Volume 3 (2015): Issue 2 (December 2015)

Recent spaceflight experiments have provided many new insights into the role of gravity in plant growth and development. Scientists have been taking seeds and plants into space for decades in an effort to understand how the stressful environment of space affects them. The resultant data have yielded significant advances in the development of advanced life-support systems for long-duration spaceflight and a better understanding of the fundamental role of gravity in directing the growth and development of plants. Experiments have improved as new spaceflight hardware and technology paved the way for progressively more insightful and rigorous plant research in space. The International Space Station (ISS) has provided an opportunity for scientists to both monitor and control their experiments in real-time. Experiments on the ISS have provided valuable insights into endogenous growth responses, light responses, and transcriptomic and proteomic changes that occur in the microgravity environment. In recent years most studies of plants in space have used Arabidopsis thaliana, but the single-celled, Ceratopteris richardii spore is also a valuable model system that has been used to understand plant gravity response. Experiments using these fern spores have revealed a dynamic and gravity-responsive trans-cell Ca²⁺ current that directs polarization of these spores and a possible role of extracellular nucleotides in establishing or contributing to this current. As technology continues to improve, spaceflight experiments will provide many new insights into the role and effects of gravity on plant growth and development.

Gravity is a fundamental stimulus that affects plant growth and development. The gravity persistent signal (GPS) treatment uses a cold treatment to isolate the events of signal transduction. Plants are reoriented horizontally in the dark at 4°C for 1 hour and then returned to vertical at room temperature. A gene expression microarray was designed to identify genes that are regulated during the GPS treatment. Arabidopsis thaliana var. Columbia was grown to maturity with inflorescence stems of 8-10 cm. Total mRNA was collected from inflorescence stems at 2, 4, 10, and 30 min after reorientation in the cold. cDNA was synthesized from the mRNA and then probed against an Arabidopsis gene expression array with 4 replicates per time point. Analyses presented here focus on transcription factors because of their regulatory functions in response pathways. Five transcription factors (AtAIB, WRKY18, WRKY26, WRKY33, and BT2) were selected for further study based on their expression at 4 min. Quantitative real-time polymerase chain reaction ((PCR) RT-qPCR) was performed to confirm expression seen in the microarray data. Seeds of Arabidopsis lines containing T-DNA insertions in the genes were obtained, plants bred to homozygosity, and the mutants analyzed for GPS phenotype. Mutant analysis shows significant differences in curvature of inflorescence stems between mutants and wild type.

In order to meet heat rejection requirements for future NASA exploration, scientific, and discovery missions, a study is being conducted for the feasibility of integral variable conductance planar heat pipe (VCPHP) technology. This represents a novel, low technology readiness level (TRL) heat rejection technology that, when developed, could operate efficiently and reliably across a wide range of thermal environments. The concept consists of a planar heat pipe whose evaporator acquires the excess thermal energy from the thermal control system and rejects it at its condenser whose outer surface acts as a radiating surface. The heat pipe is made from thermally conductive polymers in order to minimize its mass. It has a non-condensable gas that changes the active radiator surface depending on the heat load. A mathematical model of steady-state variable conductance heat pipe is developed. Two planar heat pipes are designed, fabricated, and tested to validate the theoretical model. The feasibility of the proposed VCPHP working in a space environment is discussed, based on the model.

The effects of spaceflight on yeast have high concordance with agents that induce a very low intracellular redox state and induce a massive efflux of glutathione. These results raise important issues. Can the reduced redox state during spaceflight be reproduced and modulated in ground-based simulations? Will this allow definition of unique drug pathways as a low redox potential state mirrors the electrophilic properties of mitochondria where many drugs are metabolized? Unfortunately, assays for redox status and its major cellular determinant—glutathione—are diverse and often cell-type-specific. Currently, an accepted redox probe set for yeast studies is not available. This paper validates fluorescent probes for glutathione and reactive oxygen status in yeast to support mechanistic studies of microgravity and drug metabolism. The plethora of fluorescent reagents for reactive oxygen species and glutathione makes head-to-head comparisons of all the alternatives impractical. These reagents measure the physiological milieu of reactive oxygen species and diverse thiols, rather than specific individual molecules. We report that in yeast, monochlorobimane (mBCL) and 2’,7’-dichlorodihydrofluorescein diacetate (DC-FDA) are suitable for fluorometric and flow cytometry studies of glutathione and reactive oxygen species, respectively. Both dyes have low background fluorescence, predictable loading, good retention, and are not acutely toxic to Saccharomyces cerevisiae. Both dyes show concordance with other fluorescent and biochemical assays of reactive oxygen species.

Vascular patterning is a key, genetically responsive phylogenetic classifier of tissues in major organisms flown in space, such as the wings of Drosophila melanogaster (the fruit fly), mouse retina, and leaves of Arabidopsis thaliana. Phenotypes of increasingly abnormal ectopic wing venation in the highly stereotyped Drosophila wing generated by overexpressing the H-C2 construct of Notch antagonist Hairless (Johannes and Preiss, 2002) were mapped and quantified by NASA’s VESsel GENeration Analysis (VESGEN) software. By several confirming vascular parameters, the eight stereotyped wing veins remained quite constant in wild type compared to Class 5 H-C2, the most perturbed category of the H-C2 overexpression phenotypes. However, ectopic veins increased in number from 1 in the wild type, to 18 in Class 5 H-C2. We therefore demonstrate the feasibility of using VESGEN to quantify microscopic images of altered wing venation in Drosophila melanogaster. We further determined that several of the signal transduction pathways affecting wing vein patterning were altered by spaceflight, according to gene expression differences observed in our transcriptomic data from a previous shuttle flight experiment. Future studies will help characterize the extent to which these gene expression changes can cause even subtle developmental changes using model organisms, such as Drosophila. Therefore, we propose that the sensitive analyses provided by VESGEN software will not only serve as a useful tool to map the genetics of wing vein patterning for terrestrial applications, but also for future phenotypic studies with Drosophila for spaceflight missions.