 42 Lab members Jens Fangerau Mouli Ghosh-Dastidar Ira Mägele Amaya Vilches-Barro Daniel von Wangenheim Virginie Jouannet Ana Beatriz Moreno Eva Stauffer Collaborators Malcolm J. Bennett (Nottingham university) N. Geldner & J.Vermeer (Lausanne university) Jürgen Kleine-Vehn (BOKU Vienna) Heike Leitte (Heidelberg university) Steffen Lemke (Heidelberg university) Ernst H.K. Stelzer (Francfort university) Martin Crespi (CNRS) Hervé Vaucheret (INRA) Funding A. Maizel l Montpellier 27-03-14 43
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