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At least 19 records
All sourcesNASA NTRSDOE OSTI
NASA NTRS2015

Transient Gene and miRNA Expression Profile Changes of Confluent Human Fibroblast Cells in Space

Microgravity or an altered gravity environment from the static 1 gravitational constant has been shown to influence global gene expression patterns and protein levels in cultured cells. However, most of the reported studies conducted in space or using simulated microgravity on the ground have focused on the growth or differentiation of the cells. Whether non-dividing cultured cells will sense the presence of microgravity in space has not been specifically addressed. In an experiment conducted on the International Space Station, confluent human fibroblast cells were fixed after being cultured in space for 3 and 14 days for investigations of gene and miRNA (microRNA) expression profile changes in these cells. A fibroblast is a type of cell that synthesizes the extracellular matrix and collagen, the structural framework for tissues, and plays a critical role in wound healing and other functions. Results of the experiment showed that on Day 3, both the flown and ground cells were still proliferating slowly even though they were confluent, as measured by the expression of the protein Ki-67 positive cells, and the cells in space grew slightly faster. Gene and miRNA expression data indicated activation of NF(sub kappa)B (nuclear factor kappa-light-chain-enhancer of activated B cells) and other growth related pathways involving HGF and VEGF in the flown cells. On Day 14 when the cells were mostly non-dividing, the gene and miRNA expression profiles between the flight and ground samples were indistinguishable. Comparison of gene and miRNA expressions in the Day 3 samples in respect to Day 14 revealed that most of the changes observed on Day 3 were related to cell growth for both the flown and ground cells. Analysis of cytoskeleton changes by immunohistochemistry staining of the cells with antibodies for alpha-tubulin showed no difference between the flight and ground samples. Results of our study suggest that in true non-dividing human fibroblast cells, microgravity in space has little effect on the gene and miRNA expression. Gene and miRNA expression changes were observed in cells that were confluent, but still proliferating slowly. The faster growth in the flown cells was associated with the activation of NF(sub kappa)B pathways which triggers the expression of several growth factors and the suppression of the cell cycle checkpoint.

Zhang, Ye↗
NASA NTRS2015

Wetlab-2 - Quantitative PCR Tools for Spaceflight Studies of Gene Expression Aboard the International Space Station

Wetlab-2 is a research platform for conducting real-time quantitative gene expression analysis aboard the International Space Station. The system enables spaceflight genomic studies involving a wide variety of biospecimen types in the unique microgravity environment of space. Currently, gene expression analyses of space flown biospecimens must be conducted post flight after living cultures or frozen or chemically fixed samples are returned to Earth from the space station. Post-flight analysis is limited for several reasons. First, changes in gene expression can be transient, changing over a timescale of minutes. The delay between sampling on Earth can range from days to months, and RNA may degrade during this period of time, even in fixed or frozen samples. Second, living organisms that return to Earth may quickly re-adapt to terrestrial conditions. Third, forces exerted on samples during reentry and return to Earth may affect results. Lastly, follow up experiments designed in response to post-flight results must wait for a new flight opportunity to be tested.

ISS research platform↗
NASA NTRS1999

High-efficiency Agrobacterium-mediated transformation of Norway spruce (Picea abies) and loblolly pine (Pinus taeda)

Agrobacterium-mediated gene transfer is the method of choice for many plant biotechnology laboratories; however, large-scale use of this organism in conifer transformation has been limited by difficult propagation of explant material, selection efficiencies and low transformation frequency. We have analyzed co-cultivation conditions and different disarmed strains of Agrobacterium to improve transformation. Additional copies of virulence genes were added to three common disarmed strains. These extra virulence genes included either a constitutively active virG or extra copies of virG and virB, both from pTiBo542. In experiments with Norway spruce, we increased transformation efficiencies 1000-fold from initial experiments where little or no transient expression was detected. Over 100 transformed lines expressing the marker gene beta-glucuronidase (GUS) were generated from rapidly dividing embryogenic suspension-cultured cells co-cultivated with Agrobacterium. GUS activity was used to monitor transient expression and to further test lines selected on kanamycin-containing medium. In loblolly pine, transient expression increased 10-fold utilizing modified Agrobacterium strains. Agrobacterium-mediated gene transfer is a useful technique for large-scale generation of transgenic Norway spruce and may prove useful for other conifer species.

