Growth In Microgravity Research Articles

Whole genome-scale assessment of gene fitness of Novosphingobium aromaticavorans during spaceflight

In microgravity, bacteria undergo intriguing physiological adaptations. There have been few attempts to assess global bacterial physiological responses to microgravity, with most studies only focusing on a handful of individual systems. This study assessed the fitness of each gene in the genome of the aromatic compound-degrading Alphaproteobacterium Novosphingobium aromaticavorans during growth in spaceflight. This was accomplished using Comparative TnSeq, which involves culturing the same saturating transposon mutagenized library under two different conditions. To assess gene fitness, a novel comparative TnSeq analytical tool was developed, named TnDivA, that is particularly useful in leveraging biological replicates. In this approach, transposon diversity is represented numerically using a modified Shannon diversity index, which was then converted into effective transposon density. This transformation accounts for variability in read distribution between samples, such as cases where reads were dominated by only a few transposon inserts. Effective density values were analyzed using multiple statistical methods, including log2-fold change, least-squares regression analysis, and Welch’s t-test. The results obtained across applied statistical methods show a difference in the number of significant genes identified. However, the functional categories of genes important to growth in microgravity showed similar patterns. Lipid metabolism and transport, energy production, transcription, translation, and secondary metabolite biosynthesis and transport were shown to have high fitness during spaceflight. This suggests that core metabolic processes, including lipid and secondary metabolism, play an important role adapting to stress and promoting growth in microgravity.

Equilibration of precipitants in a counter-diffusion apparatus for protein crystallization.

A cost-effective capillary dialysis apparatus (Toledo Capillary Box, TCB) developed for biomacromolecule crystal growth in microgravity and unit gravity environments can provide slow equilibration between the precipitant reservoir and capillary solutions, nurturing growth of neutron-diffraction-quality crystals. Under microgravity conditions, mass transfer of precipitants and biomacro-mol-ecules occurs under diffusion-controlled conditions, promoting slow growth and suppressing defect formation. The equilibration of common precipitants (polyethyl-ene glycol and salts such as ammonium sulfate) between capillary and reservoir solutions was measured for capillaries oriented horizontally or vertically with respect to the gravitational field at unit gravity. Precipitants equilibrated less rapidly in the vertical orientation when capillary solution densities were lower than those of the reservoir solutions. A plug filled with agarose gel was introduced in the TCB apparatus for salt precipitants since salts often exhibit relatively high free diffusion. Equilibration of the capillaries with reservoir solutions was significantly delayed for many of the salt precipitants tested. Analytical and semi-analytical models allow the prediction of precipitant equilibration of capillary and reservoir solutions under diffusion-controlled transport and show good agreement with experimental results.

Root growth direction in simulated microgravity is modulated by a light avoidance mechanism mediated by flavonols.

In a microgravity environment, without any gravitropic signal, plants are not able to define and establish a longitudinal growth axis. Consequently, absorption of water and nutrients by the root and exposure of leaves to sunlight for efficient photosynthesis is hindered. In these conditions, other external cues can be explored to guide the direction of organ growth. Providing a unilateral light source can guide the shoot growth, but prolonged root exposure to light causes a stress response, affecting growth and development, and also affecting the response to other environmental factors. Here, we have investigated how the protection of the root from light exposure, while the shoot is illuminated, influences the direction of root growth in microgravity. We report that the light avoidance mechanism existing in roots guides their growth towards diminishing light and helps establish the proper longitudinal seedling axis in simulated microgravity conditions. This process is regulated by flavonols, as shown in the flavonoid‐accumulating mutant transparent testa 3, which shows an increased correction of the root growth direction in microgravity, when the seedling is grown with the root protected from light. This finding may improve the efficiency of water and nutrient sourcing and photosynthesis under microgravity conditions, as they exist in space, contributing to better plant fitness and biomass production in space farming enterprises, necessary for space exploration by humans.

