The wild type WS plants and the Sku5 mutant plants in the WS background were germinated on ISS or on the ground and the gene expression profiles in roots at 4 days or 8 days were established. The Sku5 gene (AT4G12420) codes for a multi-copper oxidase-like protein SKU5 protein that is involved in directed root tip growth. The Sku5 protein is glycosylated GPI-anchored and localizes to the plasma membrane and the cell wall. The Sku5 gene is expressed most strongly in expanding tissues. The development of WS and Sku5 plants on orbit differs from that on the ground as demonstrated by the comparison of the gene expression profiles between 4 days old to 8 days old plant in the two environments. However the development of Sku5 mutant plants also differs from WS at either age and in either environment suggesting the role of the genetic background in these developmental decisions. The 4 days old Sku5 roots in orbit engaged substantially more genes than the 4 days old WS roots in orbit but also more than the 4 days old Sku5 roots on the ground. Overall the 4 days old roots differentially expressed more genes in spaceflight relative to ground than the 8 days old roots of either genotype. Finally the 4 days old Sku5 roots in orbit differ in 862 genes from the 8 days Sku5 roots in orbit while on the ground there is merely half of the number of genes differentially expressed between the 4 days and 8 days developmental stages of the Sku5 roots. APEX03-2 (Advanced Plant Experiment 03-2) also identified as TAGES-Isa (Transgenic Arabidopsis Gene Expression System xe2 x80 x94Intracellular Signaling Architecture) was launched on SpaceX mission CRS-5 on 10 January 2015. Dry sterilized Arabidopsis seeds were planted aseptically on the surface of 10-cm2 solid media plates and remained dormant until removed from cold stowage and exposed to light at the initiation of the experiment on the ISS (International Space Station). The plates were grown in the Vegetable Production System (VPS/Veggie) hardware on the Columbus Module of the ISS with the overhead LED lighting of the VPS. At four or for the second experimental set at eight days seedlings were harvested by an astronaut into KFT (Kennedy Fixation Tube) containing RNAlater solutions. Upon return to Earth the harvested material was used to compare the transcriptomes of each genotype and each age using RNAseq technology. The patterns of gene expression was compared between treatments (spaceflight versus ground control) within each genotype of same age or between two ages within same genotype and within same treatment. For 4 day old roots 5-8 roots were combined to serve as one biological replica. For 8 day old roots 2-3 roots were used to form one biological replica. In each case the four biological replicas were used for the transcriptomic analysis.

Interaction of COP1 and UVR8 which regulate UV-B-induced photomorphogenesis and stress acclimation in Arabidopsis thaliana.

The root apex is an important section of the plant root involved in environmental sensing and cellular development. Analyzing the gene profile of root apex in diverse environments is important and challenging especially when the samples are limiting and precious such as in spaceflight. The feasibility of using tiny root sections for transcriptome analysis was examined in this study. To understand the gene expression profiles of the root apex Arabidopsis thaliana Col-0 roots were sectioned into Zone-I (0.5 mm root cap and meristematic zone) and Zone-II (1.5 mm transition elongation and growth terminating zone). Gene expression was analyzed using microarray and RNA seq. Both the techniques arrays and RNA-Seq identified 4180 common genes as differentially expressed (with > two-fold changes) between the zones. In addition 771 unique genes and 19 novel TARs were identified by RNA-Seq as differentially expressed which were not detected in the arrays. Single root tip zones can be used for full transcriptome analysis; further the root apex zones are functionally very distinct from each other. RNA-Seq provided novel information about the transcripts compared to the arrays. These data will help optimize transcriptome techniques for dealing with small rare samples.

The Heat Shock Factor A2 (HsfA2) as a part of the HSF network is essential to the plant s response to almost any environmental stress and to the cellular homeostatic control mechanisms. Plant cell cultures disabled in HsfA2 function were grown aboard the International Space Station (ISS) in order to ascertain whether or not they use the same terrestrially effective systems to adapt to the novel environment of spaceflight. Cultured lines of Arabidopsis thaliana derived from wild type (WT) cultivar Col-0 and from a knock-out line deficient in the gene encoding HSFA2 (HSFA2 KO) were launched to the ISS on SpaceX-2 as part of the Cellular Expression Logic (CEL) experiment of the BRIC17 spaceflight mission and were fixed in-flight after 10 days on orbit. Microarray gene expression data were analyzed using a two-part comparative approach. First differentially expressed genes were identified between the environments (spaceflight to ground) within cells of the same genotype which represented physiological adaptation to the spaceflight environment. Second gene expression profiles were identified between the genotypes (HSFA2 KO to WT) within the same environment defining genes uniquely required by the two genotypes in the ground and spaceflight adapted states. The physiological state of the cells as a result of disabling a gene has tremendous control over the mechanisms induced to adapt to the environment of spaceflight. The HsfA2 demonstrated a role in the physiological adaptation to the spaceflight environment since the cells disabled in the HsfA2 gene used substantially different genes to achieve the spaceflight adapted state than the WT cells. The endoplasmic reticulum (ER) stress and unfolded protein response (UPR) define the HSFA2 KO cells physiological state regardless of the environment and likely result from the deficiency in the chaperone-mediated protein folding machinery. HsfA2 seems to have a universal stress response role but also specific roles in the physiological adaptation to spaceflight through cell wall remodeling signal perception and transduction and starch biosynthesis. Implementation of knock-out cells identified a set of genes with a required expression level in order for a cell to achieve a spaceflight-adapted state. The HSFA2 KO cells helped to unravel the HsfA2-dependent genes of the adaption of wild type cells to the environment of spaceflight.

