Non-conservative Elements Research Articles

549. Nonconserved Sequence Motifs outside the Catalytic Core Improve the Trans-Splicing Efficiency of Anti-HIV Hammerhead Ribozymes

Trans-cleaving hammerhead ribozymes (hhRzs), where stem-loops I and III are open-ended (I/III), have been widely used for the endonucleolytic cleavage of any specific RNA substrate. However, the application of hhRzs for use in gene therapy, especially for use in the targeted knockdown of HIV gene expression and replication, has been hampered by a low intracellular catalytic efficiency. As a result, therapeutic applications that exploit the RNA Interference (RNAi) pathway are presently more attractive than ribozymes. Recent studies have shown that the catalytic efficiency of naturally-derived cis-cleaving hhRzs is several orders of magnitude higher when compared to trans-cleaving counterparts. Moreover, changes made to stem-loops I and II, which are nonconserved sequence elements peripheral to the minimal essential catalytic core, greatly affect the catalytic turnover of cis-acting hhRzs. In this study, we designed two hhRzs variants with alterations intended to mimic stem-loop interactions found in the hhRz derived from satellite RNA of tobacco ringspot virus (sTRSV) to generate trans-cleaving hhRzs targeted to an accessible region within the HIV-1 rev sequence. Kinetic analyses were performed under multiple turnover conditions using an 18-mer synthetic RNA substrate at submillimolar levels. Under standard conditions (pH 7, 37°C, 10 mM MgCl2) the rate of cleavage (kobs) for hhRz variant sII-RV2 was significantly better than that of the conventional I/III hhRz, sII-RV3, with kobs of 0.24 min-1 and 0.094 min-1 respectively. HhRz variant sII-RV1, which differed from sII-RV2 only in the position of the peripheral stem, but with an identical loop, showed almost no substrate cleavage. These results may reflect the presence of a tertiary interaction between stem-loops I and 2 that facilitate cleavage. Alternatively, the addition of a mimicked stem-loop I possibly stabilizes active hhRz conformers.

Genome-wide identification of high-affinity estrogen response elements in human and mouse.

Although estrogen receptors (ERs) recognize 15-bp palindromic estrogen response elements (EREs) with maximal affinity in vitro, few near-consensus sequences have been characterized in estrogen target genes. Here we report the design of a genome-wide screen for high-affinity EREs and the identification of approximately 70000 motifs in the human and mouse genomes. EREs are enriched in regions proximal to the transcriptional start sites, and approximately 1% of elements appear conserved in the flanking regions (-10 kb to +5 kb) of orthologous human and mouse genes. Conserved and nonconserved elements were also found, often in multiple occurrences, in more than 230 estrogen-stimulated human genes previously identified from expression studies. In genes containing known EREs, we also identified additional distal elements, sometimes with higher in vitro binding affinity and/or better conservation between the species considered. Chromatin immunoprecipitation experiments in breast cancer cell lines indicate that most novel elements present in responsive genes bind ERalpha in vivo, including some EREs located up to approximately 10 kb from transcriptional start sites. Our results demonstrate that near-consensus EREs occur frequently in both genomes and that whereas chromatin structure likely modulates access to binding sites, far upstream elements can be evolutionarily conserved and bind ERs in vivo.

Anthropogenic radionuclides in Indian Ocean surface waters—the Indian Ocean transect 1998

