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NASA NTRS1985

NASA Developmental Biology Workshop: A summary

The Life Sciences Division of the National Aeronautics and Space Administration (NASA) as part of its continuing assessment of its research program, convened a workshop on Developmental Biology to determine whether there are important scientific studies in this area which warrant continued or expanded NASA support. The workshop consisted of six panels, each of which focused on a single major phylogenetic group. The objectives of each panel were to determine whether gravity plays a role in the ontogeny of their subject group, to determine whether the microgravity of spaceflight can be used to help understand fundamental problems in developmental biology, to develop the rationale and hypotheses for conducting NASA-relevant research in development biology both on the ground and in space, and to identify any unique equipment and facilities that would be required to support both ground-based and spaceflight experiments.

Souza, K. A.↗
NASA NTRS1985

What are mycoplasmas - The relationship of tempo and mode in bacterial evolution

In phenotype the mycoplasmas are very different from ordinary bacteria. However, genotypically (i.e., phylogenetically) they are not. On the basis of ribosomal RNA homologies the mycoplasmas belong with the clostridia, and indeed have specific clostridial relatives. Mycoplasmas are, however, unlike almost all other bacteria in the evolutionary characteristics of their ribosomal RNAs. These RNAs contain relatively few of the highly conserved oligonucleotide sequences characteristic of normal eubacterial ribosomal RNAs. This is interpreted to be a reflection of an elevated mutation rate in mycoplasma lines of descent. A general consequence of this would be that the variation associated with a mycoplasma population is augmented both in number and kind, which in turn would lead to an unusual evolutionary course, one unique in all respects. Mycoplasmas, then, are actually tachytelic bacteria. The unusual evolutionary characteristics of their ribosomal RNAs are the imprints of their rapid evolution.

Woese, C. R.↗
NASA NTRS1986

The rRNA evolution and procaryotic phylogeny

Studies of ribosomal RNA primary structure allow reconstruction of phylogenetic trees for prokaryotic organisms. Such studies reveal major dichotomy among the bacteria that separates them into eubacteria and archaebacteria. Both groupings are further segmented into several major divisions. The results obtained from 5S rRNA sequences are essentially the same as those obtained with the 16S rRNA data. In the case of Gram negative bacteria the ribosomal RNA sequencing results can also be directly compared with hybridization studies and cytochrome c sequencing studies. There is again excellent agreement among the several methods. It seems likely then that the overall picture of microbial phylogeny that is emerging from the RNA sequence studies is a good approximation of the true history of these organisms. The RNA data allow examination of the evolutionary process in a semi-quantitative way. The secondary structures of these RNAs are largely established. As a result it is possible to recognize examples of local structural evolution. Evolutionary pathways accounting for these events can be proposed and their probability can be assessed.

Fox, G. E.↗
NASA NTRS1986

Functional and evolutionary relationships between bacteriorhodopsin and halorhodopsin in the archaebacterium, halobacterium halobium

The archaebacteria occupy a unique place in phylogenetic trees constructed from analyses of sequences from key informational macromolecules, and their study continues to yield interesting ideas on the early evolution and divergence of biological forms. It is now known that the halobacteria among these species contain various retinal-proteins, resembling eukaryotic rhodopsins, but with different functions. Two of these pigments, located in the cytoplasmic membranes of the bacteria, are bacteriorhodopsin (a light-driven proton pump) and halorhodopsin (a light-driven chloride pump). Comparison of these systems is expected to reveal structure/function relationships in these simple (primitive?) energy transducing membrane components and evolutionary relationships which had produced the structural features which allow the divergent functions. Findings indicate that very different primary structures are needed for these proteins to accomplish their different functions. Indeed, analysis of partial amino acid sequences from halo-opsin shows already that few if any long segments exist which are homologous to bacterio-opsin. Either these proteins diverged a very long time ago to allow for the observed differences, or the evolutionary clock in the halobacteria runs faster than usual.

