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NASA NTRS2007

Deep Space Network Scheduling Using Evolutionary Computational Methods

The paper presents the specific approach taken to formulate the problem in terms of gene encoding, fitness function, and genetic operations. The genome is encoded such that a subset of the scheduling constraints is automatically satisfied. Several fitness functions are formulated to emphasize different aspects of the scheduling problem. The optimal solutions of the different fitness functions demonstrate the trade-off of the scheduling problem and provide insight into a conflict resolution process.

Deep Space Network (DSN)↗
NASA NTRS2007

Testing Prospects for Reliable Diatom Nanotechnology in Microgravity

The worldwide effort to grow nanotechnology, rather than use lithography, focuses on diatoms, single cell eukaryotic algae with ornate silica shells, which can be replaced by oxides and ceramics, or reduced to elemental silicon, to create complex nanostructures with compositions of industrial and electronics importance. Diatoms produce an enormous variety of structures, some of which are microtubule dependent and perhaps sensitive to microgravity. The NASA Single Loop for Cell Culture (SLCC) for culturing and observing microorganisms permits inexpensive, low labor in-space experiments. We propose to send up to the International Space Station diatom cultures of the three diatom species whose genomes are being sequenced, plus the giant diatoms of Antarctica (up to 2 mm diameter for a single cell) and the unique colonial diatom, Bacillaria paradoxa. Bacillaria cells move against each other in partial synchrony, like a sliding deck of cards, by a microfluidics mechanism. Will normal diatoms have aberrant pattern and shape or motility compared to ground controls? The generation time is typically one day, so that many generations may be examined from one flight. Rapid, directed evolution may be possible running the SLCC as a compustat. The shell shapes and patterns are preserved in hard silica, so that the progress of normal and aberrant morphogenesis may be followed by drying samples on a moving filter paper "diatom tape recorder". With a biodiversity of 100,000 distinct species, diatom nanotechnology may offer a compact and portable nanotechnology toolkit for exploration anywhere.

Gordon, Richard↗
NASA NTRS2006

The 'Biologically-Inspired Computing' Column

The field of Biology changed dramatically in 1953, with the determination by Francis Crick and James Dewey Watson of the double helix structure of DNA. This discovery changed Biology for ever, allowing the sequencing of the human genome, and the emergence of a "new Biology" focused on DNA, genes, proteins, data, and search. Computational Biology and Bioinformatics heavily rely on computing to facilitate research into life and development. Simultaneously, an understanding of the biology of living organisms indicates a parallel with computing systems: molecules in living cells interact, grow, and transform according to the "program" dictated by DNA. Moreover, paradigms of Computing are emerging based on modelling and developing computer-based systems exploiting ideas that are observed in nature. This includes building into computer systems self-management and self-governance mechanisms that are inspired by the human body's autonomic nervous system, modelling evolutionary systems analogous to colonies of ants or other insects, and developing highly-efficient and highly-complex distributed systems from large numbers of (often quite simple) largely homogeneous components to reflect the behaviour of flocks of birds, swarms of bees, herds of animals, or schools of fish. This new field of "Biologically-Inspired Computing", often known in other incarnations by other names, such as: Autonomic Computing, Pervasive Computing, Organic Computing, Biomimetics, and Artificial Life, amongst others, is poised at the intersection of Computer Science, Engineering, Mathematics, and the Life Sciences. Successes have been reported in the fields of drug discovery, data communications, computer animation, control and command, exploration systems for space, undersea, and harsh environments, to name but a few, and augur much promise for future progress.

