Microarray (one-color) gene expression analysis of Euglena gracilis subjected to altered acceleration during parabolic flight
ABSTRACT: Euglena gracilis is a unicellular freshwater flagellate, which uses the gravitational vector for orientation in the water column in the dark. This allows the cell to reach areas in the water column for reproduction and growth. How exactly the gravitational vector is perceived, and which intracellular pathways are involved in the signaling is not very well understood so far. In the past, parabolic flight campaigns were used to study the swimming behavior of Euglena gracilis under altered gravitational accelerations. It was shown that cells adapt their swimming direction very fast: in the dark under 1xg cell show negative gravitaxis, i.e. they move upwards in the water column of the experiment hardware and against the gravitational vector. With onset of the first hypergravity period of 1.8xg the precision of upward swimming increases slightly. This first hyper gravity lasts for only 20 seconds and is followed by 22 sec of microgravity. During this period no gravitational vector is perceived by the cells, therefore they lack a cue for orientation and move randomly in all direction. In the subsequent hyper gravity period of 1.8xg, which also lasts for 20 sec, cells direct their movement again and swim upwards. Over different parabolic flight campaigns and other experiments it was shown that this gravitactic behavior is linked to changes in membrane potential, calcium and cAMP concentration. However, due to the lack of genomic and transcriptomic data, it was so far not possible to link the differential movement to the abundance of distinct mRNA transcripts. In contrast, other model organisms, such as Arabidopsis thaliana, have been analyzed by means of gene expression with respect to the effects of altered gravitational accelerations. Also various human cell lines have shown to adapt their gene expression in dependence of the prevailing acceleration. With the recently published Euglena gracilis transcriptome, we now aimed at analyzing effects of altered acceleration on the gene expression in the flagellate. Therefore, Euglena gracilis samples were taken in the course of the 29th DLR parabolic flight campaign during parabola 1 and 31 (time difference of 2 hours and 30 additional parabolas). During both parabolas samples were fixed with TRIzol at 1xg just before onset of the first hyper gravity period, 20 sec into hyper gravity (1.8xg), 20 sec into microgravity (µg) and 20 sec into the last hypergravity period.
Project description:The general objective of the study was to determine modulation of gene expression by environmental factors, with a special emphasis on bone formation. For this reason, the specific period of treatment was chosen between 5-6 days post-fertilization (dpf), when bone formation and calcification are taking place. This experiment was designed as a new type of gravitational experiment, which we like to call \relative microgravity\, referring to the fact that the larvae first grow in hyper gravity for 5 days and are then returned to 1g normal gravity for 1 day. Zebrafish embryos were placed on a Large Diameter Centrifuge at 3 hpf, brought to a gravitational force of 3 g until 5 dpf. Reference embryos were kept in parallel at 1g (Inc). At 5dpf, one batch was left at 3g (3g), one batch was returned to 1g (3g>1g), while a third batch was returned to 1g, but left on the axis of the centrifuge (Axe; 3g>Axe). The experiment was repeated 4 times, each time with 4 batches of 60 larvae.
Project description:R. rubrum S1H inoculated on solid agar rich media was sent to the ISS in October 2003 (MESSAGE-part 2 experiment). After 10 days flight, R. rubrum cultures returned back to Earth. These cultures were then subjected to both transcriptomic and proteomic analysis and compared with the corresponding ground control. Whole-genome oligonucleotide microarray and high throughput proteomics, which offer the possibility to survey respectively the global transcriptional and translational response of an organism, were used to test the effect of space flight. Moreover, in an effort to identify a specific stress response of R. rubrum to space flight, ground simulation of space ionizing radiation and space gravity were performed under identical culture setup and growth conditions encountered during the actual space journey. This study is unique in combining the results from an actual space experiment with the corresponding space ionizing radiation and modeled microgravity ground simulations, which lead to a more solid dissection of the different factors contribution acting in space flight conditions. Total RNA was extracted from R. rubrum S1H grown after 10 days in space flight or after 10 days in simulated ionizing radiation or simulated microgravity. Each microarray slide contained 3 technical repeats.