Lanyi, J. K.↗
NASA NTRS1986

Evolutionary significance of osmoregulatory mechanisms in cyanobacteria

Physiological processes of all life forms on this planet are intrinsically related to their intracellular water potential. The overall goal was the elucidation of the mechanism(s) whereby the first oxygenic phtoautotrophs (the cyanobacteria) adjust their water potential to that of a changing external water potential (that is, osmoregulate). Osmoregulation is achieved by intracellular adjustment of inorganic and/or organic solutes (osmolytes) involving specific biochemical mechanisms. Structural and biochemical evolution within the cyanobacteria is believed completed (and fixed in present day forms) by the end of the Precambrain eon. Therefore, research using cyanobacteria of all three structural types (unicellular, filamentous, and branched), each grown in the photoautotrophic (PA), photoheterotrophic (PG), and chemotrophic (CH) modes of nutrition, should provide insight into the origin and evolution of the photosynthetically related osmoregulatory mechanisms of eukaryotic organisms. The chloroplasts of these organisms are phylogenetically related to the cyanobacteria.

Yopp, J. H.↗
NASA NTRS1987

A possible biochemical missing link among archaebacteria

The characteristics of the newly discovered strain of archaebacteria, VC-16, the only archaebacterium known to reduce sulfate, suggest that VC-16 might represent a transitional form between an anaerobic thermophilic sulfur-based type of metabolism and methanogenesis. It is shown here, using a matrix of evolutionary distances derived from an alignment of various archaebacterial 16S rRNAs and the phylogenetic tree derived from these evolutionary distances, that the lineage represented by strain VC-16 arises from the archaebacterial tree precisely where such an interpretation would predict that it would, between the Methanococcus lineage and that of Thermococcus.

Achenbach-Richter, Laurie↗
NASA NTRS1990

Characterization of marine macroalgae by fluorescence signatures

The feasibility of distinguishing macroalgal classes by their fluorescence signatures was investigated using narrow-waveband light to excite groups of accessory pigments in brown, red, and green macroalgae and measuring fluorescence emission at 685 nm. Results obtained on 20 marine macroalgae field-collected samples showed that fluorescence excitation signatures were relatively uniform within phylogenetic classes but were substantially different for different classes. It is suggested that it may be possible to characterize the type and the abundance of subtidal macroalgae from low-flying aircraft using existing laser-induced fluorescence methodology.

Topinka, J. A.↗
NASA NTRS1992

The solar system: Importance of research to the biological sciences

An attempt is made to describe the scope of scientific areas that comprise the current field of exobiology in the United States. From investigations of astrophysical phenomena that deal with the birth of stars and planetary systems to questions of molecular biology involving phylogenetic relationships among organisms, from attempts to simulate the synthesis of biological precursor molecules in the chemistry laboratory to making measurements of the organic constituents of Titan's atmosphere, these researches all converge toward a common objective--answering the question of how life came about in the universe.

Klein, Harold P.↗
NASA NTRS1994

Spectral composition of light and growing of plants in controlled environments

The main conclusions of many investigations about general requirements of plants for spectral composition of PAR (photosynthetically active radiation) are based on phylogenetic aspects of plant growth. We think that these aspects are not the main criteria in choosing the spectral composition required for growing plants in controlled conditions. Our approach to this problem is based on plant and crop reaction under long duration growth with specific spectra and intensity. Only in this way can we determine correctly the role of light characteristics for developing crops.

Tikhomirov, Alexander A.↗
NASA NTRS1996

Basic Gravitational Reflexes in the Larval Frog

This investigation was designed to determine how a primitive vertebrate, the bullfrog tadpole, is able to sense and process gravitational stimuli. Because of the phylogenetic similarities of the vestibular systems in all vertebrates, the understanding of the gravitational reflexes in this relatively simple vertebrate should elucidate a skeletal framework on a elementary level, upon which the more elaborate reflexes of higher vertebrates may be constructed. The purpose of this study was to understand how the nervous system of the larval amphibian processes gravitational information. This study involved predominantly electrophysiological investigations of the isolated, alert (forebrain removed) bullfrog tadpole head. The focus of these experiments is threefold: (1) to understand from whole extraocular nerve recordings the signals sent to the eye following static gravitational tilt of the head; (2) to localize neuronal centers responsible for generating these signals through reversible pharmacological ablation of these centers; and (3) to record intracellularly from neurons within these centers in order to determine the single neuron's role in the overall processing of the center. This study has provided information on the mechanisms by which a primitive vertebrate processes gravitational reflexes.

