Life sciences: radiobiological advanced biostack experiment.

The radiobiological properties of the heavy ions of cosmic radiation were investigated on Spacelab 1 by use of biostacks, monolayers of biological test organisms sandwiched between thin foils of different types of nuclear track detectors. Biostacks were exposed to cosmic radiation at several locations with different shielding environments in the module and on the pallet. Evaluations of the physical and biological components of the experiment to date indicate that in general they survived the spaceflight in good condition. Dosimetric data are presented for the different shielding environments.

Genetic and physiological damage induced by cosmic radiation on dry plant seeds during space flight.

Total evaluation of cosmic radiation effect with or without discrimination of individualized HZE-ion effects in dry Arabidopsis seeds flown for 10 days on STS-9, yielded significant evidence for radiation damage in space. They depend on the biological criteria tested (seed germination, morphogenesis, embryo lethality, mutation rate) which stand for early, physiological and late genetic effects. They are also related to the radiation shielding environment in the space shuttle. Proceeding from these results three direct questions can be posed for present (LDEF-1) and future (ERA-1, D-2) experiments in space: What is the influence of cosmic radiation on cytogenetic repair and ontogenetic restitution processes? Does microgravity disorder the morphogenesis (i.e. growth and cell differentiation)? Is there an interaction between the effects of cosmic radiation and microgravity in eukaryotic plant systems?

Early and late damages induced by heavy charged particle irradiation in embryonic tissue of Arabidopsis seeds.

Early and late effects of accelerated heavy ions (HZE) on the embryonic tissue of Arabidopsis thaliana seeds were investigated seeing that initial cells of the plant eumeristems resemble the original cells of animal and human tissues with continuous cell proliferation. The endpoints measured were lethality and tumorization in the M1-generation for early effects and embryonic lethality in the M2-generation for late effects. The biological endpoints are plotted as functions of the physical parameters of the irradiation i.e. ion fluence (p/cm2), dose (Gray), charge Z and linear energy transfer (LET). The results presented contribute to the estimation of the principles of biological HZE effects and thus may help to develop a unified theory which could explain the whole sequence from physical and chemical reactions to biological responses connected with heavy ion radiation. Additionally, the data of this paper may be used for the discussion of the quality factor for heavy ion irradiation needed for space missions and for HZE-application in radio-therapy by use of accelerators (UNILAC, (SIS/ESR), BEVALAC).

Heavy ion induced mutations in genetic effective cells of a higher plant.

Arabidopsis thaliana offers different possibilities for investigating heavy ion induced early and late damage. Mutations in genetic effective cells can yield early damage, in the form of reduced vitality of the descending cell-lines and/or late damage, such as mutation induction visible in the following generations. Investigation is possible on different levels of ploidy (4n, 2n, n). Different genetic effective cells with equal genomes are available. Additionally, several different biological endpoints for each level of genome ploidy can be observed. Recent results of work in this field are presented.

Cosmic ionizing radiation effects in plant seeds after short and long duration exposure flights.

Recently, comparison of biophysical data obtained from orbital flights of short and long duration led to results which will be significant for long and/or repeated stay of man in space. Under orbital conditions biological stress is induced in dry seeds of Arabidopsis thaliana by cosmic radiation especially its high energetic, densely ionizing component, the heavy ions (HZE). For comparison of radiation impact during different space flights a biological attempt at estimating the impact of single particles with high mass and energy (HZE-particles) on seeds was developed. Subdivision into LET-groups showed a remarkable contribution of an intermediate group (LET = 35 to 100 keV/micrometer) due to medium heavy ions (Z = 6 to 10). Efficiency factors for radiation damage experimentally determined and assigned to different LET-classes were compared to radiation quality factors discussed in literature.

Initial approach to comparative studies on the evolutionary potentials of space radiation effects in a plant system.

The role of cosmic ionizing radiation, including heavy ions (HZE-particles) in the induction of mutations at the molecule-, chromosome-, genome- and cell-level is discussed on the basis of different DNA organization in a pro- and eukaryotically compartmented plant system (Arabidopsis thaliana (L.) Heynh.). Data recently obtained on the biological effects of ionizing radiation make it timely to discuss comparatively the evolutionary potentials of space radiation effects in the pro- and eukaryotic genomes (plasmon, plastidom, chondriom, and nucleom) during long duration exposure on space flights.