Project description:The effect of microgravity on gene expression in C.elegans was comprehensively analysed by DNA microarray. This is the first DNA microarray analysis for C.elegans grown under microgravity. Hyper gravity and clinorotation experiments were performed as reference against the flight experiment. Total RNA was extracted from 100,000 worms of each experimental group
Project description:We address a key baseline question of whether gene expression changes are induced by the orbital environment, and then we ask whether undifferentiated cells, cells presumably lacking the typical gravity response mechanisms, perceive spaceflight. Arabidopsis seedlings and undifferentiated cultured Arabidopsis cells were launched in April, 2010, as part of the BRIC-16 flight experiment on STS-131. Biologically replicated DNA microarray and averaged RNA digital transcript profiling revealed several hundred genes in seedlings and cell cultures that were significantly affected by launch and spaceflight. The response was moderate in seedlings; only a few genes were induced by more than 7-fold, and the overall intrinsic expression level for most differentially expressed genes was low. In contrast, cell cultures displayed a more dramatic response, with dozens of genes showing this level of differential expression, a list comprised primarily of heat shock-related and stress-related genes. This baseline transcriptome profiling of seedlings and cultured cells confirms the fundamental hypothesis that survival of the spaceflight environment requires adaptive changes that are both governed and displayed by alterations in gene expression. The comparison of intact plants with cultures of undifferentiated cells confirms a second hypothesis: undifferentiated cells can detect spaceflight in the absence of specialized tissue or organized developmental structures known to detect gravity.
Project description:osteocyte is the mechanosensor in bone, taking up pivotal position in mediating the mechano-induced bone remodeling. Dimagnetic levitation has been used to stimulating a reduced gravity environment for studing the effects of changed gravity to different organisms.we constructed a superconducting magnet based platform with a large gradient high magnetic field(LG-HMF),which can provide three apparent gravity levels (μ-g,1-g and 2-g). osteocytes are sensetive to gravitational changes, our aim is to explore what responses do osteocytes exists under different gravitational environments in gene level, together with filtering the up- and down-regulated genes. mouse osteocyte-like cell line MLO-Y4 were cultured under three different apparent gravity levels (μ-g,1-g and 2-g) and normal gravity environment (control) for 48 hours, after which total RNA was extracted . And then RNA samples hybridized on affymetrix microarrays to obtain the whole genome expression profiles. the aim that we selected 48 hours as the cell culture time was to make a comparison with our previous researches of osteocytes.
Project description:We investigated differentially regulated genes in human Jurkat T lymphocytic cells in 20s and 5min microgravity and in hypergravity and compared expression profiles to identify potential gravity-regulated genes and adaptation processes. Overall design: Human Jurkat T cells were exposed to altered gravity during a parabolic flight
Project description:We investigated differentially regulated genes in human Jurkat T lymphocytic cells in 20s and 5min microgravity and in hypergravity and compared expression profiles to identify potential gravity-regulated genes and adaptation processes. Overall design: Human Jurkat T cells were exposed to altered gravity during a sounding rocket flight
Project description:The study was aimed at bioinformatic analysis of transcriptome changes in lumbar spinal cords of mice after the 30-day space flight aboard biosatellite Bion-M1 and subsequent 7-day re-adaptation to the Earth's gravity when compared with control group.
Project description:In prospective human exploration of outer space, the need to maintain a species over several generations under changed gravity conditions may arise. This paper reports the analysis of the third generation of fruit fly Drosophila melanogaster obtained during the 44.5-day space flight (Foton-M4 satellite, 2014, Russia), followed by the fourth generation on Earth and the fifth generation under conditions of a 12-day space flight (2014, in the Russian Segment of the ISS). The obtained results show that it is possible to obtain the third-fifth generations of a complex multicellular Earth organism under changed gravity conditions (in the cycle “weightlessness – Earth – weightlessness”), which preserves fertility and normal development. However, there were a number of changes in the expression levels and content of cytoskeletal proteins that are the key components of the spindle apparatus and the contractile ring of cells. Overall design: Examination of drosophila genes expression change during space flight by RNA-Seq technique
Project description:Using diamagnetic levitation, we have exposed A. thaliana in vitro callus cultures to five environments with different levels of effective gravity (from levitation i.e. simulated mg* to 2g*) and magnetic fields (10.1 to 16.5 Tesla) and we have compared the results with those of similar experiments done in a Random Position Machine (simulated micro g) and a Large Diameter Centrifuge (2g) free of high magnetic fields. Microarray analysis indicates that there are changes in overall gene expression of the cultured cells exposed to these unusual environments but also that gravitational and magnetic field produce synergic variations in the steady state of the transcriptional profile of A. thaliana. Significant changes in the expression of structural, abiotic stress and secondary metabolism genes were observed into the magnet field. These results confirm that the strong magnetic field, both at micro g* or 2g*, has a significant effect on the expression of these genes but subtle gravitational effects are still observable. These subtle responses to microgravity environments are opposite to the ones observed in a hypergravity one. Overall design: seven-condition experiment, MM2D Arabidopsis culture callus control vs. Treatment (altered gravity simulation, GBF). Three GBF were used (LDC (2g) + control, RPM (mg) + control and Magnet (mg*, 0.1g*, 1g*, 1.9g*, 2g*) + control). Biological replicates: 3 replicates in all conditions and controls except 1.9g* (2 replicates)