Cochran, Stephen L.↗
NASA NTRS1988

Phenotype and Age Differences in Blood Gas Characteristics, Electrolytes, Hemoglobin, Plasma Glucose and Cortisol in Female Squirrel Monkeys

Due to its small size, lower cost, tractable nature, successful breeding in captivity and its status near the middle of the primate phylogenetic scale, the squirrel monkey has become an attractive primate model for basic and biomedical research. Although the squirrel monkey now is being used more extensively in many laboratories with diverse interests, only fragmentary reports have been published regarding basic physiological characteristics, or baseline blood reference values of different phenotypes, particularly blood gases, hematology and serum chemical constituents. It is becoming recognized increasingly that these baseline blood reference values are important not only in the care and maintenance of the squirrel monkey, but are critical for assessing normal physiological status, as well as the effects of various experimental treatments. The purpose of this study was to compare differences in blood gases, electrolytes, hematology, blood glucose and cortisol among young and old Bolivian (Roman type) and Colombian (Gothic type) phenotypes of the squirrel monkey.

Brizzee, K. R.↗
NASA NTRS1993

The RNA World: Life Before DNA and Protein

All of the life that is known, all organisms that exist on Earth today or are known to have existed on Earth in the past, are of the same life form: a life form based on DNA and protein. It does not necessarily have to be that way. Why not have two competing life forms on this planet? Why not have biology as we know it and some other biology that occupies its own distinct niche? Yet that is not how evolution has played out. From microbes living on the surface of antarctic ice to tube worms lying near the deep-sea hydrothermal vents, all known organisms on this planet are of the same biology. Looking at the single known biology on Earth, it is clear that this biology could not have simply sprung forth from the primordial soup. The biological system that is the basis for all known. life is far too complicated to have arisen spontaneously. This brings us to the notion that something else something simpler, must have preceded life based on DNA and protein. One suggestion that has gained considerable acceptance over the past decade is that DNA and protein-based life was preceded by RNA-based life in a period referred to as the 'RNA world'. Even an RNA-based life form would have been fairly complicated - not as complicated as our own DNA- and protein-based life form - but far too complicated, according to prevailing scientific thinking, to have arisen spontaneously from the primordial soup. Thus, it has been argued that something else must have preceded RNA-based life, or even that there was a succession of life forms leading from the primordial soup to RNA-based life. The experimental evidence to support this conjecture is not strong because, after all, the origin of life was a historical event that left no direct physical record. However, based on indirect evidence in both the geological record and the phylogenetic record of evolutionary history on earth, it is possible to reconstruct a rough picture of what life was like before DNA and protein.

Joyce, Gerald F.↗
NASA NTRS1995

The Origin of Life--Did It Occur at High Temperatures

A high-temperature origin of life has been proposed, largely for the reason that the hyperthermophiles are claimed to be the last common ancestor of modern organisms. Even if they are the oldest extant organisms, which is in dispute, their existence can say nothing about the temperatures of the origin of life, the RNA world, and organisms preceding the hyperthermophiles. There is no geological evidence for the physical setting of the origin of life because there are no unmetamorphosed rocks from that period. Prebiotic chemistry points to a low-temperature origin because most biochemicals decompose rather rapidly at temperatures of 100 C (e.g., half-lives are 73 min for ribose, 21 days for cytosine, and 204 days for adenine). Hyperthermophiles may appear at the base of some phylogenetic trees because they outcompeted the mesophiles when they adapted to lower temperatures, possibly due to enhanced production of heat-shock proteins.

