This study examined tissue-specific early transcriptional responses upon reorientation in wild type and starchless pgm-1 mutant seedlings of Arabidopsis. Seedlings were grown in vertical Petri dishes for four days. Treatment samples were reoriented 90 degrees for ten minutes while control plants remained vertical. All seedlings were flash frozen in liquid nitrogen and RNA was extracted from root tips, mature root, hypocotyl, and cotyledon fractions of the seedlings. RNA was sequenced via NovaSeq.

On Earth plants are constantly exposed to a gravitational field of 1G. Gravity affects a plant in every step of its development. Germinating seedlings orient their radicle and hypocotyl and growing plants position organs at a specific Gravitropic Set-point Angle dictated by the asymmetric distribution of auxin depending on the gravity vector. Hence gravitropism is one of the fundamental growth responses in plants. For any experiment studying the effects of gravity on plants, the ultimate control is the microgravity in space. In this study, Arabidopsis seeds were flown to the International Space Station and allowed to germinate and grow for 3 days in microgravity. Arabidopsis Wild Type Col-0 seeds were plated onto twenty-two 60mm Petri plates, loaded into PDFUs and inserted 4 Biological Research in Canisters (BRICs). Approximately 800 seeds were sterilized, plated on each 60mm Petri plates and cold stratified for 16 hours followed by 2 hours of white light treatment. The BRICs were maintained at 4C until spaceflight to ensure seed germination in microgravity. After 3 days of germination and growth, the seedlings were fixed by injecting RNAlater into the chamber. They were kept at ambient temperature for 12 hours followed by freezing at -80C. An additional 22 plates were used as ground controls. After the spaceflight, tissue from five plates was pooled to make each of three replicates. Both membrane and soluble proteins were extracted from the pooled seedlings. Proteins were trypsin digested, labelled with iTRAQ and identified using tandem mass spectrometry.

Traveling to nearby extraterrestrial objects having a reduced gravity level (partial gravity) compared to Earth's gravity is becoming a realistic objective for space agencies. The use of plants as part of life support systems will require a better understanding of the interactions among plant growth responses including tropisms, under partial gravity conditions. Here, we present results from our latest space experiments on the ISS, in which seeds of Arabidopsis thaliana were germinated, and seedlings grew for six days under different gravity levels, namely micro-g, several intermediate partial-g levels, and 1g, and were subjected to irradiation with blue light for the last 48 hours. RNA was extracted from 20 samples for subsequent RNAseq analysis. Transcriptomic analysis was performed using the HISAT2-Stringtie-DESeq pipeline. Differentially expressed genes were further characterized for global responses using the GEDI tool, gene networks and for Gene Ontology (GO) enrichment.

These investigations studied the fundamentals of how plants perceive gravity and develop in microgravity. It informs how gene regulation is altered by spaceflight conditions. We noted expression changes in genes involved in hypoxia and heat shock responses, DNA repair, and cell wall structure between spaceflight samples compared to the ground controls. In addition, glycome profiling supported our expression analyses in that there was a difference in cell wall components between ground control and spaceflight-grown plants. Comparing our studies to those of the other BRIC-16 experiments demonstrated that, even with the same hardware and similar biological materials, differences in results in gene expression were found among these spaceflight experiments.

Physical forces greatly influence the growth and function of an organism. Altered gravity can perturb normal development and induce corresponding changes in gene expression. Understanding this relationship between the physical and biological realms is important for NASA's space travel goals. We use combined RNA-Seq and qRT-PCR to profile changes in early Drosophila melanogaster pupae exposed to chronic hypergravity (3 g, three times Earth's gravity) to highlight gravity-dependent pathways and gene products. Robust transcriptional response was evident among the pupae developed in a hypergravity environment compared to control. 1,513 genes showed significantly (p less than 0.05) altered gene expression in the 3 g samples. These findings were supported with qRT-PCR data. Major biological processes affected include ion transport, redox homeostasis, immune and humoral stress response, proteolysis, and cuticle development.

This study's objective was to develop an understanding of the biological effects of space radiation on plants with the goal of producing fresh food during long duration space missions to support astronauts' nutritional and psychological needs.10-day-old Arabidopsis seedlings were exposed to simulated Galactic Cosmic Rays (GCR) and assessed for transcriptomic changes. The simulated GCR irradiation was carried out in the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Lab (BNL). The exposures were conducted acutely for two dose points at 40 cGy or 80 cGy, with sequential delivery of proton, helium, oxygen, silicon, and iron ions. Control and irradiated seedlings were then harvested and stabilized in RNAlater at 3 hrs. post irradiation. Total RNA was isolated for transcriptomic analyses using RNAseq. The data revealed that the transcriptomic responses were dose-dependent, with significant upregulation of DNA repair pathways and downregulation of glucosinolate biosynthetic pathways. Glucosinolates are important for plant pathogen defense and for the taste of a plant, which are both relevant to growing plants for spaceflight. These findings fill in knowledge gaps of how plants respond to radiation in beyond-Earth environments.