Hinchey, Mike↗
NASA NTRS2009

Biomorphic Multi-Agent Architecture for Persistent Computing

A multi-agent software/hardware architecture, inspired by the multicellular nature of living organisms, has been proposed as the basis of design of a robust, reliable, persistent computing system. Just as a multicellular organism can adapt to changing environmental conditions and can survive despite the failure of individual cells, a multi-agent computing system, as envisioned, could adapt to changing hardware, software, and environmental conditions. In particular, the computing system could continue to function (perhaps at a reduced but still reasonable level of performance) if one or more component( s) of the system were to fail. One of the defining characteristics of a multicellular organism is unity of purpose. In biology, the purpose is survival of the organism. The purpose of the proposed multi-agent architecture is to provide a persistent computing environment in harsh conditions in which repair is difficult or impossible. A multi-agent, organism-like computing system would be a single entity built from agents or cells. Each agent or cell would be a discrete hardware processing unit that would include a data processor with local memory, an internal clock, and a suite of communication equipment capable of both local line-of-sight communications and global broadcast communications. Some cells, denoted specialist cells, could contain such additional hardware as sensors and emitters. Each cell would be independent in the sense that there would be no global clock, no global (shared) memory, no pre-assigned cell identifiers, no pre-defined network topology, and no centralized brain or control structure. Like each cell in a living organism, each agent or cell of the computing system would contain a full description of the system encoded as genes, but in this case, the genes would be components of a software genome.

Lodding, Kenneth N.↗
NASA NTRS2008

Mice Drawer System

The Mice Drawer System (MDS) is an Italian Space Agency (ASI) facility which is able to support mice onboard the International Space Station during long-duration exploration missions (from 100 to 150-days) by living space, food, water, ventilation and lighting. Mice can be accommodated either individually (maximum 6) or in groups (4 pairs). MDS is integrated in the Space Shuttle middeck during transportation (uploading and downloading) to the ISS and in an EXPRESS Rack in Destiny, the US Laboratory during experiment execution. Osteoporosis is a debilitating disease that afflicts millions of people worldwide. One of the physiological changes experienced by astronauts during space flight is the accelerated loss of bone mass due to the lack of gravitational loading on the skeleton. This bone loss experienced by astronauts is similar to osteoporosis in the elderly population. MDS will help investigate the effects of unloading on transgenic (foreign gene that has been inserted into its genome to exhibit a particular trait) mice with the Osteoblast Stimulating Factor-1, OSF-1, a growth and differentiation factor, and to study the genetic mechanisms underlying the bone mass pathophysiology. MDS will test the hypothesis that mice with an increased bone density are likely to be more protected from osteoporosis, when the increased bone mass is a direct effect of a gene involved in skeletogenesis (skeleton formation). Osteoporosis is a debilitating disease that afflicts millions worldwide. One of the physiological changes experienced by astronauts during space flight is the accelerated loss of bone mass due to the lack of gravitational loading on the skeleton, a loss that is similar to osteoporosis in the elderly population on Earth. Osteoblast Stimulating Factor-1 (OSF-1), also known as pleiotrophin (PTN) or Heparin-Binding Growth- Associated Molecule (HB-GAM) belongs to a family of secreted heparin binding proteins..OSF-1 is an extracellular matrix-associated growth and differentiation factor that is normally expressed in cartilage; it can stimulate the proliferation and differentiation of human osteoprogenitor cells (cell that differentiate into an osteoblast) in vitro. The Mice Drawer System will study the effects of microgravity on transgenic mouse bones in order to identify genetic mechanisms playing a role in the reduction of the bone mass observed in humans and animals as a consequence of long-duration (greater than 100 days) microgravity exposure. Onboard the ISS, MDS is relatively self-sufficient; a crewmember will check the health status of the rodents on a daily basis, by assessing them through the viewing window. Water levels will be assessed by the crew daily and refilled as needed. Replacement of the food bars and replacement of the waste filters will be conducted inflight by crewmembers every 20-days.