NASA Discipline Plant Biology↗
NASA NTRS2011

Stress-Driven Selection of Novel Phenotypes

A process has been developed that can confer novel properties, such as metal resistance, to a host bacterium. This same process can also be used to produce RNAs and peptides that have novel properties, such as the ability to bind particular compounds. It is inherent in the method that the peptide or RNA will behave as expected in the target organism. Plasmid-born mini-gene libraries coding for either a population of combinatorial peptides or stable, artificial RNAs carrying random inserts are produced. These libraries, which have no bias towards any biological function, are used to transform the organism of interest and to serve as an initial source of genetic variation for stress-driven evolution. The transformed bacteria are propagated under selective pressure in order to obtain variants with the desired properties. The process is highly distinct from in vitro methods because the variants are selected in the context of the cell while it is experiencing stress. Hence, the selected peptide or RNA will, by definition, work as expected in the target cell as the cell adapts to its presence during the selection process. Once the novel gene, which produces the sought phenotype, is obtained, it can be transferred to the main genome to increase the genetic stability in the organism. Alternatively, the cell line can be used to produce novel RNAs or peptides with selectable properties in large quantity for separate purposes. The system allows for easy, large-scale purification of the RNAs or peptide products. The process has been reduced to practice by imposing sub-inhibitory concentrations of NiCl2 on cells of the bacterium Escherichia coli that were transformed separately with the peptide library and RNA library. The evolved resistant clones were isolated, and sequences of the selected mini-gene variants were established. Clones resistant to NiCl2 were found to carry identical plasmid variants with a functional mini-gene that specifically conferred significant nickel tolerance on the host cells. Sequencing of the selected mini-gene revealed a propensity of the encoded peptide to bind transient metal ions. Expression of the mini-gene markedly improved growth parameters of the evolved clones at sub-inhibitory concentrations of NiCl2 while being slightly detrimental in the absence of stress. Similar results have been obtained with the RNA libraries. Overall, the results demonstrate a very natural outcome of the selection experiments in which the mini-genes were expected to be either successfully integrated into bacterial genetic networks, or rejected depending upon their effect on host fitness. This described approach can be useful as a laboratory model to study the dynamics of bacterial adaptive evolution on the molecular level. It can also provide a strategy for screening expressed DNA libraries in search of novel genes with desirable properties.

Fox, George E.↗
NASA NTRS2011

System Re-set: High LET Radiation or Transient Musculoskeletal Disuse Cause Lasting Changes in Oxidative Defense Pathways Within Bone

Six months post-IR, there were no notable changes in skeletal expression of 84 principal genes in the p53 signaling pathway due to low dose IR (0.5Gy), HU, or both. In contrast, numerous genes relevant to oxidative stress were regulated by the treatments, typically in a direction indicative of increased oxidative stress and impaired defense. IR and HU independently reduced (between 0.46 to 0.88 fold) expression levels of Noxa1, Gpx3, Prdx2, Prdx3, and Zmynd17. Surprisingly, transient HU alone (sham-irradiated) decreased expression of several redox-related genes (Gpx1,Gstk1, Prdx1, Txnrd2), which were not affected significantly by IR alone. Irradiation increased (1.13 fold) expression of a gene responsible for production of superoxides by neutrophils (NCF2). Of interest, only combined treatment with HU and IR led to increased expression levels of Ercc2, (1.19 fold), a DNA excision repair enzyme. Differences in gene expression levels may reflect a change in gene expression on a per cell basis, a shift in the repertoire of specific cell types within the tissue, or both. Serum nitrite/nitrate levels were elevated to comparable levels (1.6-fold) due to IR, HU or both, indicative of elevated systemic nitrosyl stress. CONCLUSIONS The magnitude of changes in skeletal expression of oxidative stress-related genes six months after irradiation and/or transient unloading tended to be relatively modest (0.46-1.15 fold), whereas the p53 pathway was not affected. The finding that many different oxidative stress-related genes differed from controls at this late time point implicates a generalized impairment of oxidative defense within skeletal tissue, which coincides with both profound radiation damage to osteoprogenitors/stem cells in bone marrow and impaired remodeling of mineralized tissue.