Tracing transport of protein aggregates in microgravity versus unit gravity crystallization

Microgravity conditions have been used to improve protein crystallization from the early 1980s using advanced crystallization apparatuses and methods. Early microgravity crystallization experiments confirmed that minimal convection and a sedimentation-free environment is beneficial for growth of crystals with higher internal order and in some cases, larger volume. It was however realized that crystal growth in microgravity requires additional time due to slower growth rates. The progress in space research via the International Space Station (ISS) provides a laboratory-like environment to perform convection-free crystallization experiments for an extended time. To obtain detailed insights in macromolecular transport phenomena under microgravity and the assumed reduction of unfavorable impurity incorporation in growing crystals, microgravity and unit gravity control experiments for three different proteins were designed. To determine the quantity of impurity incorporated into crystals, fluorescence-tagged aggregates of the proteins (acting as impurities) were prepared. The recorded fluorescence intensities of the respective crystals reveal reduction in the incorporation of aggregates under microgravity for different aggregate quantities. The experiments and data obtained, provide insights about macromolecular transport in relation to molecular weight of the target proteins, as well as information about associated diffusion behavior and crystal lattice formation. Results suggest one explanation why microgravity-grown protein crystals often exhibit higher quality. Furthermore, results from these experiments can be used to predict which proteins may benefit more from microgravity crystallization.

Loss of physical contact in space alters the dopamine system in C. elegans

SummaryProgressive neuromuscular decline in microgravity is a prominent health concern preventing interplanetary human habitation. We establish functional dopamine-mediated impairments as a consistent feature across multiple spaceflight exposures and during simulated microgravity in C. elegans. Animals grown continuously in these conditions display reduced movement and body length. Loss of mechanical contact stimuli in microgravity elicits decreased endogenous dopamine and comt-4 (catechol-O-methyl transferase) expression levels. The application of exogenous dopamine reverses the movement and body length defects caused by simulated microgravity. In addition, increased physical contact made comt-4 and dopamine levels rise. It also increased muscular cytoplasmic Ca2+ firing. In dop-3 (D2-like receptor) mutants, neither decrease in movement nor in body length were observed during simulated microgravity growth. These results strongly suggest that targeting the dopamine system through manipulation of the external environment (contact stimuli) prevents muscular changes and is a realistic and viable treatment strategy to promote safe human deep-space travel.

Plant Gravitropism: From Mechanistic Insights into Plant Function on Earth to Plants Colonizing Other Worlds.

Gravitropism, the growth of roots and shoots toward or away from the direction of gravity, has been studied for centuries. Such studies have not only led to a better understanding of the gravitropic process itself, but also paved new paths leading to deeper mechanistic insights into a wide range of research areas. These include hormone biology, cell signal transduction, regulation of gene expression, plant evolution, and plant interactions with a variety of environmental stimuli. In addition to contributions to basic knowledge about how plants function, there is accumulating evidence that gravitropism confers adaptive advantages to crops, particularly under marginal agricultural soils. Therefore, gravitropism is emerging as a breeding target for enhancing agricultural productivity. Moreover, research on gravitropism has spawned several studies on plant growth in microgravity that have enabled researchers to uncouple the effects of gravity from other tropisms. Although rapid progress on understanding gravitropism witnessed during the past decade continues to be driven by traditional molecular, physiological, and cell biological tools, these tools have been enriched by technological innovations in next-generation omics platforms and microgravity analog facilities. In this chapter, we review the field of gravitropism by highlighting recent landmark studies that have provided unique insights into this classic research topic while also discussing potential contributions to agriculture on Earth and beyond.

Spaceflight induces novel regulatory responses in Arabidopsis seedling as revealed by combined proteomic and transcriptomic analyses