Cancedda, Ranieri↗
NASA NTRS2009

Development of a GCR Event-based Risk Model

A goal at NASA is to develop event-based systems biology models of space radiation risks that will replace the current dose-based empirical models. Complex and varied biochemical signaling processes transmit the initial DNA and oxidative damage from space radiation into cellular and tissue responses. Mis-repaired damage or aberrant signals can lead to genomic instability, persistent oxidative stress or inflammation, which are causative of cancer and CNS risks. Protective signaling through adaptive responses or cell repopulation is also possible. We are developing a computational simulation approach to galactic cosmic ray (GCR) effects that is based on biological events rather than average quantities such as dose, fluence, or dose equivalent. The goal of the GCR Event-based Risk Model (GERMcode) is to provide a simulation tool to describe and integrate physical and biological events into stochastic models of space radiation risks. We used the quantum multiple scattering model of heavy ion fragmentation (QMSFRG) and well known energy loss processes to develop a stochastic Monte-Carlo based model of GCR transport in spacecraft shielding and tissue. We validated the accuracy of the model by comparing to physical data from the NASA Space Radiation Laboratory (NSRL). Our simulation approach allows us to time-tag each GCR proton or heavy ion interaction in tissue including correlated secondary ions often of high multiplicity. Conventional space radiation risk assessment employs average quantities, and assumes linearity and additivity of responses over the complete range of GCR charge and energies. To investigate possible deviations from these assumptions, we studied several biological response pathway models of varying induction and relaxation times including the ATM, TGF -Smad, and WNT signaling pathways. We then considered small volumes of interacting cells and the time-dependent biophysical events that the GCR would produce within these tissue volumes to estimate how GCR event rates mapped to biological signaling induction and relaxation times. We considered several hypotheses related to signaling and cancer risk, and then performed simulations for conditions where aberrant or adaptive signaling would occur on long-duration space mission. Our results do not support the conventional assumptions of dose, linearity and additivity. A discussion on how event-based systems biology models, which focus on biological signaling as the mechanism to propagate damage or adaptation, can be further developed for cancer and CNS space radiation risk projections is given.

Cucinotta, Francis A.↗
NASA NTRS2010

Developing Biological ISRU: Implications for Life Support and Space Exploration

Main findings: 1) supplementing very dilute media for cultivation of CB with analogs of lunar or Martian regolith effectively supported the proliferation of CB; 2) O2 evolution by siderophilic cyanobacteria cultivated in diluted media but supplemented with iron-rich rocks was higher than O2 evolution by same strain in undiluted medium; 3) preliminary data suggest that organic acids produced by CB are involved in iron-rich mineral dissolution; 4) the CB studied can accumulate iron on and in their cells; 4) sequencing of the cyanobacterium JSC-1 genome revealed that this strain possesses molecular features which make it applicable for the cultivation in special photoreactors on Moon and Mars. Conclusion: As a result of pilot studies, we propose, to develop a concept for semi-closed integrated system that uses CB to extract useful elements to revitalize air and produce valuable biomolecules. Such a system could be the foundation of a self-sustaining extraterrestrial outpost (Hendrickx, De Wever et al., 2005; Handford, 2006). A potential advantage of a cyanobacterial photoreactor placed between LSS and ISRU loops is the possibility of supplying these systems with extracted elements and compounds from the regolith. In addition, waste regolith may be transformed into additional products such as methane, biomass, and organic and inorganic soil enrichment for the cultivation of higher plants.

Brown, I. I.↗
NASA NTRS2010

Studying Prokaryotic Communities in Iron Depositing Hot Springs (IDHS): Implication for Early Mars Habitability

We speculate that both external and intracellular iron precipitate in iron-tolerant CB might be involved in oxidative stress suppression shown by [9]. Significant differences are apparent between a set of proteins involved in the maintenance of Fe homeostasis and oxidative stress protection in iron-tolerant and fresh-water and marine CB. Correspondingly, these properties may help to make iron-tolerant CB as dominant organisms in IDHS and probably on early Earth and Mars. Further comparative analyses of hot springs metagenomes and the genomes of iron-tolerant microbes versus fresh-water/marine ones may point out to different habitable zones on early Mars.

Sarkisova, S. A.↗
NASA NTRS2010

Distributions of Low- and High-LET Radiation-Induced Breaks in Chromosomes are Associated with Inter- and Intrachromosome Exchanges

To study the breakpoint along the length of the chromosome induced by low- and high-LET radiations, we exposed human epithelial cells in vitro to Cs-137 rays at both low and high dose rates, secondary neutrons at a low dose rate, and 600 MeV/u Fe ions at a high dose rate. The location of the breaks was identified using the multicolor banding in situ hybridization (mBAND) that paints Chromosome 3 in 23 different colored bands. The breakpoint distributions were found to be similar between rays of low and high dose rates and between the two high-LET radiation types. Detailed analysis of the chromosome break ends involved in inter- and intrachromosome exchanges revealed that only the break ends participating in interchromosome exchanges contributed to the hot spots found for low-LET. For break ends participating in intrachromosome exchanges, the distributions for all four radiation scenarios were similar with clusters of breaks found in three regions. Analysis of the locations of the two break ends in Chromosome 3 that joined to form an intrachromosome exchange demonstrated that two breaks with a greater genomic separation may be more likely to rejoin than two closer breaks, indicating that chromatin folding can play an important role in the rejoining of chromosome breaks. Our study demonstrated that the gene-rich regions do not necessarily contain more breaks. The breakpoint distribution depends more on the likelihood that a break will join with another break in the same chromosome or in a different chromosome.