Miller, Stanley L.↗
NASA NTRS2000

Bridging Two Worlds: From the Archean to the Proterozoic

As now known, the Archean and Proterozoic appear to have been different worlds: the geology (tectonic style, basinal distribution, dominant rock types), atmospheric composition (O2, CO21, CH4), and surface environment (day-length, solar luminosity, ambient temperature) all appear to have changed over time. And virtually all paleobiologic indicators can be interpreted as suggesting there were significant biotic differences as well: (1) Stromatolites older than 2.5 Ga are rare relative to those of the Proterozoic; their biotic components are largely unknown; and the biogenicity of those older than approx. 3.2 Ga has been questioned. (2) Bona fide microfossils older than approx. 2.4 Ga are rare, poorly preserved, and of uncertain biological relations. Gaps of hundreds of millions of years in the known record make it impossible to show that Archean microorganisms are definitely part of the 2.4 Ga-to-present evolutionary continuum. and (3) In rocks older than approx. 2.2 Ga, the sulfur isotopic record is subject to controversy; phylogenetically distinctive bio-markers are unknown; and nearly a score of geologic units contain organic carbon anomalously light isotopically (relative to that of the Proterozoic and Phanerozoic) that may reflect the presence of Archaeans ("Archaebacteria of earlier classifications) but may not (since cellularly preserved Archean-age Archaeans have never been identified).

Schopf, J. William↗
NASA NTRS2000

The Evolution of Sulfide Tolerance in the Cyanobacteria

Understanding how the function of extant microorganisms has recorded both their evolutionary histories and their past interactions with the environment is a stated goal of astrobiology. We are taking a multidisciplinary approach to investigate the diversification of sulfide tolerance mechanisms in the cyanobacteria, which vary both in their degree of exposure to sulfide and in their capacity to tolerate this inhibitor of photosynthetic electron transport. Since conditions were very reducing during the first part of Earth's history and detrital sulfides have been found in Archean sediments, mechanisms conferring sulfide tolerance may have been important for the evolutionary success of the ancestors of extant cyanobacteria. Two tolerance mechanisms have been identified in this group: (1) resistance of photosystem II, the principal target of sulfide toxicity; and (2) maintenance of the ability to fix carbon despite photosystem II inhibition by utilizing sulfide as an electron donor in photosystem I - dependent, anoxygenic photosynthesis. We are presently collecting comparative data on aspects of sulfide physiology for laboratory clones isolated from a variety of habitats. These data will be analyzed within a phylogenetic framework inferred from molecular sequence data collected for these clones to test how frequently different mechanisms of tolerance have evolved and which tolerance mechanism evolved first. In addition, by analyzing these physiological data together with environmental sulfide data collected from our research sites using microelectrodes, we can also test whether the breadth of an organism's sulfide tolerance can be predicted from the magnitude of variation in environmental sulfide concentration it has experienced in its recent evolutionary past and whether greater average sulfide concentration and/or temporal variability in sulfide favors the evolution of a particular mechanism of sulfide tolerance.