Kumar, Akhilesh↗
NASA NTRS2020

The NASA Twins Study: The Effect of One Year in Space on Long-Chain Fatty Acid Desaturases and Elongases

Background: To date, there is no clear understanding of the effect of long-duration spaceflight on the major enzymes that govern the metabolism of omega-6 and omega-3 fatty acids. To address this gap in knowledge, we used data from the NASA Twins Study, which includes a multi-scale omic investigation of the changes that occurred during a year-long (340 days) human spaceflight. Embedded within the NASA Twins data are specific analytes associated with fatty acid metabolism. Objectives: To examine the long-chain fatty acid desaturases and elongases in a single human during one year in space. Method: One male twin was on board the International Space Station (ISS) for one year, while his monozygotic twin served as a genetically matched ground control. Longitudinal assessments included the genome, epigenome, transcriptome, proteome, metabolome, microbiome, and immunome during the mission, as well as six months before and after. The gene-specific fatty acid desaturase and elongase transcriptome data (FADS1, FADS2, ELOVL2 and ELOVL5) were extracted from untargeted RNA-seq measurements derived from white blood cell fractions. Results: Most data from the elongases and desaturases exhibited relatively similar expression profiles (R2>0.6) over time for the CD8, CD19, and LD cell fractions, indicating overall conservation of function within and between the subjects. Both cell-type and temporal specificity was observed in some cases, and some differences were also apparent between the poly-adenylated fraction (polyA) of processed RNAs vs. the ribo-depleted (ribo-) fraction. The flight subject showed a stronger enrichment of the Fatty Acid Metabolic processes pathway across almost all cell types (columns, CD4, CD8, CPT, LD), most especially in the ribodepleted fraction of RNA, but also with the polyA+ fraction of RNA. GSEA enrichment measures across three related Fatty Acid Metabolism pathways showed a differential between the ground and flight subject. Conclusions: There appears to be no persistent alteration of desaturase and elongase gene expression associated with one year in space. However, these data provide evidence that cellular lipid metabolism can be responsive and dynamic to spaceflight, even though it appears cell-type- and context-specific, most notably in terms of the fraction of RNA measured and the collection protocols. These results also provide new evidence of mid-flight spikes in expression of selected genes, which may indicate transient responses to specific insults during spaceflight.

Elongase↗
NASA NTRS1997

Targeted gene expression without a tissue-specific promoter: creating mosaic embryos using laser-induced single-cell heat shock

We have developed a method to target gene expression in the Drosophila embryo to a specific cell without having a promoter that directs expression in that particular cell. Using a digitally enhanced imaging system to identify single cells within the living embryo, we apply a heat shock to each cell individually by using a laser microbeam. A 1- to 2-min laser treatment is sufficient to induce a heat-shock response but is not lethal to the heat-shocked cells. Induction of heat shock was measured in a variety of cell types, including neurons and somatic muscles, by the expression of beta-galactosidase from an hsp26-lacZ reporter construct or by expression of a UAS target gene after induction of hsGAL4. We discuss the applicability of this technique to ectopic gene expression studies, lineage tracing, gene inactivation studies, and studies of cells in vitro. Laser heat shock is a versatile technique that can be adapted for use in a variety of research organisms and is useful for any studies in which it is desirable to express a given gene in only a distinct cell or clone of cells, either transiently or constitutively, at a time point of choice.

