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Changes in the effective gravitational field strength affect the state of phosphorylation of stress-related proteins in callus cultures of Arabidopsis thaliana.

ABSTRACT: In a recent study it was shown that callus cell cultures of Arabidopsis thaliana respond to changes in gravitational field strengths by changes in protein expression. Using ESI-MS/MS for proteins with differential abundance after separation by 2D-PAGE, 28 spots which changed reproducibly and significantly in amount (P <0.05) after 2 h of hypergravity (18 up-regulated, 10 down-regulated) could be identified. The corresponding proteins were largely involved in stress responses, including the detoxification of reactive oxygen species (ROS). In the present study, these investigations are extended to phosphorylated proteins. For this purpose, callus cell cultures of Arabidopsis thaliana were exposed to hypergravity (8 g) and simulated weightlessness (random positioning; RP) for up to 30 min, a period of time which yielded the most reliable data. The first changes, however, were visible as early as 10 min after the start of treatment. In comparison to 1 g controls, exposure to hypergravity resulted in 18 protein spots, and random positioning in 25, respectively, with increased/decreased signal intensity by at least 2-fold (P <0.05). Only one spot (alanine aminotransferase) responded the same way under both treatments. After 30 min of RP, four spots appeared, which could not be detected in control samples. Among the protein spots altered in phosphorylation, it was possible to identify 24 from those responding to random positioning and 12 which responded to 8 g. These 12 proteins (8 g) are partly (5 out of 12) the same as those changed in expression after exposure to 2 h of hypergravity. The respective proteins are involved in scavenging and detoxification of ROS (32%), primary metabolism (20.5%), general signalling (14.7%), protein translation and proteolysis (14.7%), and ion homeostasis (8.8%). Together with our recent data on protein expression, it is assumed that changes in gravitational fields induce the production of ROS. Our data further indicate that responses toward RP are more by post-translational protein modulation (most changes in the degree of phosphorylation occur under RP-treatment) than by protein expression (hypergravity).

SUBMITTER: Barjaktarovic Z

PROVIDER: S-EPMC2652066 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Changes in the effective gravitational field strength affect the state of phosphorylation of stress-related proteins in callus cultures of Arabidopsis thaliana.

Barjaktarović Zarko Z Schütz Wolfgang W Madlung Johannes J Fladerer Claudia C Nordheim Alfred A Hampp Rüdiger R

Journal of experimental botany 20090106 3

In a recent study it was shown that callus cell cultures of Arabidopsis thaliana respond to changes in gravitational field strengths by changes in protein expression. Using ESI-MS/MS for proteins with differential abundance after separation by 2D-PAGE, 28 spots which changed reproducibly and significantly in amount (P <0.05) after 2 h of hypergravity (18 up-regulated, 10 down-regulated) could be identified. The corresponding proteins were largely involved in stress responses, including the detox ...[more]

PMID: 19129159

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Project description: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.

Dataset Information

Changes in the effective gravitational field strength affect the state of phosphorylation of stress-related proteins in callus cultures of Arabidopsis thaliana.

Publications

Changes in the effective gravitational field strength affect the state of phosphorylation of stress-related proteins in callus cultures of Arabidopsis thaliana.

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