First radiobiological results of LDEF-1 experiment A0015 with Arabidopsis seed embryos and Sordaria fungus spores.

This article highlights the first results of investigations on the general vitality and damage endpoints caused by cosmic ionizing radiation in dry, dormant plant seeds of the crucifer plant Arabidopsis thaliana (L.) Heynh. and the ascomycete Sordaria fimicola after 69 month stay in space. Wild-type and mutant gene marker lines were included in Free Flyer Biostack containers and exposed on earth and side tray of the LDEF-1 satellite. The damage in biological endpoints observed in the seeds increased in the side tray sample compared to the earth tray sample. For the ascospores we found different effects depending on the biological endpoints investigated for both expositions.

Biological effects of heavy ions in Arabidopsis seeds.

Irradiation of dry seeds of Arabidopsis with heavy ions (HZE-particles) produced by UNILAC-accelerator (GSI, Darmstadt) yielded aberrations in varied developmental endpoints such as survival rate and embryo vitality. The damage increased with particle density and charge. Cross sections in the range of 0.2-1.0 micrometer2 for Ne and Ar and 2.0-10.0 micrometers2 for Xe were estimated. Soaked seeds were more sensitive than dry seeds (cross-section 2.0-10.0 micrometers2 for Ar). The induced total damage in the irradiated seeds was estimated adding the different damages weighted by certain factors. These results will be used as base data for the interpretation and evaluation of spaceflight experiments on the biological effects of cosmic radiation.

Biophysical effect of cosmic heavy ions of distinct LET-classes in a plant model system.

Results presented from recent space flight BION 9 show biological effects of different LET-classes of HZE-particles in different target regions of the seed (meristem and the whole embryo) of Arabidopsis thaliana (L.) Heyhn. HZE-one hit events and non-hit events, i.e. only hit by the low-LET background radiation, and their combined effects on the biological damage endpoint lethality are distinguished. This procedure is opening the opportunity of an approach to comparative studies of the biological effects induced by cosmic HZE-particles of different LET-ranges interacting in the complex cosmic radiation spectrum and with other space flight conditions.

Advanced biostack: experiment 1 ES 027 on Spacelab-1.

The radiobiological properties of the heavy ions of cosmic radiation were investigated on Spacelab 1 by use of biostacks, monolayers of biological test organisms sandwiched between thin foils of different types of nuclear track detectors. Biostacks were exposed to cosmic radiation at several locations with different shielding environments in the module and on the pallet. Evaluations of the physical and biological components of the experiment to date indicate that in general they survived the spaceflight in good condition. Dosimetric data are presented for the different shielding environments.

Heavy ion and cosmic radiation effects in different targets of the Arabidopsis seed.

The seed embryo of Arabidopsis thaliana, the favorite plant of up-to-date molecular developmental biology, represents a valuable model system for space radiation research with heavy ions. For several years, densely ionizing particles have been available for basic studies in nuclear physics and radiation biology through special accelerators as well as through space flight missions. Because of the great variability of the available radiation spectra, the induced relative biological effect (RBE-value) has to be differentiated with respect to the effective charge (z) and the linear energy transfer (LET) deposited in a certain volume of matter, and to the biological targets which differ in size, structure and function. The results of comparative studies of 8 space flight experiments, here by example show increasing damage produced by single heavy ions and/or cosmic background radiation (protons, neutrons, X-rays). These results offer new prospects for molecular investigation of the RBE of heavy ions in space.

Recent results of the joint ESA-DARA/IBMP experiments Biokosmos "Seeds".

Comparison of experimental data obtained from short (SDEF) and long duration exposure flights (LDEF) have recently led to results which will be significant for longer and/or repeated sojourn of man in space. Under orbital conditions biological stress and damage are induced in test subjects by cosmic radiation, especially the high energetic, densely ionizing component of heavy ions. Plant seeds were successful model systems for a biotest in studying the physiological damages and mutagenic effect caused by ionizing cosmic radiation in particular stem cells. Dosimetrically, the subdivision into charge- and Let-groups reveals the contribution of the intermediate group (LET = 350-1000 MeV/cm) due to the medium heavy ions (Z = 6-10). Their relative contribution increases with the lower inclination of the orbit of LDEF-1; on the other hand, the total fluence becomes higher with longer duration of the flight. The observed endpoints of the biological radiation damage hint at a correlation with particle dose rate rather than with the dose; additionally, data on shielding effects inside and outside the space craft and its exposure were gained from the different SDEF- and LDEF-missions.