In this study transcriptome profiling was used to gain insight into the spaceflight adaptation role of Altered response to gravity-1 (Arg1), a gene known to affect gravity responses in plants on Earth. The study compared expression profiles of cultured lines of Arabidopsis thaliana derived from wild type (WT) cultivar Col-0 to profiles from a knock-out line deficient in the gene encoding (ARG1 KO), both on the ground and in space. The cell lines were launched on SpaceX CRS-2 as part of the Cellular Expression Logic (CEL) experiment of the BRIC17 spaceflight mission. The cultured cell lines were grown within 60mm Petri plates in Petri Dish Fixation Units (PDFUs) that were housed within the Biological Research In Canisters (BRIC) hardware. Spaceflight samples were fixed on orbit. Differentially expressed genes were identified between the two environments (spaceflight and comparable ground controls) and the two genotypes (WT and ARG1 KO). Each genotype engaged unique genes during physiological adaptation to the spaceflight environment, with little overlap. Most of the genes altered in expression in spaceflight in WT cells were found to be Arg1-dependent, suggesting a major role for that gene in the physiological adaptation of undifferentiated cells to spaceflight.

These investigations studied the fundamentals of how plants perceive gravity and develop in microgravity. It informs how gene regulation is altered by spaceflight conditions.

The PESTO experiment was launched on April 8, 2002 onboard STS-110 (Atlantis) and was returned on June 19, 2002 onboard STS-111 (Endeavor), a total of 73 days in space. The ground control was conducted on a 14-day delay, and was harvested on July 3, 2002. During this period there were a total of 7 harvests of PESTO wheat and priming of 6 wheat root modules on orbit. A total of 18 plants were fixed on orbit for morphological and structural analysis, 4 plants were fixed for RNA and molecular genetic analysis and greater than 150 plants were frozen for biochemical analysis. A total of 3018 images of wheat chambers were collected from the BPS cameras during flight, and 3,640 images were obtained from the ground controls. A total of 65 digital images of the wheat harvest were collected during flight, with 75 being collected from ground controls. In addition, crew members recorded approximately 50 images and over 3 hrs of video of flight operations. Over 2 gigabytes of environmental data were obtained from the flight and ground control units. A ground control experiment was conducted in the BPS ground control chamber in the Orbiter Environment Simulator (OES) at the Kennedy Space Center Launch Site Support Facility in Hangar L. The ground control was delayed by 14-days from the real- time flight operations in order to duplicate the flight operations as closely as possible in a 1 g environment. At launch, two chambers were dedicated to the PESTO study, which consisted of three growth cycles using Apogee wheat. The third chamber was dedicated to the TVT study for the first growth cycle and used Apogee wheat. For the subsequent two wheat growth cycles, the third chamber was used for the PESTO study. For the first growth cycle in all Plant Growth Chambers, seeds were germinated pre-launch. In PESTO, the ages of the plants were staggered at launch so that comparisons of physiological and biochemical properties between mature plants, which were well developed and photosynthetically competent, and young plants could be studied. In-flight, plants were harvested at a specific age, and the initiation of the next cycle of growth was staggered between the chambers

Understanding the molecular mechanisms by which plants sense and adapt to changes in the space environment is essential for generating plants that are better adapted to withstand space flight, microgravity, and other adverse conditions encountered in space. The objective of our spaceflight experiment “Plant Signaling in Microgravity” (carried out on the International Space Station, ISS), was to compare transcript profiles of wild type and transgenic InsP 5-ptase plants with compromised InsP3 signaling. The transgenic Arabidopsis plants constitutively express the mammalian type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme that specifically hydrolyzes the lipid-derived second messenger inositol 1,4,5-trisphosphate (InsP3). These transgenic plants exhibit normal growth and morphology; however, their responses to environmental stimuli including gravity and drought are altered. Seedlings were grown for 5 days under continuous light in experimental containers placed in the European Modular Cultivation system (EMCS) onboard the ISS. The EMCS consists of two rotors within a controlled chamber, allowing for a “1g” control in space. After sample retrieval from the ISS, RNA was isolated from shoot and root tissue and subjected to RNA sequencing. Two-way comparisons of micro g versus “1”g have uncovered regulatory mechanisms that are both conserved and altered between the wild type and transgenic seedlings.