BackgroundUnderstanding of gravity sensing and response is critical to long-term human habitation in space and can provide new advantages for terrestrial agriculture. To this end, the altered gene expression profile induced by microgravity has been repeatedly queried by microarray and RNA-seq experiments to understand gravitropism. However, the quantification of altered protein abundance in space has been minimally investigated.ResultsProteomic (iTRAQ-labelled LC-MS/MS) and transcriptomic (RNA-seq) analyses simultaneously quantified protein and transcript differential expression of three-day old, etiolated Arabidopsis thaliana seedlings grown aboard the International Space Station along with their ground control counterparts. Protein extracts were fractionated to isolate soluble and membrane proteins and analyzed to detect differentially phosphorylated peptides. In total, 968 RNAs, 107 soluble proteins, and 103 membrane proteins were identified as differentially expressed. In addition, the proteomic analyses identified 16 differential phosphorylation events. Proteomic data delivered novel insights and simultaneously provided new context to previously made observations of gene expression in microgravity. There is a sweeping shift in post-transcriptional mechanisms of gene regulation including RNA-decapping protein DCP5, the splicing factors GRP7 and GRP8, and AGO4,. These data also indicate AHA2 and FERONIA as well as CESA1 and SHOU4 as central to the cell wall adaptations seen in spaceflight. Patterns of tubulin-α 1, 3,4 and 6 phosphorylation further reveal an interaction of microtubule and redox homeostasis that mirrors osmotic response signaling elements. The absence of gravity also results in a seemingly wasteful dysregulation of plastid gene transcription.ConclusionsThe datasets gathered from Arabidopsis seedlings exposed to microgravity revealed marked impacts on post-transcriptional regulation, cell wall synthesis, redox/microtubule dynamics, and plastid gene transcription. The impact of post-transcriptional regulatory alterations represents an unstudied element of the plant microgravity response with the potential to significantly impact plant growth efficiency and beyond. What’s more, addressing the effects of microgravity on AHA2, CESA1, and alpha tubulins has the potential to enhance cytoskeletal organization and cell wall composition, thereby enhancing biomass production and growth in microgravity. Finally, understanding and manipulating the dysregulation of plastid gene transcription has further potential to address the goal of enhancing plant growth in the stressful conditions of microgravity.

Effect of macromolecular mass transport in microgravity protein crystallization

Abstract To investigate the effect of macromolecular transport and the incorporation of protein aggregate impurities in growing crystals, experiments were performed on the International Space Station (ISS) and compared with control experiments performed in a 1G laboratory environment. Crystal growth experiments for hen egg-white lysozyme (HEWL) and Plasmodium falciparum glutathione S-transferase (PfGST) were monitored using the ISS Light Microscopy Module (LMM). Experiments were performed applying the liquid–liquid counter diffusion crystallization method using rectangular, optically transparent capillaries. To analyze the quantity of impurity incorporated into growing crystals, stable fluorescently labeled protein aggregates were prepared and subsequently added at different percent concentrations to nonlabeled monomeric protein suspensions. For HEWL, a covalent cross-linked HEWL dimer was fluorescently labeled, and for PfGST, a stable tetramer was prepared. Crystallization solutions containing different protein aggregate ratios were prepared. The frozen samples were launched on 19.02.2017 via SpaceX-10 mission and immediately transferred to a -80°C freezer on the ISS. Two series of crystallization experiments were performed on ISS, one during 26.02.2017 to 10.03.2017 and a second during 16.06.2017 to 23.06.2017. A comparison of crystal growth rate and size showed different calculated average growth rates as well as different dimensions for crystals growing in different positions along the capillary. The effect of macromolecular mass transport on crystal growth in microgravity was experimentally calculated. In parallel, the percentage of incorporated fluorescent aggregate into the crystals was monitored utilizing the fluorescent LMM and ground-based fluorescent microscopes.

Crystal growth in microgravity for neutron diffraction studies

Crystal growth in microgravity for neutron diffraction studies

30 Years of Crystal Growth Under Microgravity Conditions in Freiburg: An Overview of Past Activities

AbstractIn the framework of this overview, the past four decades of research activities from the Freiburg Crystallography in the field of crystal growth in microgravity (µg) environments is presented. Crystal growth from the melt and solution phases of different materials was investigated onboard various manned and unmanned µg platforms such as parabolic flights, shuttle missions, satellite missions, and sounding rocket flights. The focus of material research in Freiburg was and is still in semiconducting materials; elemental semiconductors like Si, Ge, and GeSi as well as various compound semiconductors like InP, GaAs, and CdTe. The objectives of the single experiments, the results, and their further application and utilizations are summarized in this overview.