[Physiological genetics of quantitative characters in Arabidopsis thaliana (L.) Heynh. : Part 1: segregation and biosynthesis of pigments in chlorophyll-b defect mutants]. / Genphysiologie quantitativer Merkmale bei Arabidopsis thaliana (L.) Heynh. : Teil I: Spaltungsanalyse und Pigmentbiosynthese in quantitativen Chlorophyll b-Mangelmutanten.

In biometric studies on the genetics of quantitative characters the problem of regulation of the activity of genes is rarely considered. Mutants of Arabidopsis thaliana (L.) Heynh. (Cruciferae), defective for chlorophyll b, permit a direct biochemical and physiological determination of their quantitative gene effects in the biosynthetic pathway of the plastid pigments. In agreement with other authors, the detectable genotypes are found to be based on multiple alleles (ch (+), ch (1) and ch (2)) at the ch locus. This evidence was obtained by test crosses of two mutants with the wild type and by a phaenotypic classification of the progeny on the basis of preparatory thin layer chromatography, paper chromatography tests of single plants, and quantitative spectrophotometry of the lines. In ch (1)/ch (1) the synthesis of chlorophyll b is completely blocked, in ch (2)/ch (2) only about 10 percent of the wild-type pigment is present, and in the heterozygote ch (2)/ch (1) a complementation effect is observed (i. e. superdominance of ch (2) over ch (1)) resulting in still more pigment production. In the segregating generations the variances for chlorophyll and carotinoids of all the genotypes are higher than in the parents and therefore suggest a possible polygenic background for pigment development. However, the physiological effect of the genetic background is slight as compared with the effect of the major gene ch. Preliminary experiments show that in tissue extracts of wild type leaves (ch (+)/ch (+)) there is an effective agent which enables irradiated chloroplasts of the defective ch (1)/ch (1) and ch (2)/ch (2) genotypes to synthesize chlorophyll b in vitro. Further information on the mechanism, extent and the time of action of the active agent should be obtained by varying the conditions of irradiation and by refining the tests for pigment precursors.

[Physiological genetics of quantitative characters in Arabidopsis thaliana (L.) Heynh. : Part 2: Modification of primary and secondary genetic effects of single-gene chlorophyll-b mutants by long-wave irradiation]. / Genphysiologie quantitativer Merkmale bei Arabidopsis thaliana (L.) Heynh. : Teil 2: Modifikation primärer und sekundärer genwirkungen durch langwellige strahlung bei monogenen chlorophyll b-defektmutanten.

An analysis of the quantitative molecular effects of genes requires information on the norm of reaction ("Reaktionsnorm") and on its variability in the genotype tested. 1) Experiments were carried out in an automatically controlled growth chamber to determine the optimal conditions for the norm of reaction of the plants to be tested. Genotypically determined variances between the characters studied and differences of means between test samples were most frequently significant when the growth conditions were those of an intermittent 16 hour day. 2) Changes in the transmitted radiation to wavelengths of above 500 nm or 600 nm respectively, while keeping the total radiant energy input constant, led to certain characteristic modifications in three of the mutants and in the wild type of Arabidopsis thaliana (L.) Heynh. Four ontogenetic stages were affected. The mutants are monogenic defect mutants for synthesis of chlorophyll b; two different gene loci are involved. 3) Treatment with radiation of wavelengths above 600 nm (filtered red light) results in marked developmental disorders, after 100 days (three times the normal growth period) leaf rosettes had not yet developed. Moreover, leaf pigment and dry matter production of the wild type decreased by about 20 to 30 percent as compared with production in unfiltered normal light. 4) Filtered yellow light of wavelengths above 500 nm decreases the biosynthesis of chlorophyll a and of the carotinoids and the net assimilation rate, but does not influence chlorophyll b production, leaf area index and apparent use of radiant energy. Each of the genotypes, however, shows a different reaction to the same radiation treatment. Furthermore, each shows characteristic differences in response to altered quality of radiation. The wild type and the mutant ch (2)/ch (2) show certain similarities which are not shared by the two other genotypes ch (1)/ch (1) and ch 3/ch 3. The latter two are characterized by more serious plastid defects. 5) The ch-locus has a primary effect on biosynthesis of chlorophyll b and secondary physiological effects on production of other leaf pigments and dry matter. When chlorophyll b is absent, there is no increase, but rather a reduction of the amount of chlorophyll a; however, there is no quantitative correlation between production of the two chlorophylls. The primary genetic effect of the two mutant loci is discussed, in accordance with Shibata's model of chlorophyll transformations. The secondary pleiotropic gene effects on dry matter production are probably due to additional binding of radiant energy by chlorophyll b and by the carotenoids. Complex interactions between the effects of the genes, and between gene effects and environmental effects must also be considered.

