1) To determine the modifications of cellular processes and mechanisms caused by red light stimulation under environmental conditions of microgravity or fractional gravity in space. Our interest is focused on root meristematic cell growth and proliferation, two basic and essential cell processes which are fundamental for the plant developmental programme. Specifically, we want to study:
a) The effect of phytochrome activation by red light, as a potentially proliferative stimulus, on the compensation of the alterations in cell cycle and ribosome biogenesis caused by the gravitational stress;2) To investigate the effects of red-light-stimulation in the space environment on the alteration of auxin transport and perception, and of auxin distribution in the root.
b) The concerted action of light and gravity in controlling plant growth and development through effects on cell proliferation, especially focused in the root development.
Auxins are regulators and coordinators of meristematic cell growth and proliferation and are key factors affecting plant development.
3) To know the alterations in the phototropic behavior after red light stimulation, under different gravity conditions, of plants defective in essential components of the apparatus of auxin transport and/or perception, or lacking essential factors of cell growth and proliferation.
4) To detect gravity thresholds of the cellular effects of the gravitational stress, by means of the investigation of the relationship between light and gravity in fractional gravity. This investigation may also serve to approach a simulation of the gravity conditions in Moon and Mars.
1. To confirm and characterise the novel red light-dependent phototropic response in flowering plants;
2. To investigate the relationship between light and gravity by measuring thresholds in fractional gravity;
3. To determine whether the red-light-effect on blue-light-based phototropism is a direct or indirect effect;
- Run 1: 6 days duration, microgravityduring ISS Increment 41/42
- Run 2: 6 days duration, 0.1 g
- Run 3: 6 days duration, 0.3 g
- Run 4: 6 days duration, 1 g
- Run 1: 6 days duration, 0.5 gduring ISS Increment 47/48
- Run 2: 6 days duration, 0.8 g
- Run 3: 6 days duration, microgravity and 1 g
- Run 1: 6 days duration, microgravity and 1g(The use of a dedicated hardware (Fixation Box, FixBox for short) providing 3 LoCs is foreseen during the Seedling Growth-3 part to achieve the PFA (Paraformaldehyde) and GA (Glutaraldehyde) chemical fixation of the samples.)
- Run 2: 6 days duration, 0.3 g
1. Light and electron microscopy (parameters related to root meristematic cell growth and proliferation):
- Immunodetectionof nucleolin, RNA pol1 and cyclinB1
- Rate of local cell production
- Nucleolar size
- Nucleolar ultrastructure-proportion of granular component
- Number of cells per mm in the root meristem
2. Molecular Biology (RNA and Protein extraction)
- Global gene expression: microarray technique
- Expression of relevant genes (nucleolin, cyclins, auxin responsive elements): RT-PCR or qPCR
- Protein analysis: two-dimensional electrophoresis, proteomic techniques
- Definition with a precision higher than presently available of the factors involved, either as causes or effects, in the uncoupling of cell proliferation and cell growth which has previously described in root meristematic cells from plants grown in a microgravity environment.
- Determination of the role of the auxin polar transport in the cellular alterations caused by the altered gravity stress.
- Evaluation of the effect of a pulse of red light on counteracting the alterations in cell growth and proliferation caused by the gravitational stress.
- Estimation of the effects of fractional gravity on altering cell growth and proliferation parameters.
I. Matía, F. González-Camacho, R. Herranz, J.Z. Kiss, G. Gasset, J.J.W.A. van Loon, R. Marco, F.J. Medina, (2010), "Plant cell proliferation and growth are altered by microgravity conditions in spaceflight", Journal of Plant Physiology, 167, 3, pp. 184-193.
F.J. Medina, R. Herranz, (2010), "Microgravity environment uncouples cell growth and cell proliferation in root meristematic cells: the mediator role of auxin", Plant Signaling & Behavior, 5, 2, pp. 176-179.
F.J. Medina, (2010), "Sensing of gravity by plant cells and its effects on plant growth and development", in: Cell Mechanochemistry, J.J.W.A. van Loon, M. Monici, ISBN: 978-81-7895-458-5, pp. 97-112.
A.I. Manzano, I. Matía, F. González-Camacho, E. Carnero-Díaz, J.J.W.A. van Loon, C. Dijkstra, O. Larkin, P. Anthony, M.A. Davey, R. Marco, F.J. Medina, (2009), "Germination of Arabidopsis seed in space and in simulated microgravity: alterations in root cell growth and proliferation", Microgravity Science and Technology, 21, pp. 293-297.
I. Matía, J.J.W.A. van Loon, E. Carnero-Díaz, R. Marco, F.J. Medina, (2009), "Seed germination and seedling growth under simulated microgravity causes alterations in plant cell proliferation and ribosome biogenesis", Microgravity Science and Technology, 21, 2, pp. 169-174.