Terrestrial development of the experiments on the fullerite C60 crystal growth in microgravity

Terrestrial development of the experiments on the fullerite C60 crystal growth in microgravity

Crystal growth of Cd1−xZnxTe by the traveling heater method in microgravity on board of Foton-M4 spacecraft

Cadmium zinc telluride crystals were grown using the traveling heater method (THM) under microgravity conditions on board of Foton-M4 spacecraft, and a reference crystal was grown on Earth under gravity conditions. Structure, chemical and phase compositions of these crystals, their optical characteristics and microhardness were compared. It can be concluded that the THM growth in microgravity has a positive effect on CZT crystals, since they have more homogeneous composition and their structural perfection is improved as compared with the crystals grown under terrestrial conditions, which results in improvement of electric and optical characteristics.

JAXA protein crystallization in space: ongoing improvements for growing high-quality crystals

The Japan Aerospace Exploration Agency (JAXA) started a high-quality protein crystal growth project, now called JAXA PCG, on the International Space Station (ISS) in 2002. Using the counter-diffusion technique, 14 sessions of experiments have been performed as of 2012 with 580 proteins crystallized in total. Over the course of these experiments, a user-friendly interface framework for high accessibility has been constructed and crystallization techniques improved; devices to maximize the use of the microgravity environment have been designed, resulting in some high-resolution crystal growth. If crystallization conditions were carefully fixed in ground-based experiments, high-quality protein crystals grew in microgravity in many experiments on the ISS, especially when a highly homogeneous protein sample and a viscous crystallization solution were employed. In this article, the current status of JAXA PCG is discussed, and a rational approach to high-quality protein crystal growth in microgravity based on numerical analyses is explained.

Fundamental Plant Biology Enabled by The Space Shuttle

The relationship between fundamental plant biology and space biology was especially synergistic in the era of the Space Shuttle. While all terrestrial organisms are influenced by gravity, the impact of gravity as a tropic stimulus in plants has been a topic of formal study for more than a century. And while plants were parts of early space biology payloads, it was not until the advent of the Space Shuttle that the science of plant space biology enjoyed expansion that truly enabled controlled, fundamental experiments that removed gravity from the equation. The Space Shuttle presented a science platform that provided regular science flights with dedicated plant growth hardware and crew trained in inflight plant manipulations. Part of the impetus for plant biology experiments in space was the realization that plants could be important parts of bioregenerative life support on long missions, recycling water, air, and nutrients for the human crew. However, a large part of the impetus was that the Space Shuttle enabled fundamental plant science essentially in a microgravity environment. Experiments during the Space Shuttle era produced key science insights on biological adaptation to spaceflight and especially plant growth and tropisms. In this review, we present an overview of plant science in the Space Shuttle era with an emphasis on experiments dealing with fundamental plant growth in microgravity. This review discusses general conclusions from the study of plant spaceflight biology enabled by the Space Shuttle by providing historical context and reviews of select experiments that exemplify plant space biology science.

Homogeneous SiGe crystal growth in microgravity by the travelling liquidus-zone method

Homogeneous SiGe crystal growth experiments will be performed on board the ISS "Kibo" using a gradient heating furnace (GHF). A new crystal growth method invented for growing homogeneous mixed crystals named "travelling liquidus-zone (TLZ) method" is evaluated by the growth of Si0.5Ge0.5 crystals in space. We have already succeeded in growing homogeneous 2mm diameter Si0.5Ge0.5 crystals on the ground but large diameter homogeneous crystals are difficult to be grown due to convection in a melt. In microgravity, larger diameter crystals can be grown with suppressing convection. Radial concentration profiles as well as axial profiles in microgravity grown crystals will be measured and will be compared with our two-dimensional TLZ growth model equation and compositional variation is analyzed. Results are beneficial for growing large diameter mixed crystals by the TLZ method on the ground. Here, we report on the principle of the TLZ method for homogeneous crystal growth, results of preparatory experiments on the ground and plan for microgravity experiments.

Recombinant formate dehydrogenase from Arabidopsis thaliana: Preparation, crystal growth in microgravity, and preliminary X-ray diffraction study

Crystals of high-purity recombinant NAD+-dependent formate dehydrogenase from the higher plant Arabidopsis thaliana (AraFDH) were grown in microgravity in the Modul’-1 protein crystallization apparatus on the International Space Station. The space-grown crystals have larger sizes than those grown on Earth. X-ray diffraction data suitable for determining the three-dimensional structure were collected from the space-grown crystals to a resolution of 1.22 A using an X-ray synchrotron source. The crystals belong to sp. gr. P43212; the unit-cell parameters are a = b = 107.865 A, c = 71.180 A, α = β = γ = 90°.

