Gravitational and magnetic field variations synergize to cause subtle variations in the global transcriptional state of Arabidopsis in vitro callus cultures

Abstract

BackgroundBiological systems respond to changes in both the Earth's magnetic and gravitational fields, but as experiments in space are expensive and infrequent, Earth-based simulation techniques are required. A high gradient magnetic field can be used to levitate biological material, thereby simulating microgravity and can also create environments with a reduced or an enhanced level of gravity (g), although special attention should be paid to the possible effects of the magnetic field (B) itself.ResultsUsing diamagnetic levitation, we exposed Arabidopsis thaliana in vitro callus cultures to five environments with different levels of effective gravity and magnetic field strengths. The environments included levitation, i.e. simulated μg* (close to 0 g* at B = 10.1 T), intermediate g* (0.1 g* at B = 14.7 T) and enhanced gravity levels (1.9 g* at B = 14.7 T and 2 g* at B = 10.1 T) plus an internal 1 g* control (B = 16.5 T). The asterisk denotes the presence of the background magnetic field, as opposed to the effective gravity environments in the absence of an applied magnetic field, created using a Random Position Machine (simulated μg) and a Large Diameter Centrifuge (2 g).Microarray analysis indicates that changes in the overall gene expression of cultured cells exposed to these unusual environments barely reach significance using an FDR algorithm. However, it was found that gravitational and magnetic fields produce synergistic variations in the steady state of the transcriptional profile of plants. Transcriptomic results confirm that high gradient magnetic fields (i.e. to create μg* and 2 g* conditions) have a significant effect, mainly on structural, abiotic stress genes and secondary metabolism genes, but these subtle gravitational effects are only observable using clustering methodologies.ConclusionsA detailed microarray dataset analysis, based on clustering of similarly expressed genes (GEDI software), can detect underlying global-scale responses, which cannot be detected by means of individual gene expression techniques using raw or corrected p values (FDR). A subtle, but consistent, genome-scale response to hypogravity environments was found, which was opposite to the response in a hypergravity environment.

Transcriptome profiling of peanut (Arachis hypogaea) gynophores in gravitropic response

“…Similar to our results, transcriptome analysis in Arabidopsis exposed to gravity stimulation also revealed that metabolism is one of major gene groups responding to gravitropism (Kimbrough et al 2004;Kittang et al 2004) Among the metabolism pathways identified in this study, energy metabolism was also found to function in plants under gravitropic stimulation according to proteomic analysis (Azri et al 2009;Herrera et al 2010). In Arabidopsis, secondary metabolism genes were altered by magnetically induced hyperand microgravity at both transcription and translation levels (Manzano et al 2012;Herranz et al 2013). Proteome analysis of poplar stems exposed to gravitropic stimulation has led to determination of several pathways also identified in our study, including nucleotide metabolism, carbohydrate metabolism, photosynthesis and primary metabolism (energy) (Azri et al 2009).…”

Ground-Based Facilities for Simulation of Microgravity: Organism-Specific Recommendations for Their Use, and Recommended Terminology

“…In general, data obtained from cell cultures reveal alterations in cell growth and proliferation that could be compared to those found in seedlings, indicating that a significant part of the response to altered gravity does not depend on specialized cells containing mechanoreceptors (Manzano et al, 2012a;Herranz et al, 2013). In cell cultures we verified (i) the alteration of the relative proportion of cell cycle phases (which probably leads to a change in the duration of the cycle), (ii) the change in expression of numerous genes acting as regulators in cell cycle checkpoints of phase transitions, and (iii) the deregulation of ribosome biogenesis by means of the alteration of nucleolar structure and of transcriptional and post-translational changes of key proteins of this process, such as nucleolin and fibrillarin.…”

“…In addition to the work carried out with seedlings, solid in vitro cell cultures of Arabidopsis thaliana were used and exposed to the HFML magnet (maximum field of 16.5 T from 0g* to 2g*), the RPM (real random mode), and the LDC (2g, hypergravity), as a homogeneous proliferating cellular material with the use of agar-filled tubes/plates of the required size (Manzano et al, 2012a). This material has been rarely used in real spaceflight experiments (Paul et al, 2012), but its use as source material for altered gravity research has the purpose of testing whether cells not specialized in gravity perception respond to gravity alteration.…”

“…In fact, the intensity of alterations caused by the gravitational stress was strengthened by an amplified environmental stress caused by the synergy between the altered gravity and the high magnetic field, both at the gene expression and proteomic level (Manzano et al, 2012a;Herranz et al, 2013).…”

Mechanisms of disruption of meristematic competence by microgravity inArabidopsisseedlings

“…Actually, in both plant and animal cells it has been reported the perception of mechanical signals by cells not apparently specialized in gravity sensing [20][21][22] and, in several laboratories, different research groups, including us, have reported genomic and proteomic effects of an altered gravity environment (even of spaceflight) on Arabidopsis callus cell cultures. [23][24][25][26][27] In the case of the magnetic levitation, the response at the meristematic cell level is associated to an altered polar auxin transport; this means that there should be an intermediate factor capable of linking the signal sensed in the cell by the diamagnetic levitation of water and the alteration of the polar auxin transport. In turn, cell cultures lack both, gravitropic signals induced by statolith movements and polar auxin transport However, gravity alteration is sensed by cells and results in cellular and molecular effects, also including disruption of meristematic competence (unpublished results).…”