J. Sáez-Vásquez, F.J. Medina, (2008), "The Plant Nucleolus", Advances in Botanical Research, 47, J.C. Kader, M. Delseny, pp. 1-46.
M.L. Molas, J.Z. Kiss, (2008), "PKS1 plays a role in red-light-based positive phototropism in roots", Plant Cell and Environment, 31, pp. 842-849.
I. Matía, F. González-Camacho, R. Marco, J.Z. Kiss, G. Gasset, J.J.W.A. van Loon, F.J. Medina, (2007), "The "root" experiment of the "cervantes" spanish soyuz mission: Cell proliferation and nucleolar activity alterations in arabidopsis roots germinated in real or simulated microgravity", Microgravity Science and Technology, 19, 5-6, DOI: 10.1007/BF02919467, pp. 128-132.
F. Pontvianne, I. Matía, J. Douet, S. Tourmente, F.J. Medina, M. Echeverría, J. Sáez-Vásquez, (2007), "Characterization of AtNUC-L1 reveals a central role of nucleolin in nucleolus organization and silencing of AtNUC-L2 gene in Arabidopsis", Molecular Biology of the Cell, 18, pp. 369-379.
M. Sobol, F. González-Camacho, V. Rodríguez-Vilariño, E. Kordyum, F.J. Medina, (2006), "Subnucleolar location of fibrillarin and NopA64 in Lepidium sativum root meristematic cells is changed in altered gravity", Protoplasma, 228, pp. 209-219.
J.Z. Kiss, K.D.L. Millar, P. Kumar, R.E. Edelmann, M.J. Correll, (2011), "Improvements in the re-flight of spaceflight experiments on plant tropisms", Advances in Space Research, 47, pp. 545-552.
K.D.L. Millar, P. Kumar, M.J. Correll, J. Mullen, R.P. Hangarter, R.E. Edelmann, J.Z. Kiss, (2010), "A novel phototropic response to red light is revealed in microgravity", The New Phytologist, 186, pp. 648-656.
J.Z. Kiss, (2014), "Plant biology in reduced gravity on the Moon and Mars", Plant Biology, 16, S1, DOI: 10.1111/plb.12031, pp. 12-17.
J.Z. Kiss, G. Aanes, M. Schiefloe, L.H.F. Coelho, K.D.L. Millar, R.E. Edelmann, (2014), "Changes in operational procedures to improve spaceflight experiments in plant biology in the European Modular Cultivation System", Advances in Space Research, 53, 5, DOI: 10.1016/j.asr.2013.12.024, pp. 818-827.
J.P. Vandenbrink, J.Z. Kiss, R. Herranz, F.J. Medina, (2014), "Light and gravity signals synergize in modulating plant development", Frontiers in Plant Science, 5, DOI: 10.3389/fpls.2014.00563, pp. 563.
J.Z. Kiss, (2015), "Conducting plant experiments in space", Methods in Molecular Biology, 1309, DOI: 10.1007/978-1-4939-2697-8_19, pp. 255-283.
J.P. Vandenbrink, R. Herranz, F.J. Medina, R.E. Edelmann, J.Z. Kiss, (2016), "A novel blue-light phototropic response is revealed in roots of Arabidopsis thaliana in microgravity", Planta, 244, 6, DOI: 10.1007/s00425-016-2581-8, pp. 1201-1215.
J.P. Vandenbrink, J.Z. Kiss, (2016), "Space, the final frontier: a critical review of recent experiments performed in microgravity", Plant Science, 243, DOI: 10.1016/j.plantsci.2015.11.004, pp. 115-119.
(2018), "3.12 Seedling Growth 3", N-USOC - History, Achievements and Reflections 2003-2018, Centre for Interdisciplinary Research in Space (CIRiS) Trondheim, Norway, pp. 41-42.
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Figure 1: Schematic representation of the effects of gravity and light on the roots of developing plants and, specifically, on root meristematic cells, and the pending questions which are approached in this experiment. The main hypothesis to test in the project is whether or not the combined stimuli of altered gravity (microgravity) and modulated light (red light) produce effects on the auxin distribution in roots which may result in alterations of essential functions of meristematic cells, such as cell cycle and ribosome biogenesis.
Summary Overview Flow Diagrams: Seedling Growth-1
Seedling Growth-1 - Run #1
Summary Overview Flow Diagrams: Seedling Growth-2
Seedling Growth-2 - Run #1
Seedling Growth-2 - Run #2
Seedling Growth-2 - Run #3
Summary Overview Flow Diagrams: Seedling Growth-3
Seedling Growth-3 - Run #1