Miller, Scott R.↗
NASA NTRS2001

A Structural and Molecular Approach for the Study Biomarkers

Investigation of the nucleation and growth of crystals in both abiotic and biotic systems is critical to seemingly diverse disciplines of geology, biology, environmental science, and astrobiology. While there are abundant studies devoted to the determination of the structure and composition of inorganic crystals, as well as to the development of thermodynamic and kinetic models, it is only recently that research efforts have been directed towards understanding mineralization in biological systems (i.e., biomineralization). Biomineralization refers to the processes by which living organisms form inorganic solids. Studies of the processes of biomineralization under low temperature aqueous conditions have focused primarily on magnetite forming bacteria and shell forming marine organisms. Many of the biological building materials consist of inorganic minerals (calcium carbonate, calcium phosphate, silica or iron oxide) intricately combined with organic polymers (like proteins). More recently, efforts have been undertaken to explore the nature of biological activities in ancient rocks. In the absence of well-preserved microorganisms or genetic material required for the polmerase chain reaction (PCR) method in molecular phylogenetic studies, using biominerals as biomarkers offers an alternative approach for the recognition of biogenic activity in both terrestrial and extraterrestrial environments. The primary driving force in biomineralization is the interaction between organic and inorganic phases. Thus, the investigation of the ultrastructure and the nature of reactions at the molecular level occurring at the interface between inorganic and organic phases is essential to understanding the processes leading to the nucleation and growth of crystals. It is recognized that crystal surfaces can serve as the substrate for the organization of organic molecules that lead to the formation of polymers and other complex organic molecules, and in discussions of the origins of life, is referred to as organic synthesis on mineral surfaces. Furthermore, it is suggested that the interaction between mineral surfaces and simple organic molecules resulted in the formation of amino acids, RNA, and perhaps other more complex molecules such as proteins. On the other hand, in natural systems, it is recognized that functional groups on cell walls or membranes of microorganisms serve as sites of nucleation and crystallization. The precise replication of biominerals with controlled structure, morphology, size and texture is not confined to higher organisms as it also occurs in primitive prokaryotic cells such as magnetotactic bacteria and cyanobacteria. This suggests that the principal strategies of biomineralization were established early on in the evolutionary history of organisms. It is critical, therefore, to search for common mechanisms within diverse biological systems. One such common factor is the capability for organization and self-assembly. Organic macromolecules such as proteins and lipids can aggregate and polymerize forming membranes or extracellular matrix. At the organic-inorganic interface, several factors such as lattice geometry, polarity, stereochemistry and topography may act in concert to control nucleation and growth of crystals. Although several models have been proposed that discuss the significance of these factors for biomineralization, no comprehensive experimental data are available. In contrast to crystallization in exclusively inorganic systems, the kinetics of reaction and structural relationships between organic and inorganic phases in biominerals or biomimetic material is poorly understood. For example, it is not clear if the concept of epitactic growth (geometrical matching of unit cells at the interface of a secondary crystal growing on a primary crystal) applies to organic-inorganic systems. In contrast to inorganic templates that often have a smooth and rigid surface that promotes epitactic growth, biological substrates are usually rough and result in a large degree of mismatch. It is apparent that factors controlling the reaction at the crystal-matrix interface are strongly dependent upon the nature of the substrate. Therefore, characterization of the assembled organic surface and surface structure of the inorganic phase is crucial to understanding the processes of biomineralization. The focus of our research is the investigation of the processes leading to the nucleation and growth of crystals on both natural and synthetic systems through an interdisciplinary approach that integrates molecular biology, morphology and mineralogy using advanced preparation and analytical techniques. We have studied run-products, particularly magnetite, siderite and other carbonates, that resulted from extracellular biomineralization by extremophiles isolated from a variety of extreme environments ranging from permafrost to hydrothermal vent systems. The results of this study are critical to recognizing biomarkers in terrestrial and extraterrestrial environments.