Non-NASA Center↗
NASA NTRS1999

Rapid and systemic accumulation of chloroplast mRNA-binding protein transcripts after flame stimulus in tomato

It has been shown that tomato (Lycopersicon esculentum) plants respond to flame wounding and electrical stimulation by a rapid (15 min) and systemic up-regulation of proteinase inhibitor (pin) genes. To find other genes having a similar expression pattern, we used subtractive cDNA screening between flamed and control plants to select clones up-regulated by flame wounding. We report the characterization of one of them, a chloroplast mRNA-binding protein encoded by a single gene and expressed preferentially in the leaves. Systemic gene expression in response to flaming in the youngest terminal leaf exhibited three distinct phases: a rapid and transient increase (5-15 min) in transcript accumulation, a decline to basal levels (15-45 min), and then a second, more prolonged increase (60-90 min). In contrast, after a mechanical wound the rapid, transient increase (5 min) was followed by a rapid decline to basal levels but no later, prolonged accumulation. In the petiole, the initial flame-wound-evoked transient increase (15 min) was followed by a continuous decline for 3 h. The nature of the wound signal(s) causing such rapid changes in transcript abundance is discussed in relation to electrical signaling, which has recently been implicated in plant responses to wounding.

NASA Discipline Plant Biology↗
NASA NTRS2010

Immune System Dysregulation and Herpesvirus Reactivation Persist During Long-Duration Spaceflight

Background: Immunity, latent herpesvirus reactivation, physiological stress and circadian rhythms were assessed during six month spaceflight onboard ISS. Blood and saliva samples were collected early, mid and late in-flight and returned for immediate analysis. Mid-point study data (10 of 17 planned subjects) will be presented. Results: Some shifts in leukocyte distribution occurred during flight, including alterations in CD8+ T cell maturation. General T cell function was consistently reduced early in-flight. Levels CD8+/IFNg+ producing T cells were depressed early in-flight, and immediately upon landing. Persistent mitogen-dependant reductions were observed in IFNg, IL-17a, IL-10, TNFa and IL-6 production. Monocyte production of IL-10 was reduced, whereas IL-8 levels were increased. Levels of mRNA for the TNFa, IL-6 and IFNg were transiently elevated early in-flight, and the dynamics of TNF and IL-6 gene expression were somewhat antagonistic to their corresponding receptors during flight. The number of virus-specific CD8+ T-cells was measured using MHC tetramers, while their function was measured using intracellular cytokine analysis following peptide stimulation. Both the number and function of EBV-specific cells decreased during flight as compared to preflight levels. The number of CMV-specific T-cells generally increased as the mission progressed while their function was variable. Viral (EBV) load in blood was elevated postflight. Anti-EBV VCA antibodies were significantly elevated by R+0; anti-EA antibodies were not significantly elevated at landing; and anti-CMV antibodies were somewhat elevated during flight. Higher levels of salivary EBV DNA were found during flight. VZV DNA reactivation occurred in ~50 % of astronauts during flight, continuing for up to 30 days post-flight. CMV was shed in 35 % the in-flight and 30% of postflight urine samples of the crewmembers. There was generally a higher level of cortisol as measured in urine and saliva in the astronauts during flight, but plasma cortisol was relatively unchanged during flight. Circadian rhythm of salivary cortisol was altered during flight. Conclusion. Some alterations in immunity do not resolve during six month spaceflight, consequentially resulting in persistent herpesvirus reactivation. Ongoing immune dysregulation may represent specific clinical risks for exploration-class space missions.

Crucian, B. E.↗
NASA NTRS2002

Regulation of collagenase-3 and osteocalcin gene expression by collagen and osteopontin in differentiating MC3T3-E1 cells

Both collagenase-3 and osteocalcin mRNAs are expressed maximally during the later stages of osteoblast differentiation. Here, we demonstrate that collagenase-3 mRNA expression in differentiating MC3T3-E1 cells is dependent upon the presence of ascorbic acid, is inhibited in the presence of the collagen synthesis inhibitor, 3,4-dehydroproline, and is stimulated by growth on collagen in the absence of ascorbic acid. Transient transfection studies show that collagenase-3 promoter activity increases during cell differentiation and requires the presence of ascorbic acid. Additionally, we show that, in differentiating MC3T3-E1 cells, collagenase-3 gene expression increases in the presence of an anti-osteopontin monoclonal antibody that binds near the RGD motif of this protein, whereas osteocalcin expression is inhibited. Furthermore, an RGD peptidomimetic compound, designed to block interaction of ligands to the alpha(v) integrin subunit, increases osteocalcin expression and inhibits collagenase-3 expression, suggesting that the RGD peptidomimetic initiates certain alpha(v) integrin signaling in osteoblastic cells. Overall, these studies demonstrate that stimulation of collagenase-3 expression during osteoblast differentiation requires synthesis of a collagenous matrix and that osteopontin and alpha(v) integrins exert divergent regulation of collagenase-3 and osteocalcin expression during osteoblast differentiation.