Melt Structural Self-Organization and Viscosity within the Transient Layer during a Single Crystal Growth in Microgravity

A brief review is given of the results obtained and published in 2003–2007 by IChPM and IPPE during their joint study and modeling of Ge:Ga, Ge:Sb, GaSb:Te, InP:S single crystal growth from stoichiometric and non-stoichiometric melts on board the Photon satellite series. The use of microgravity is shown to be justified and holding promise for research into the structural self-organization processes (cluster forming) taking place within the transient layer of the melt during the solidification. The mathematical model of convective heat and mass transfer taking into account the dual-phase character of matter in the boundary layers near the interface has been created and used as an independent tool for the study of such processes. Prospects are discussed for this new area of space material science.

Plant cell proliferation and growth are altered by microgravity conditions in spaceflight

Seeds of Arabidopsis thaliana were sent to space and germinated in orbit. Seedlings grew for 4 d and were then fixed in-flight with paraformaldehyde. The experiment was replicated on the ground in a Random Positioning Machine, an effective simulator of microgravity. In addition, samples from a different space experiment, processed in a similar way but fixed in glutaraldehyde, including a control flight experiment in a 1 g centrifuge, were also used. In all cases, comparisons were performed with ground controls at 1 g. Seedlings grown in microgravity were significantly longer than the ground 1 g controls. The cortical root meristematic cells were analyzed to investigate the alterations in cell proliferation and cell growth. Proliferation rate was quantified by counting the number of cells per millimeter in the specific cell files, and was found to be higher in microgravity-grown samples than in the control 1 g. Cell growth was appraised through the rate of ribosome biogenesis, assessed by morphological and morphometrical parameters of the nucleolus and by the levels of the nucleolar protein nucleolin. All these parameters showed a depletion of the rate of ribosome production in microgravity-grown samples versus samples grown at 1 g. The results show that growth in microgravity induces alterations in essential cellular functions. Cell growth and proliferation, which are strictly associated functions under normal ground conditions, appeared divergent after gravity modification; proliferation was enhanced, whereas growth was depleted. We suggest that the cause of these changes could be an alteration in the cell cycle regulation, at the levels of checkpoints regulating cell cycle progression, leading to a shortened G2 period.

Low-shear modelled microgravity alters expression of virulence determinants of Staphylococcus aureus

Microbiological monitoring of air and surfaces within the ISS indicate that bacteria of the genus Staphylococcus are found with high frequency. Staphylococcus aureus, an opportunistic pathogen with the capacity to cause severe debilitating infection, constitutes a significant proportion of these isolates. Experiments conducted during short-term flight suggest that growth in microgravity leads to increases in bacterial antibiotic resistance and to cell wall changes. Growth under low-shear modelled microgravity (LSMMG) indicated that a reduced gravitational field acts as an environmental signal for expression of enhanced bacterial virulence in gram-negative pathogens. We therefore examined the effect of simulated microgravity on parameters of antibiotic susceptibility and virulence in methicillin-susceptible S. aureus isolates RF1, RF6 and RF11; these strains were grown in a high aspect ratio vessel under LSMMG and compared with cells grown under normal gravity (NG). There were no significant differences in antibiotic susceptibility of staphylococci grown under LSMMG compared to NG. LSMMG-induced reductions in synthesis of the pigment staphyloxanthin and the major virulence determinant α-toxin were noted. Significant changes in global gene expression were identified by DNA microarray analysis; with isolate RF6, the expression of hla and genes of the regulatory system saeR/saeS were reduced approximately two-fold. These data provide strong evidence that growth of S. aureus under modelled microgravity leads to a reduction in expression of virulence determinants.

Melt Structural Self-Organization and Viscosity within the Transient Layer during Crystal Growth in Microgravity

Melt Structural Self-Organization and Viscosity within the Transient Layer during Crystal Growth in Microgravity