A Hypergravity Environment Induced by Centrifugation Alters Plant Cell Proliferation and Growth in an Opposite Way to Microgravity

Microgravity Science and Technology, Dec 2012

Seeds of Arabidopsis thaliana were exposed to hypergravity environments (2g and 6g) and germinated during centrifugation. Seedlings grew for 2 and 4 days before fixation. In all cases, comparisons were performed against an internal (subjected to rotational vibrations and other factors of the machine) and an external control at 1g. On seedlings grown in hypergravity the total length and the root length were measured. The cortical root meristematic cells were analyzed to investigate the alterations in cell proliferation, which were quantified by counting the number of cells per millimeter in the specific cell files, and cell growth, which were appraised through the rate of ribosome biogenesis, assessed by morphological and morphometrical parameters of the nucleolus. The expression of cyclin B1, a key regulator of entry in mitosis, was assessed by the use of a CYCB1:GUS genetic construction. The results showed significant differences in some of these parameters when comparing the 1g internal rotational control with the 1g external control, indicating that the machine by itself was a source of alterations. When the effect of hypergravity was isolated from other environmental factors, by comparing the experimental conditions with the rotational control, cell proliferation appeared depleted, cell growth was increased and there was an enhanced expression of cyclin B1. The functional meaning of these effects is that cell proliferation and cell growth, which are strictly associated functions under normal 1g ground conditions, are uncoupled under hypergravity. This uncoupling was also described by us in previous experiments as an effect of microgravity, but in an opposite way. Furthermore, root meristems appear thicker in hypergravity-treated than in control samples, which can be related to changes in the cell wall induced by altered gravity.

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A Hypergravity Environment Induced by Centrifugation Alters Plant Cell Proliferation and Growth in an Opposite Way to Microgravity

Ana I. Manzano 0 Ral Herranz 0 Jack J. W. A. van Loon 0 F. Javier Medina 0 0 J. J. W. A. van Loon Dutch Experiment Support Center (DESC) @ OCB-ACTA, VU-University Amsterdam , Amsterdam, Netherlands Seeds of Arabidopsis thaliana were exposed to hypergravity environments (2g and 6g) and germinated during centrifugation. Seedlings grew for 2 and 4 days before fixation. In all cases, comparisons were performed against an internal (subjected to rotational vibrations and other factors of the machine) and an external control at 1g. On seedlings grown in hypergravity the total length and the root length were measured. The cortical root meristematic cells were analyzed to investigate the alterations in cell proliferation, which were quantified by counting the number of cells per millimeter in the specific cell files, and cell growth, which were appraised through the rate of ribosome biogenesis, assessed by morphological and morphometrical parameters of the nucleolus. The expression of cyclin B1, a key regulator of entry in mitosis, was assessed by the use of a CYCB1:GUS genetic construction. The results showed significant differences in some of these parameters when comparing the 1g internal rotational control with the 1g external control, indicating that the machine by itself was a source of alterations. When the effect of hypergravity was isolated from other environmental factors, by comparing the experimental conditions with the rotational control, cell proliferation appeared depleted, cell growth was increased and there was an enhanced expression of cyclin B1. The functional meaning of these effects is that cell proliferation and cell growth, which are strictly associated functions under normal 1g ground conditions, are uncoupled under hypergravity. This uncoupling was also described by us in previous experiments as an effect of microgravity, but in an opposite way. Furthermore, root meristems appear thicker in hypergravity-treated than in control samples, which can be related to changes in the cell wall induced by altered gravity. - All living organisms are well-adapted to the different conditions on Earth, including the gravity force. Longterm modifications of this parameter would undoubtedly lead to evolutionary changes, whereas physiological changes are expected for short-term alterations in the gravity vector. In addition to space flight, ground based facilities are used to create altered gravity environments, both simulating microgravity on a random positioning machine (RPM) by clinorotation (Van Loon 2007) and increasing gravitational load in hypergravity habitats produced by centrifugation (Van Loon et al. 2008). Although a lot of research efforts have been dedicated to microgravity research, hypergravity is an unavoidable partner in this endeavour, being part of spaceflights during rockets or spacecrafts launching and reappearing upon recovery of samples to the Earth or an eventual travel to the Moon or Mars. Modification of the g vector quantity by centrifugation is also needed to simulate other celestial bodies gravitational loads different from 1g force on Earth. Also, during parabolic flights, hypergravity periods alternate with microgravity ones. Therefore, it is highly advisable to be capable of detecting and isolating the effects of both types of altered gravity environments. Previous experiments on the effects of hypergravity on plants have described the reorientation of microtubules in the epidermic layer of cells of the Arabidopsis hypocotyls (Hoson et al. 2010; Soga et al. 1999). Other authors have shown that hypergravity produces changes in the cell surface, intracellular transduction pathways and in the localization of subcellular organelles (Ingber 2006; Monshausen and Sievers 2002; Nickerson et al. 2004) The processes of growth, differentiation and development affect the whole plant, but they rely on cellular mechanisms, including cell proliferation and growth, which are basic and essential functions for the cell life. It is well known that signals transduced between different plant organs are capable of activating key modulators of cell growth and cell division in a coordinated manner, in meristems; the reception of these signals and the response to them is indeed called meristematic competence (Mizukami 2001). Furthermore, cell proliferation at the root meristem constitutes the source of cells for root growth and differentiation and the cellular basis for the developmental program of the plant (Dolan et al. 1993; Scheres et al. 2002). The alteration of environmental conditions such as gravity can modulate the activity of meristematic cells (Medina and Herranz 2010). Results obtained in space experiments have shown that cell proliferation parameters are modified in plants grown in space, including maize (Barmicheva et al. 1989), lentils (Darbelley et al. 1989) and also Arabidopsis (Mata et al. 2010), in which a decoupling between cell growth and cell proliferation in meristems h (...truncated)


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Ana I. Manzano, Raúl Herranz, Jack J. W. A. van Loon, F. Javier Medina. A Hypergravity Environment Induced by Centrifugation Alters Plant Cell Proliferation and Growth in an Opposite Way to Microgravity, Microgravity Science and Technology, 2012, pp. 373-381, Volume 24, Issue 6, DOI: 10.1007/s12217-012-9301-1