Results of the analyses of 3H, 90Sr, 137Cs, 239,240Pu and 241Am in surface-water samples collected during the 1998 Indian Ocean Transect cruise from New Zealand to Italy are reported and discussed. Latitudinal variations in the concentrations of these radionuclides still can be seen over 40 yr after the main input, but the observed distribution no longer corresponds to the atmospheric deposition in the study area. Low levels of conservative radionuclides characterise the Tasman Sea and the South Australian Basin. At about 30°S the concentrations increase and remain very similar until about 10°N. A slight decrease is observed in the Gulf of Aden and in the Red Sea. The distribution of transuranic elements shows marked differences with respect to that of conservative radionuclides, particularly in the Tasman Sea and in the south-eastern Indian Ocean. The observed latitudinal trend is mainly dependent on the surface oceanic circulation for conservative radionuclides, while for the non-conservative transuranic elements it is also related to primary productivity and enhanced scavenging. For all investigated radionuclides, the Mediterranean is the area with the highest concentrations. The observed radionuclide concentrations in addition to 137Cs/3H, 90Sr/3H, 137Cs/90Sr, and 241Am/239,240Pu activity ratios confirm the global origin of the radionuclides studied in surface waters, except for the area below 30°S, where the average 238Pu/239,240Pu ratios (0.20±0.04) suggest the possible influence of the SNAP Satellite that burned-up over the Indian Ocean in 1964.

Sequence elements outside the hammerhead ribozyme catalytic core enable intracellular activity.

The hammerhead ribozyme (HHRz) is a small, naturally occurring ribozyme that site-specifically cleaves RNA and has long been considered a potentially useful tool for gene silencing. The minimal conserved HHRz motif derived from natural sequences consists of three helices that intersect at a highly conserved catalytic core of 11 nucleotides. The presence of this motif is sufficient to support cleavage at high Mg2+ concentrations, but not at the low Mg2+ concentrations characteristic of intracellular environments. Here we demonstrate that natural HHRzs require the presence of additional nonconserved sequence elements outside of the conserved catalytic core to enable intracellular activity. These elements may stabilize the HHRz in a catalytically active conformation via tertiary interactions. HHRzs stabilized by these interactions cleave efficiently at physiological Mg2+ concentrations and are functional in vivo. The proposed role of these tertiary interacting motifs is supported by mutational, functional, structural and molecular modeling analysis of natural HHRzs.

Nutrients and carbon budgets for the Gulf of Lion during the Moogli cruises

Shelf-slope exchanges and budgets of organic and inorganic nutrients are calculated for the Gulf of Lion continental margin in the northwestern Mediterranean. Computations are based on data from three seasonal marine surveys performed in March 1998, June 1998 and January 1999 in the framework of the French Programme National d’Environnement Côtier. A Land-Ocean Interactions in the Coastal Zone type box model is used to calculate the advective exchanges between the shelf and the adjacent open sea and to estimate the budgets of non-conservative elements (DIP, DIN, DOP, DON, DOC, POC, PON, POP). These budgets consider river discharges, urban sewage supply, atmospheric deposition, and fluxes at the water-sediment and shelf-slope interfaces. Uncertainties on the fluxes and budgets are estimated to assess the robustness of the results with respect to the spatial variability of the system. Results indicate that shelf-slope exchanges by mixing predominate with respect to the major input terms (river discharge and sediment release). Budgets for inorganic nutrients, that show a strong concentration gradient between the shelf and the slope waters, are significantly different from zero and indicate an excess of these elements in the shelf water. For all surveys, these surpluses suggest (i) that the whole shelf system is autotrophic (1.6–4.3 × 10 3 mol C s –1) and acts as a sink of CO 2, and (ii) that it is a site of net denitrification (119 mol N s –1) and acts as a sink of N 2. Average shelf-slope fluxes of dissolved and particulate organic elements generally indicate an export to the open sea. However, the large uncertainty on these fluxes yields budgets for the shelf not significantly different from zero.