Thomas-Keprta, Kathie↗
NASA NTRS2001

Organization of P, S, and Fe Inclusions in a Freshwater Magnetococcus

Magnetotactic bacteria are a heterogeneous group of motile, mainly aquatic procaryotes that align and swim along geomagnetic field lines. They are of interest to astrobiologists because of the magnetite crystals found in the Mars meteorite ALH84001 which share many characteristics with the magnetite produced intracellularly by magnetotactic bacteria. These bacteria are diverse morphologically, physiologically and phylogenetically, sharing a few key characteristics: 1) the presence of intracellular membrane-bound magnetic crystals (magnetosomes), usually but not always arranged in chains; 2) motility by means of flagella; and 3) microaerophillic or anaerobic physiology. The bilophotrichous (having two flagella bundles) magnetotactic cocci (MC) are ubiquitous in aquatic habitats but have proven extremely difficult to cultivate. Because only several marine strains have been isolated and grown in axenic culture, little is known about the physiology and the biogeochemical roles of the MC. We studied the composition and distribution of intracellular structures in an uncultured MC, designated ARB-1. To do this, a combination of light microscopy, environmental scanning electron microscopy (ESEM), transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDS) were used. Cells of ARB-1 were separated from sediments collected from Baldwin Lake (Los Angeles Arboretum, Arcadia, CA). They are large spherical to oblate spheroidal Gram-negative cells, ranging from 1 to 4 micrometers along the maximum dimension, which is perpendicular to the direction of swimming. Cells have two large phosphorus-containing inclusions that comprise a large percentage of the cell volume. Many smaller sulfur inclusions are located at the convex end of the cell. Most of the cellular Fe is present in the magnetosomes. These may be arranged as a clump at the concave end of the cell, near the two flagella bundles, or as chains, or as both a clump and chains. The magnetosomes were identified as magnetite (Fe3O4) by selected area electron diffraction (SAED) and high resolution TEM. We saw a trend between cell size and organization of the magnetosomes. Smaller, more spherical cells were more likely to have chains than were larger, more oblate cells. This may indicate different populations of cells, or it may be attributed to variations in cell growth cycle. The size distribution (length) of magnetosomes in chains was similar to that of magnetosomes in clusters, except that there was a larger size range for clustered magnetosomes. Magnetosomes from ARB-1 cells average 82 nm in length. If plotted on a graph of length as a function of aspect ratio, they fall within the single domain region of the plot. If compared with the size distributions of magnetite from ALH84001 and magnetosomes from the cultured magnetotactic vibrio MV-1, the magnetites produced by ARB-1 cells are, on average, larger and have a wider range of aspect ratio. ARB-1 cells have a specific organization of the P, S, and Fe inclusions. The P inclusions always occupy the majority of the cell volume and separate the S inclusions from the disorganized clumps of magnetosomes and the flagella bundles. The P inclusions may contain polyphosphate, which could play several roles in motility, adaptation to stress, growth and division, buoyancy, and energy. The S inclusions might be a way to store S, a potential energy source, when the cells move from sulfide to oxygen zones. The consistency of P, S, and Fe organization in ARB-1 cells suggests that these inclusions have some specific and interactive functions.

Cox, Lea↗
NASA NTRS2001

Anaerobic Psychrophiles from Alaska, Antarctica, and Patagonia: Implications to Possible Life on Mars and Europa

Microorganisms preserved within the permafrost, glaciers, and polar ice sheets of planet Earth provide analogs for microbial life forms that may be encountered in ice or permafrost of Mars, Europa, Callisto, Ganymede, asteroids, comets or other frozen worlds in the Cosmos. The psychrophilic and psychrotolerant microbes of the terrestrial cryosphere help establish the thermal and temporal limitations of life on Earth and provide clues to where and how we should search for evidence of life elsewhere in the Universe. For this reason, the cold-loving microorganisms are directly relevant to Astrobiology. Cryo-preserved microorganisms can remain viable (in deep anabiosis) in permafrost and ice for millions of years. Permafrost, ice wedges, pingos, glaciers, and polar ice sheets may contain intact ancient DNA, lipids, enzymes, proteins, genes, and even frozen and yet viable ancient microbiota. Some microorganisms carry out metabolic processes in water films and brine, acidic, or alkaline channels in permafrost or ice at temperatures far below 0 T. Complex microbial communities live in snow, ice-bubbles, cryoconite holes on glaciers and ancient microbial ecosystems are cryopreserved within the permafrost, glaciers, and polar caps. In the Astrobiology group of the NASA Marshall Space Flight Center and the University of Alabama at Huntsville, we have employed advanced techniques for the isolation, culture, and phylogenetic analysis of many types of microbial extremophiles. We have also used the Environmental Scanning Electron Microscope to study the morphology, ultra-microstructure and chemical composition of microorganisms in ancient permafrost and ice. We discuss several interesting and novel anaerobic microorganisms that we have isolated and cultured from the Pleistocene ice of the Fox Tunnel of Alaska, guano of the Magellanic Penguin, deep sea sediments from the vicinity of the Rainbow Hydrothermal Vent and enrichment cultures from ice of the Patriot Hills of Antarctica. The microbial extremophiles recovered from permafrost, ice, cold pools and deep sea sediments may provide information relevant to the question of how and where we should search for evidence of extant or extinct microbial life elsewhere in the Cosmos.

Hoover, Richard B.↗