NASA Discipline Musculoskeletal↗
NASA NTRS1993

Molecular Basis of the Increase in Invertase Activity Elicited by Gravistimulation of Oat-Shoot Pulvini

An asymmetric (top vs. bottom) increase in invertase activity is elicited by gravistimulation in oatshoot pulvini starting within 3h after treatment. In order to analyze the regulation of invertase gene expression in this system, we examined the effect of gravistimulation on invertase mRNA induction. Total RNA and poly(A)(+)RNA, isolated from oat pulvini, and two oligonucleotide primers, corresponding to two conserved amino-acid sequences (NDPNG and WECPD) found in invertase from other species, were used for the Polymerase Chain Reaction (PCR). A partial-length cDNA (550 base pairs) was obtained and characterized. There was a 52 % deduced amino-acid sequence homology to that of carrot beta-fructosi- dase and a 48 % homology to that of tomato invertase. Northern blot analysis showed that there was an obvious transient accumulation of invertase mRNA elicited by gravistimulation of oat pulvini. The mRNA was rapidly induced to a maximum level at 1h following gravistimulation treatment and gradually decreased afterwards. The mRNA level in the bottom half of the oat pulvinus was significantly higher (five-fold) than that in the top half of the pulvinus tissue. The induction of invertase mRNA was consistent with the transient enhancement of invertase activity during the graviresponse of the pulvinus. These data indicate that the expression of the invertase gene(s) could be regulated by gravistimulation at the transcriptional and/or translational levels. Southern blot analysis showed that there were four genomic DNA fragments hybridized to the invertase cDNA. This suggests that an invertase gene family may exist in oat plants.

Wu, Liu-Lai↗
NASA NTRS1993

Kinetic Induction of Oat Shoot Pulvinus Invertase mRNA by Gravistimulation and Partial cDNA Cloning by the Polymerase Chain Reaction

An asymmetric (top vs. bottom halves of pulvini) induction of invertase mRNA by gravistimulation was analyzed in oat shoot pulvini. Total RNA and poly(A)(+) RNA, isolated from oat pulvini, and two oli-gonucleotide primers, corresponding to two conserved amino acid sequences (NDPNG and WECPD) found in invertase from other species, were used for the polymerase chain reaction (PCR). A partial length cDNA (550 bp) was obtained and characterized. A 62% nucleotide sequence homology and 58% deduced amino acid sequence homology, as compared to beta-fructosidase of carrot cell wall, was found. Northern blot analysis showed that there was an obviously transient induction of invertase mRNA by gravistimulation in the oat pulvinus system. The mRNA was rapidly induced to a maximum level at 1 hour after gravistimulation treatment and gradually decreased afterwards. The mRNA level in the bottom half of the oat pulvinus was significantly higher than that in the top half of the pulvinus tissue. The kinetic induction of invertase mRNA was consistent with the transient accumulation of invertase activity during the graviresponse of the pulvinus. This indicates that the expression of the invertase gene(s) could be regulated by gravistimulation at the transcriptional level. Southern blot analysis showed that there were two to three genomic DNA fragments which hybridized with the partial-length invertase cDNA.

Wu, Liu-Lai↗
NASA NTRS2001

TP53-dependent chromosome instability is associated with transient reductions in telomere length in immortal telomerase-positive cell lines