Whole seedlings of wild type (4d) and atm mutants (4d) have been analyzed after a gamma ray irradiation of 0.75h 1.5h 3h & 5h (time course). Roots of wt (4d) atm (3d) and atr (4d) mutants have been analyzed after a 1h irradiation. Ataxia Telangiectasia Mutated (ATM) encodes a large protein with a phosphatidylinositol 3-kinase (PI3K)-like domain at the C terminus (reviewed by Rotman and Shiloh 1998). PI3K-related proteins make up a large family of Ser-Thr protein kinases numerous members of which are involved in the regulation of cell cycle progression responses to DNA damage and the maintenance of genomic stability (Hoekstra 1997). AtATM plays an essential role in meiosis and in the somatic response to DNA damage in plants similar to the function of ATM in mammals and other eukaryotes. Ataxia telangiectasia-mutated and Rad3-related (ATR) plays a central role in cell-cycle regulation transmitting DNA damage signals to downstream effectors of cell-cycle progression.

When germinated and grown on-board the ISS (International Space Station) plant do not exhibit abnormal structures but they do have altered growth habits and this project aims to investigate the molecular mechanisms that provide the foundation for the altered growth habits observed in orbit. APEX03-2 (Advanced Plant Experiment 03-2) also known as TAGES-ISA (Transgenic Arabidopsis Gene Expression System-Intracellular Signaling Architecture) specifically addresses the growth and molecular changes that occur in Arabidopsis thaliana plants during spaceflight by using molecular and genetic tools and by asking fundamental questions regarding root structure growth and cell wall remodeling may be answered. This investigation advances the fundamental understanding of the molecular biological responses to extraterrestrial environments. This understanding helps to further define the impacts of spaceflight on biological systems to better enable NASA x92s future space exploration goals.

The Cosmic Ray Exposure Sequencing Science (CRESS) payload system is a proof of concept experiment to assess the genomic impact of space radiation on seeds. CRESS was designed as a secondary payload for the December 2016 high-altitude high-latitude and long-duration balloon flight carrying the Boron And Carbon Cosmic Rays in the Upper Stratosphere (BACCUS) experimental hardware. Investigation of the biological effects of Galactic Cosmic Radiation (GCR) particularly those of ions with High-Z and Energy (HZE) is of interest due to the genomic damage this type of radiation inflicts. The biological effects of upper-stratospheric mixed radiation above Antarctica (ANT) were sampled using Arabidopsis thaliana seeds and were compared to those resulting from a controlled simulation of GCR at Brookhaven National Laboratory (BNL) and to laboratory control seed. The payload developed for Antarctica exposure was broadly designed to 1U CubeSat specifications (10cmx10cmx10cm <1.33kg) maintained 1 atm internal pressure and carried an internal cargo of four seed trays (about 580,000 seeds) and twelve CR-39 Solid-State Nuclear Track Detectors (SSNTDs). The irradiated seeds were recovered sterilized and grown on Petri plates for phenotypic screening. BNL and ANT M0 seeds showed significantly reduced germination rates and elevated somatic mutation rates when compared to non-irradiated controls with the BNL mutation rate also being significantly higher than that of ANT. Genomic DNA from mutants of interest was evaluated with whole-genome sequencing using PacBio SMRT technology. Sequence data revealed the presence of an array of genome structural variants in the genomes of M0 and M1 mutant plants.

Adult and neonatal human cardiovacular progenitor cell clonal populations were flown aboard the ISS for 12 days prior to fixation in RNAprotect. Gene expression analysis was performed against clone- patient- and passage-matched ground control samples. qPCR gene expression profiling. Human CPCs from three neonates and three adults were individually cultured on the ground or aboard the ISS.

Gene expression changes induced by acute skeletal muscle unloading which leads to physiological changes including muscle atrophy fibre-type switching and loss of ability to transition between lipid and glucose as energy source (metabolic inflexibility) was investigated by hind-limb suspension (HLS) treatment of Male ICR mice (28 x9632 g body wt; Harlan Indianapolis IN). Agilent Whole Mouse Genome Oligo Microarrays were utilised to examine the effects of HLS on mRNA expression profiles of the soleus muscle and the gastrocnemius muscle in the hindlimbs of freely ambulating control and 24h HLS treated mice.

Adaptation of humans in low gravity conditions is a matter of utmost importance when efforts are on to a gigantic leap in human space expeditions for tourism and formation of space colonies. In this connection cardiovascular adaptation in low gravity is a critical component of human space exploration. Deep high-throughput sequencing approach allowed us to analyze the miRNA and mRNA expression profiles in human umbilical cord vein endothelial cells (HUVEC) cultured under gravity (G) and stimulated microgravity (MG) achieved with a clinostat. The present study identified totally 1870 miRNAs differentially expressed in HUVEC under MG condition when compared to the cells subjected to unitary G conditions. The functional association of identified miRNAs targeting specific mRNAs revealed that miRNAs hsa-mir-496 hsa-mir-151a hsa-miR-296-3p hsa-mir-148a hsa-miR-365b-5p hsa-miR-3687 hsa-mir-454 hsa-miR-155-5p and hsa-miR-145-5p differentially regulated the genes involved in cell adhesion angiogenesis cell cycle JAK-STAT signaling MAPK signaling nitric oxide signaling VEGF signaling and wound healing pathways. Further the q-PCR based experimental studies of upregulated and downregulated miRNA and mRNAs demonstrate that the above reported miRNAs influence the cell proliferation and vascular functions of the HUVEC in MG conditions effectively. Consensus on the interactome results indicates restricted fluctuations in the transcriptome of the HUVEC exposed to short-term MG that could lead to higher levels of endothelial functions like angiogenesis and vascular patterning.