Greater GNN pattern bias in sequence elements encoding conserved residues of ancient proteins may be an indicator of amino acid composition of early proteins

The possibility that RNY pattern bias in extant sequences is a remnant of more pronounced bias of this type in early ancestors was investigated. To this end, conserved residues (those residues for which the inferred ancestral and known descendant amino acids are identical) and non-conserved residues of ancient proteins dating to the Last Universal Ancestor were identified within six species: two archaea, two eubacteria and two eukaryotes. Bias within sequence elements encoding each subset of residues, conserved and non-conserved, was then determined. In all species, GNN bias is greater within conserved than non-conserved sequence elements, whereas ANN is not. This difference is statistically significant in all six species examined. Since the relative mutability of the GNN-encoded amino acids does not explain the greater bias in conserved sequences, it is concluded that early sequences probably possessed a strong GNN bias. It is suggested that this bias may be a consequence of the GNN codons being the first introduced into the genetic code. Although NNY bias is also greater within conserved sequence elements of the six species, that difference is statistically significant in only half of them. Therefore, the evidence for early NNY bias remains inconclusive. The findings of this study do not support the proposal of Diaz-Lazcoz et al. (J. Mol. Biol. 250 (1995) 123) that the codons of the TCN four-codon block were the first assigned to serine during the evolution of the genetic code.

Biogeochemical cycles of Org-C, Tot-N and Tot-S in the sediment of the Ghar El Melh Lagoon (north of Tunisia)

Pore water analysis of chloride (Cl −), alkalinity, ammonium (NH 4 +), and sulfate (SO 4 2−) in the Lagoon of Ghar El Melh and the evolution of their relative concentrations indicate that these concentrations had changed over time reflecting the non-steady state conditions prevailing in this lagoon. Cl −, which is considered in this study as a non-conservative tracer element, was used to determine the effective diffusion rates, including bioirrigation or wind action in lagoon sediment, indicating that the effective diffusion coefficient ( D eff) varied between 10 and 20 dm 2 year −1 in summer and was equal to 50 dm 2 year −1 in autumn. The so-calculated diffusion rates clearly expressed the particular role of bioirrigation and wind action as major factors which control the biogeochemical processes in this lagoon. The examination of the 210Pb profile along the sediment column indicated that bioturbation was low in this lagoon, and did play only a very limited role. On the other hand, based on the excess of 210Pb, the sedimentation rate ( w) had been estimated to be less than 0.1 cm year −1 in this lagoon. The alkalinity and NH 4 + modeling combined with calculations for Cl − diffusion have led to the estimation of the following production rates ( R p) at the surface of the lagoon sediment: alkalinity=52–60 mmol l −1 year −1 and NH 4 +=8 mmol l −1 year −1. In spring, D eff and R p were fitted simultaneously showing the following results: D eff alk=10–50 dm 2 year −1; R p alk=57 mmol l −1 year −1; D eff NH 4 + =150 dm 2 year −1 and R p NH 4 + =8 mmol l −1 year −1. Such particularly high values of D eff reflect one more time the crucial impact of bioirrigation or wind action on the biogeochemical processes developing within the pore-water sediment. This result is however supported by the increase of micro-organisms in the sediment and by the typical windy periods occurring in spring. Fluxes calculated using the concentration gradients in pore water within the undisturbed sediment, ranged from 4.48 to 11.13 mmol m −2 day −1 for alkalinity, 0.011–0.351 mmol m −2 day −1 for NH 4 + and 4.64–11.92 mmol m −2 day −1 for T–CO 2. Nevertheless, the calculated NO 3 − fluxes were very low. Seasonal fluxes of SO 4 2−, estimated by the alkalinity budget, are found to range from 1.18 and 3.43 mmol m −2 day −1 in the undisturbed sediment. Combination of fluxes determined using pore water concentration gradients and solid phases have led to a better understanding of the biogeochemical cycles of organic carbon (Org-C), total nitrogen (Tot-N) and total sulfur (Tot-S) developing in the undisturbed sediment of the lagoon. The results showed that Org-C regeneration remained important in the deep sediment, while most of the Tot-N regeneration occurred mainly near the sediment–water interface. As regards the Tot-S, this element appeared to be mainly governed either by the SO 4 2− reduction within the undisturbed sediment and/or by the FeS oxidation/reduction processes occurring at the upper part of the sediment.