Telomere shortening in telomerase-negative somatic cells leads to the activation of the TP53 protein and the elimination of potentially unstable cells. We examined the effect of TP53 gene expression on both telomere metabolism and chromosome stability in immortal, telomerase-positive cell lines. Telomere length, telomerase activity, and chromosome instability were measured in multiple clones isolated from three related human B-lymphoblast cell lines that vary in TP53 expression; TK6 cells express wild-type TP53, WTK1 cells overexpress a mutant form of TP53, and NH32 cells express no TP53 protein. Clonal variations in both telomere length and chromosome stability were observed, and shorter telomeres were associated with higher levels of chromosome instability. The shortest telomeres were found in WTK1- and NH32-derived cells, and these cells had 5- to 10-fold higher levels of chromosome instability. The primary marker of instability was the presence of dicentric chromosomes. Aneuploidy and other stable chromosome alterations were also found in clones showing high levels of dicentrics. Polyploidy was found only in WTK1-derived cells. Both telomere length and chromosome instability fluctuated in the different cell populations with time in culture, presumably as unstable cells and cells with short telomeres were eliminated from the growing population. Our results suggest that transient reductions in telomere lengths may be common in immortal cell lines and that these alterations in telomere metabolism can have a profound effect on chromosome stability. Copyright 2000 Wiley-Liss, Inc.

Non-NASA Center↗
NASA NTRS2016

Gene Expression and Structural Skeletal Responses to Long-Duration Simulated Microgravity in Rats

Spaceflight has deleterious effects on skeletal structure and function, specifically causingprofound loss in bone mass, density, and strength, as well as changes in expression levels of genes related to oxidative stress [Hyeon et al., Smith et al.]. It is known that bone resorption remains elevated after spaceflight and that bone density and strength fail to recover completely even years following spaceflight [Smith et al., Carpenter et al.]. However, our current understanding of the signaling pathways and molecular mechanisms that control bone loss and that link oxidative stress, bone resorption, and mechanical unloading of skeletal tissue is incomplete. Here, we aim to examine skeletal responses to simulated long-duration spaceflight on bone loss using the ground-based hindlimb unloading (HU) model in adult (9 months old) male rats. We hypothesized that simulated microgravity leads to the temporal regulation of oxidative-defense genes and pro-osteoclastogenic factors, showing progression and eventual plateau during long-term unloading, and that transient changes at early timepoints in these pathways precede skeletal adaptations to long-duration unloading. We will identify oxidativestress and bone resorption-related changes using global gene expression analysis (Affymetrix arrays) for both acute (within 14 days) and long-term timepoints (90 days). We will also use quantitative PCR to examine changes in expression of genes related to oxidative metabolism (e.g. Nrf2, SOD-1), bone turnover (resorption and formation markers, e.g. TRAP, osteocalcin respectively, SOST), and osteoclastogenesis (e.g. RANKL, OPG) at both early and late timepoints. We will then use detailed microarchitectural and structural analysis through microcomputed tomography to relate gene expression changes with structural changes in bone, expecting that plateaus in gene expression correlate with long-term changes in bone microarchitecture.

Rael, Victoria E.↗
NASA NTRS1998

Calcium signals and calcium channels in osteoblastic cells

Calcium (Ca2+) channels are present in non-excitable as well as in excitable cells. In bone cells of the osteoblast lineage, Ca2+ channels play fundamental roles in cellular responses to external stimuli including both mechanical forces and hormonal signals. They are also proposed to modulate paracrine signaling between bone-forming osteoblasts and bone-resorbing osteoclasts at local sites of bone remodeling. Calcium signals are characterized by transient increases in intracellular Ca2+ levels that are associated with activation of intracellular signaling pathways that control cell behavior and phenotype, including patterns of gene expression. Development of Ca2+ signals is a tightly regulated cellular process that involves the concerted actions of plasma membrane and intracellular Ca2+ channels, along with Ca2+ pumps and exchangers. This review summarizes the current state of knowledge concerning the structure, function, and role of Ca2+ channels and Ca2+ signals in bone cells, focusing on the osteoblast.