Hada, Megumi↗
NASA NTRS2000

Three-Dimensional Transgenic Cell Models to Quantify Space Genotoxic Effects

In this paper we describe a three-dimensional, multicellular tissue-equivalent model, produced in NASA-designed, rotating wall bioreactors using mammalian cells engineered for genomic containment of mUltiple copies of defined target genes for genotoxic assessment. The Rat 2(lambda) fibroblasts (Stratagene, Inc.) were genetically engineered to contain high-density target genes for mutagenesis. Stable three-dimensional, multicellular spheroids were formed when human mammary epithelial cells and Rat 2(lambda) fibroblasts were cocultured on Cytodex 3 Beads in a rotating wall bioreactor. The utility of this spheroidal model for genotoxic assessment was indicated by a linear dose response curve and by results of gene sequence analysis of mutant clones from 400micron diameter spheroids following low-dose, high-energy, neon radiation exposure

Gonda, S.↗
NASA NTRS2007

Interpreting Chromosome Aberration Spectra

Ionizing radiation can damage cells by breaking both strands of DNA in multiple locations, essentially cutting chromosomes into pieces. The cell has enzymatic mechanisms to repair such breaks; however, these mechanisms are imperfect and, in an exchange process, may produce a large-scale rearrangement of the genome, called a chromosome aberration. Chromosome aberrations are important in killing cells, during carcinogenesis, in characterizing repair/misrepair pathways, in retrospective radiation biodosimetry, and in a number of other ways. DNA staining techniques such as mFISH ( multicolor fluorescent in situ hybridization) provide a means for analyzing aberration spectra by examining observed final patterns. Unfortunately, an mFISH observed final pattern often does not uniquely determine the underlying exchange process. Further, resolution limitations in the painting protocol sometimes lead to apparently incomplete final patterns. We here describe an algorithm for systematically finding exchange processes consistent with any observed final pattern. This algorithm uses aberration multigraphs, a mathematical formalism that links the various aspects of aberration formation. By applying a measure to the space of consistent multigraphs, we will show how to generate model-specific distributions of aberration processes from mFISH experimental data. The approach is implemented by software freely available over the internet. As a sample application, we apply these algorithms to an aberration data set, obtaining a distribution of exchange cycle sizes, which serves to measure aberration complexity. Estimating complexity, in turn, helps indicate how damaging the aberrations are and may facilitate identification of radiation type in retrospective biodosimetry.

Levy, Dan↗
NASA NTRS2010

Utilization of Alternate Chirality Enantiomers in Microbial Communities

Our previous study of chirality led to interesting findings for some anaerobic extremophiles: the ability to metabolize substrates with alternate chirality enantiomers of amino acids and sugars. We have subsequently found that not just separate microbial species or strains but entire microbial communities have this ability. The functional division within a microbial community on proteo- and sugarlytic links was also reflected in a microbial diet with L-sugars and D-amino acids. Several questions are addressed in this paper. Why and when was this feature developed in a microbial world? Was it a secondary de novo adaptation in a bacterial world? Or is this a piece of genetic information that has been left in modern genomes as an atavism? Is it limited exclusively to prokaryotes, or does this ability also occur in eukaryotes? In this article, we have used a broader approach to study this phenomenon using anaerobic extremophilic strains from our laboratory collection. A series of experiments were performed on physiologically different groups of extremophilic anaerobes (pure and enrichment cultures). The following characteristics were studied: 1) the ability to grow on alternate chirality enantiomers -- L-sugars and D- amino acids; 2) Growth-inhibitory effect of alternate chirality enantiomers; 3) Stickland reaction with alternate chirality amino acids. The results of this research are presented in this paper.

