Arabidopsis thaliana in altered gravity: WS in C-9 parabolic flight, 2015

The increasing availability of flights on suborbital rockets creates new avenues for the study of spaceflight effects on biological systems, in particular the transitions between hypergravity and microgravity. This paper presents an initial comparison of the responses of Arabidopsis thaliana to suborbital and atmospheric parabolic flights as an important step toward characterizing these emerging suborbital platforms and their effects on biology. Transcriptomic profiling of the response of the Arabidopsis ecotype Wassilewskija (WS) to the aggregate spaceflight experiences in the Blue Origin New Shepard and Virgin Galactic SpaceShipTwo rockets revealed that the transcriptomic load induced by flight differed greatly between the two flights, yet was biologically related to traditional parabolic flight responses. The sku5 skewing mutant and 14-3-3κ:GFP regulatory protein overexpression lines each showed altered intra-platform responses compared to WS in the Blue Origin and parabolic flights, respectively. An additional parabolic flight using the F-104 Starfighter showed that the response of 14-3-3κ:GFP to flight was modulated in a similar manner to the WS line. Despite the differing genotypes, experimental workflows, flight profiles and platforms, alteration of gene expression remodeling central metabolic processes was commonly observed as a response to the flights. The processes included carbon and nitrogen metabolism, branched-chain amino acid degradation, and hypoxic responses. The timing and directionality of differentially-expressed genes involved in the conserved pathways differed among the platforms. The data presented herein highlight the potential for various suborbital platforms to contribute insights into biological responses to spaceflight, and further suggest that in-flight fixation during suborbital experiments will provide insights into responses to each phase of flight. Sterilized seeds were planted on 0.5x MS Phytagel™ media, and maintained at the University of Florida at 19 ± 2 °C, with 24-h fluorescent lighting of about 80 m-2 s-1 PAR. Plates were transported to Ellington Airport, TX, inside a cooler lit with an LED light bank. A makeshift grow area was set up at the experimental location using a fluorescent light bank. Kennedy Space Center Fixation Tubes (KFTs) were preloaded with RNAlater™ fixative. The morning before the C-9 parabolic flight experiment, seedlings were transplanted to the sample chambers of the KFTs, onto moistened Kimwipes shaped roughly into a cylinder. The following day, KFTs were taken onboard the C-9 aircraft. KFTs were actuated in level flight before the onset of parabolic flight, immediately at the start of the first parabola, mid-way through the first parabola at the hypergravity to microgravity, and after the first, third, and tenth parabolas. Samples were stored at 4 °C, and transported on ice back to UF, where they were stored at -80 °C. Samples were later removed from -80 °C, roots were dissected from shoot tissues, and roots were grouped into 3 replicates per condition. Total RNA was extracted from roots and submitted for microarray analysis. The Affymetrix GeneChip Arabidopsis Gene (AraGene) 1.0 ST array was used to measure gene expression levels. The transcriptomic data from the samples fixed in parabolic flight were then compared to those fixed during level flight in order to identify differentially-expressed genes associated with parabolic flight patterns.