Non-NASA Center↗
NASA NTRS1999

Developmental regulation of the gene for chimeric calcium/calmodulin-dependent protein kinase in anthers

Chimeric Ca(2+)/calmodulin-dependent protein kinase (CCaMK) was cloned from developing anthers of lily (Lilium longiflorum Thumb. cv. Nellie White) and tobacco (Nicotiana tabacum L. cv. Xanthi). Previous biochemical characterization and structure/function studies had revealed that CCaMK has dual modes of regulation by Ca(2+) and Ca(2+)/calmodulin. The unique structural features of CCaMK include a catalytic domain, a calmodulin-binding domain, and a neural visinin-like Ca(2+)-binding domain. The existence of these three features in a single polypeptide distinguishes it from other kinases. Western analysis revealed that CCaMK is expressed in a stage-specific manner in developing anthers. Expression of CCaMK was first detected in pollen mother cells and continued to increase, reaching a peak around the tetrad stage of meiosis. Following microsporogenesis, CCaMK expression rapidly decreased and at later stages of microspore development, no expression was detected. A tobacco genomic clone of CCaMK was isolated and transgenic tobacco plants were produced carrying the CCaMK promoter fused to the beta-glucuronidase reporter gene. Both CCaMK mRNA and protein were detected in the pollen sac and their localizations were restricted to the pollen mother cells and tapetal cells. Consistent results showing a stage-specific expression pattern were obtained by beta-glucuronidase analysis, in-situ hybridization and immunolocalization. The stage- and tissue-specific appearance of CCaMK in anthers suggests that it could play a role in sensing transient changes in free Ca(2+) concentration in target cells, thereby controlling developmental events in the anther.

NASA Discipline Plant Biology↗
NASA NTRS1994

Altered skeletal pattern of gene expression in response to spaceflight and hindlimb elevation

Spaceflight leads to osteopenia, in part by inhibiting bone formation. Using an animal model (hindlimb elevation) that simulates the weightlessness of spaceflight, we and others showed a reversible inhibition of bone formation and bone mineralization. In this study, we have measured the mRNA levels of insulin-like growth factor I (IGF-I), IGF-I receptor (IGF-IR), alkaline phosphatase, and osteocalcin in the tibiae of rats flown aboard National Aeronautics and Space Administration Shuttle Flight STS-54 and compared the results with those obtained from their ground-based controls and from the bones of hindlimb-elevated animals. Spaceflight and hindlimb elevation transiently increase the mRNA levels for IGF-I, IGF-IR, and alkaline phosphatase but decrease the mRNA levels for osteocalcin. The changes in osteocalcin and alkaline phosphatase mRNA levels are consistent with a shift toward decreased maturation, whereas the rise in IGF-I and IGF-IR mRNA levels may indicate a compensatory response to the fall in bone formation. We conclude that skeletal unloading during spaceflight or hindlimb elevation resets the pattern of gene expression in the osteoblast, giving it a less mature profile.

NASA Discipline Musculoskeletal↗
NASA NTRS1992

Parathyroid hormone induces c-fos and c-jun messenger RNA in rat osteoblastic cells

PTH is a potent regulator of osteoblast gene expression, yet the nuclear events that mediate PTH action are poorly understood. We were interested in identifying immediate early genes which may regulate PTH-altered gene expression in the osteoblast. Therefore, we examined the effects of PTH on c-fos and c-jun gene expression in a rat osteoblastic cell line (UMR 106-01). Under control conditions, c-fos and c-jun mRNAs were present at low basal levels. After PTH treatment, c-fos mRNA abundance dramatically increased, with a maximal and transient response at 30 min. PTH also stimulated an increase in c-jun mRNA, but in a biphasic manner, with maximal levels at 30 min and 2 h. These responses were dose dependent, not altered by cotreatment with the protein synthesis inhibitor cycloheximide, and preceded PTH-induced expression of matrix metallo-proteinase-1 mRNA. Nuclear run-on assays demonstrated an increased rate of c-fos and c-jun transcription after PTH exposure. To determine the signal transduction pathways involved, second messenger analogs were tested for their ability to mimic the effects of PTH. 8-Bromo-cAMP and phorbol 12-myristate 13-acetate (PMA) caused increases in the abundance of c-fos and c-jun transcripts. Ionomycin had no effect on the expression of these genes. Pretreatment of the cells with PMA resulted in a decrease in basal c-jun expression, but did not alter the PTH-mediated increase in c-fos, c-jun, or matrix metalloproteinase-1 mRNAs.(ABSTRACT TRUNCATED AT 250 WORDS).

Non-NASA Center↗