Pikuta, Elena V.↗
NASA NTRS2011

New Modeling Approaches to Investigate Cell Signaling in Radiation Response

Ionizing radiation damages individual cells and tissues leading to harmful biological effects. Among many radiation-induced lesions, DNA double-strand breaks (DSB) are considered the key precursors of most early and late effects [1] leading to direct mutation or aberrant signal transduction processes. In response to damage, a flow of information is communicated to cells not directly hit by the radiation through signal transduction pathways [2]. Non-targeted effects (NTE), which includes bystander effects and genomic instability in the progeny of irradiated cells and tissues, may be particularly important for space radiation risk assessment [1], because astronauts are exposed to a low fluence of heavy ions and only a small fraction of cells are traversed by an ion. NTE may also have important consequences clinical radiotherapy [3]. In the recent years, new simulation tools and modeling approaches have become available to study the tissue response to radiation. The simulation of signal transduction pathways require many elements such as detailed track structure calculations, a tissue or cell culture model, knowledge of biochemical pathways and Brownian Dynamics (BD) propagators of the signaling molecules in their micro-environment. Recently, the Monte-Carlo simulation code of radiation track structure RITRACKS was used for micro and nano-dosimetry calculations [4]. RITRACKS will be used to calculate the fraction of cells traversed by an ion and delta-rays and the energy deposited in cells in a tissue model. RITRACKS also simulates the formation of chemical species by the radiolysis of water [5], notably the .OH radical. This molecule is implicated in DNA damage and in the activation of the transforming growth factor beta (TGF), a signaling molecule involved in NTE. BD algorithms for a particle near a membrane comprising receptors were also developed and will be used to simulate trajectories of signaling molecules in the micro-environment and characterize autocrine and paracrine cell communication and signal transduction.

Plante, Ianik↗
NASA NTRS2011

Differential Processing of Low and High LET Radiation Induced DNA Damage: Investigation of Switch from ATM to ATR Signaling

The members of the phosphatidylinositol kinase-like kinase family of proteins namely ataxia-telangiectasia mutated (ATM) and ATM- and Rad3-related (ATR) are directly responsible for the maintenance of genomic integrity by mounting DDR through signaling and facilitating the recruitment of repair factors at the sites of DNA damage along with coordinating the deployment of cell cycle checkpoints to permit repair by phosphorylating Checkpoint kinase Chk1, Chk2 and p53. High LET radiation from GCR (Galactic Cosmic Rays) consisting mainly of protons and high energy and charged (HZE) particles from SPE (Solar Particle Event) pose a major health risk for astronauts on their space flight missions. The determination of these risks and the design of potential safeguards require sound knowledge of the biological consequences of lesion induction and the capability of the cells to counter them. We here strive to determine the coordination of ATM and ATR kinases at the break sites directly affecting checkpoint signaling and DNA repair and whether differential processing of breaks induced by low and high LET radiation leads to possible augmentation of swap of these damage sensors at the sites of DNA damage. Exposure of cells to IR triggers rapid autophosphorylation of serine-1981 that causes dimer dissociation and initiates monomer formation of ATM. ATM kinase activity depends on the disruption of the dimer, which allows access and phosphorylation of downstream ATM substrates like Chk2. Evidence suggests that ATM is activated by the alterations in higher-order chromatin structure although direct binding of ATM to DSB ends may be a crucial step in its activation. On the other hand, in case of ATR, RPA (replication protein A)-coated ssDNA (single-stranded DNA) generated as a result of stalled DNA replication or during processing of chromosomal lesions is crucial for the localization of ATR to sites of DNA damage in association with ATR-interacting protein (ATRIP). Although the majority of RPA-coated ssDNA is generally present only during DNA replication, ATR activation in G1 and G2-phase might still require formation of RPA-coated ssDNA, probably initiated by the MRN-CtIP complex and then extended by the Exo1- or BLM-dependent mechanisms at the sites of DSBs. Evidence accumulates that activation of ATM and ATR are oppositely regulated by the length of single stranded overhangs generated at the break sites by processes mentioned above and these stretches of single stranded overhangs hold the clue for ATM to ATR switch at broken DNA ends. We irradiated 82-6hTERT human fibroblast cells with low LET gamma-rays and high LET Fe and Si particles. Preliminary results with cells exposed to 1Gy gamma-rays show that the kinetics of pChk2-pT68 foci formation is comparable to that of gamma-H2AX although they appear to recede quicker. The number and intensity of observed foci reaches a maximum at 30 min and 60 min post IR for Chk2-pT68 and gamma-H2AX foci respectively and all Chk2-pT68 foci colocalize with gamma-H2AX foci. The kinetics of Chk1-pS345 and ATRIP are being determined. Results of Chk2-pT68 foci kinetics was also corroborated by western blot experiments, although phosphorylation was detected as early as 10 min and started receding 30 min post IR with 2Gy of gamma-rays. On the other hand, level of ATR-pS428 reached its maximum between 60 and 120 min and was maintained until the last measured time point of 4 hours post IR as determined by western blotting. Experiments performed with high LET Fe and Si particles will be reported.