[Physiological genetics of quantitative characters in Arabidopsis thaliana (L.) Heynh. : Part 3. Dauermodifications of pleiotropic effects of homoallelic genes]. / Genphysiologie quantitativer Merkmale bei Arabidopsis thaliana (L.) Heynh. : Teil 3. Dauermodifikation in der Pleiotropie bei homoalleler Genwirkung.

The genephysiological and developmental basis of "Dauermodifications" (DM) have still not been explained. In particular definite evidence for DM in plastids is lacking up to now. Moreover no attention has been paid to the connections with the problem of quantitative gene effects. For that reason mutants of the nuclear gene ch in Arabidopsis thaliana (L.) Heynh., quantitatively defective in chlorophyll b were tested during seven generations under continuous yellow radiation (λ < 550 nm) expecting DM.The following results were obtained: 1) The main effects of the homoalleles ch (+), ch (1) and ch (2) on the reactions of chlorophyll transformation, especially on chlorophyll b-synthesis, cannot be modified permanently. 2) DM in certain accessory effects of the gene ch, i.e. carotinoid content (CC), apparent radiation energy use (EAG) and net assimilation rate (NAR) were induced by the applied radiation. These modifications are quantitative and cannot be restituted in the following generation after return to normal light.The induced DM affects phenotypic characters of the plastids which may result from biochemical regulation modified at the level of translation. The effects of multiple interactions between the allelic condition of the gene ch, the rest of the nuclear genes, the carriers of the extrachromosomal gene information, and the inducing environmental factor must be considered.

Karyotype analysis in meiosis: Giemsa banding in the genus Secale L.

Terminal bands of meiotic chromosomes stained by the Giemsa technique are permanent genetic structures of the nucleus during PMC differentiation in 8 samples of wild, primitive, and cultivated species of rye. The characteristic meiotic banding pattern is probably identical with the heterochromatic regions of mitotic chromosomes of root meristem cells (RMC) which have so far been studied. Karyotype analysis can be significantly improved by quantitative studies of the number and size of the bands combined with certain well-known chromosome characters in diplotene and diakinesis. The chromosomes involved in multivalents of some natural and synthetic species hybrids are identified for the first time. The results are discussed both in relation to the problems of chromosome evolution and their significance for marker techniques in cytogenetics.

Biological damage induced by ionizing cosmic rays in dry Arabidopsis seeds.

In September 1987 dry seeds containing embryos of the crucifer plant Arabidopsis thaliana (L.) Heynh, were flown in orbit for 13 days on the Kosmos 1887 satellite. The seeds were fixed on CNd detectors and stored in units of Biorack type I/O. One unit was exposed inside, another one outside the satellite. The temperature profile of the flown seeds inside the satellite was simulated on earth in an identical backup control sample (BC). An additional control (SC) was studied with the original seeds sample. By use of the CNd-detector, HZE-tracks were measured with a PC-assisted microscope. The biological damages were investigated by growing the seeds under controlled climatic conditions. The following biological endpoints of the cosmic radiation damage were studied: germination, radicle length, sublethality, morphological aberrations, flower development, tumorization, embryo lethality inside the siliques. The summarized damage (D) and the mutation frequencies of embyronic lethal genes were calculated. The following results were obtained: the damages increase significantly in orbit at all biological endpoints; germination and fiowerings especially, as well as embryo lethality of fruits and lethal mutation frequency, were maximum mostly for HZE-hit seeds. Additionally, an increase of damage was observed for the seeds of the outside-exposed Biorack in comparison to the inside ones, which was probably caused by less radiation shielding and free space vacuum. The significance of the results obtained is discussed with respect to stress and risk and, thus, the quality of the RBE-factors and heavy ionizing radiation all needed for the very definition of radiation protection standards in space.