Saha, Janapriya↗
NASA NTRS2011

Chromosomal Aberrations in Normal and AT Cells Exposed to High Dose of Low Dose Rate Irradiation

Ataxia telangiectasia (A-T) is a human autosomally recessive syndrome characterized by cerebellar ataxia, telangiectases, immune dysfunction, and genomic instability, and high rate of cancer incidence. A-T cell lines are abnormally sensitive to agents that induce DNA double strand breaks, including ionizing radiation. The diverse clinical features in individuals affected by A-T and the complex cellular phenotypes are all linked to the functional inactivation of a single gene (AT mutated). It is well known that cells deficient in ATM show increased yields of both simple and complex chromosomal aberrations after high-dose-rate irradiation, but, less is known on how cells respond to low-dose-rate irradiation. It has been shown that AT cells contain a large number of unrejoined breaks after both low-dose-rate irradiation and high-dose-rate irradiation, however sensitivity for chromosomal aberrations at low-dose-rate are less often studied. To study how AT cells respond to low-dose-rate irradiation, we exposed confluent normal and AT fibroblast cells to up to 3 Gy of gamma-irradiation at a dose rate of 0.5 Gy/day and analyzed chromosomal aberrations in G0 using fusion PCC (Premature Chromosomal Condensation) technique. Giemsa staining showed that 1 Gy induces around 0.36 unrejoined fragments per cell in normal cells and around 1.35 fragments in AT cells, whereas 3Gy induces around 0.65 fragments in normal cells and around 3.3 fragments in AT cells. This result indicates that AT cells can rejoin breaks less effectively in G0 phase of the cell cycle? compared to normal cells. We also analyzed chromosomal exchanges in normal and AT cells after exposure to 3 Gy of low-dose-rate rays using a combination of G0 PCC and FISH techniques. Misrejoining was detected in the AT cells only? When cells irradiated with 3 Gy were subcultured and G2 chromosomal aberrations were analyzed using calyculin-A induced PCC technique, the yield of unrejoined breaks decreased in both normal and AT cells and misrejoined breaks increased in both cell lines. The present study suggests that AT cells begin to rejoin breaks when a certain number of breaks are accumulated and an increased number of exchanges were observed in G0 AT cells, which is similar situation after high-dose-rate irradiation.

Kawata, T.↗
NASA NTRS2011

Atomic Simulation of Complex DNA DSBs and the Interactions with the Ku70/80 Heterodimer

DNA double strand breaks (DSBs) induced by ionizing radiation (IR) usually contain modified bases such as 8-oxo-7,8-dihydroguanine (8-oxoG) and thymine glycol, apurinic/apyrimidinic (AP) sites, 2-deoxyribonolactone, or single-strand breaks (SSBs). The presence of such lesions in close proximity to the DSB terminus makes the DNA nicks more difficult to repair and rejoin than endogenously induced simple DSBs, and as such a major determinant of the biological effects of high linear energy transfer (LET) radiation as encountered in space travel. In this study we conducted molecular dynamics simulations on a series of DNA duplexes with various complex lesions of 8-oxoG and AP sites, in an effort to investigate the effects of such lesions to the structural integrity and stability of DNA after insulted by IR. We also simulated the interaction of such complex DSBs with the Ku70/80 heterodimer, the first protein in mammalian cells to embark the non-homologous end joining (NHEJ) DNA repair pathway. The results indicate, compared to DNA with simple DSBs, the complex lesions can enhance the hydrogen bonds opening rate at the DNA terminus, and increase the mobility of the whole duplex, thus they present more deleterious effects to the genome integrity if not captured and repaired promptly in cells. Simulations also demonstrate the binding of Ku drastically reduces structural disruption and flexibility caused by the complex lesions, and the interactions of Ku with complex DSBs have a different potential energy landscape from the bound structure with simple DSB. In all complex DSBs systems, the binding of DSB terminus with Ku70 is softened while the binding of the middle duplex with Ku80 is tightened. This energy shift may help the Ku protein to secure at the DSB terminus for a longer time, so that other end processing factors or repair pathways can proceed at the lesions before NHEJ repair process starts. These atomic simulations may provide valuable new insight into the selective action of repair proteins on damaged DNA.

Hu, Shaowen↗
NASA NTRS2004

Oligodeoxynucleotide Probes for Detecting Intact Cells

A rapid, sensitive test using chemiluminescent oligodeoxynucleotide probes has been developed for detecting, identifying, and enumerating intact cells. The test is intended especially for use in detecting and enumerating bacteria and yeasts in potable water. As in related tests that have been developed recently for similar purposes, the oligodeoxynucleotide probes used in this test are typically targeted at either singlecopy deoxyribonucleic acid (DNA) genes (such as virulence genes) or the multiple copies (10,000 to 50,000 copies per cell) of 16S ribosomal ribonucleic acids (rRNAs). Some of those tests involve radioisotope or fluorescent labeling of the probes for reporting hybridization of probes to target nucleic acids. Others of those tests involve labeling with enzymes plus the use of chemiluminescent or chromogenic substrates to report hybridization via color or the emission of light, respectively. The present test is of the last-mentioned type. The chemiluminescence in the present test can be detected easily with relatively simple instrumentation. In developing the present test, the hybridization approach was chosen because hybridization techniques are very specific. Hybridization detects stable, inheritable genetic targets within microorganisms. These targets are not dependent on products of gene expression that can vary with growth conditions or physiological states of organisms in test samples. Therefore, unique probes can be designed to detect and identify specific genera or species of bacteria or yeast (in terms of rRNA target sequences) or can be designed to detect and identify virulence genes (genomic target sequences). Because of the inherent specificity of this system, there are few problems of cross-reactivity. Hybridization tests are rapid, but hybridization tests now available commercially lack sensitivity; typically, between 10(exp 6) and 10(exp 7) cells of the target organism are needed to ensure a reliable test. Consequently, the numbers of target bacteria in samples are usually amplified by overnight pre-enrichment growth. These tests are usually performed in laboratories by skilled technicians. The present test was designed to overcome the shortcomings of the commercial hybridization tests. The figure summarizes the major steps of the test. It is important to emphasize that the hybridization process used in this test differs from that of other hybridization tests in that it does not extract target nucleic acids. This process is based on intact-cell hybridization (so-called in situ hybridization ), wherein the intact cells act as immobilizing agents. The cells are identified and enumerated by measuring the chemiluminescence emitted from alkaline phosphatase-probe (AP-probe) hybridization; the chemiluminescence is detected or measured by use of photographic film or a luminometer, respectively.

Rosson, Reinhardt A.↗
NASA NTRS2011

Early and Late Damages in Chromosome 3 of Human Lymphocytes After Radiation Exposure

Tumor formation in humans or animals is a multi-step process. An early stage of cancer development is believed to be genomic instability (GI) which accelerates the mutation rate in the descendants of the cells surviving radiation exposure. GI is defined as elevated or persistent genetic damages occurring many generations after the cells are exposed. While early studies have demonstrated radiation-induced GI in several cell types as detected in endpoints such as mutation, apoptosis and damages in chromosomes, the dependence of GI on the quality of radiation remains uncertain. To investigate GI in human lymphocytes induced by both low- and high-LET radiation, we initially exposed white blood cells collected from healthy subjects to gamma rays in vitro, and cultured the cells for multiple generations. Chromosome aberrations were analyzed in cells collected at first mitosis post irradiation and at several intervals during the culture period. Among a number of biological endpoints planned for the project, the multi-color banding fluorescent in situ hybridization (mBAND) allows identification of inversions that were expected to be stable. We present here early and late chromosome aberrations detected with mBAND in chromosome 3 after gamma exposure. Comparison of chromosome damages in between human lymphocytes and human epithelial cells is also discussed

Sunagawa, Mayumi↗