Isolation and characterization of the human UGT2B15 gene, localized within a cluster of UGT2B genes and pseudogenes on chromosome 4

Glucuronidation is a major pathway of androgen metabolism and is catalyzed by UDP-glucuronosyltransferase (UGT) enzymes. UGT2B15 and UGT2B17 are 95 % identical in primary structure, and are expressed in steroid target tissues where they conjugate C19 steroids. Despite the similarities, their regulation of expression are different; however, the promoter region and genomic structure of only the UGT2B17 gene have been characterizedX to date. To isolate the UGT2B15 gene and other novel steroid-conjugating UGT2B genes, eight P-1-derived artificial chromosomes (PAC) clones varying in length from 30 kb to 165 kb were isolated. The entire UGT2B15 gene was isolated and characterized from the PAC clone 21598 of 165 kb. The UGT2B15 and UGT2B17 genes are highly conserved, are both composed of six exons spanning approximately 25 kb, have identical exon sizes and have identical exon-intron boundaries. The homology between the two genes extend into the 5′-flanking region, and contain several conserved putative cis-acting elements including Pbx-1, C/EBP, AP-1, Oct-1 and NF/κB. However, transfection studies revealed differences in basal promoter activity between the two genes, which correspond to regions containing non-conserved potential elements. The high degree of homology in the 5′-flanking region between the two genes is lost upstream of −1662 in UGT2B15, and suggests a site of genetic recombination involved in duplication of UGT2B genes. Fluorescence in situ hybridization mapped the UGT2B15 gene to chromosome 4q13.3-21.1. The other PAC clones isolated contain exons from the UGT2B4, UGT2B11 and UGT2B17 genes. Five novel exons, which are highly homologous to the exon 1 of known UGT2B genes, were also identified; however, these exons contain premature stop codons and represent the first recognized pseudogenes of the UGT2B family. The localization of highly homologous UGT2B genes and pseudogenes as a cluster on chromosome 4q13 reveals the complex nature of this gene locus, and other novel homologous UGT2B genes encoding steroid conjugating enzymes are likely to be found in this region of the genome.

Plutonium Activity Ratios in Plankton: New Evidence of Hold-Up Time in Irish Sea Sediments

The determination of activity ratios for radioisotopes of different half-lives can be used to estimate transit times from a point source to locations further away. For conservative elements, this time is approximately equivalent to the net hydrological transport. However, for non-conservative elements such as plutonium, the additional influence of biogeochemical processes decreases the net transport time. In this study, 241Pu and 239,240Pu concentrations in Irish Sea plankton samples, collected in May 1994, were determined and the 241Pu/239,240Pu ratios calculated. Plutonium-239,240 was measured using a standard method by ion exchange chromatography and alpha counting, and 241Pu was determined by liquid scintillation counting using the disk-supported technique. The latter showed some methodological problems, which are briefly discussed. The 241Pu/239,240Pu ratios gave an estimate of the "transit time" from Sellafield to the different sampling points. In fact, this time represents the age of plutonium in plankton, i.e., the time lag between release from Sellafield and detection at the different sampling stations. The mean plutonium age was 17±2 years (n = 10) and 18.6±0.8 years (n = 13) in phytoplankton and zooplankton, respectively. The spatial distribution was reasonably homogeneous over the Irish Sea. The assimilation-elimination processes of plutonium in plankton are rather rapid. Therefore, it may be assumed that, in this time scale, the plutonium concentrations were in equilibrium with surrounding waters. Thus, it is concluded that plutonium was rather old because resuspension-sedimentation processes had occurred that delayed its transport within the Irish Sea. Therefore, the age of plutonium in plankton represented the hold-up time of plutonium in the sediments from the Irish Sea.

High Precision Pb Isotopes in Fe-Mn crusts from the NE Atlantic: A Proxy for Palaeosources and Palaeocean Circulation

Mixing in the oceans is traced using conservative properties (salinity, temperature....) of water masses where mixing is reflected as linear arrays (Broecker and Peng, 1982). The potential of Pb isotopes as tracers of ocean circulation was first demonstrated by the consistency of Pb isotopic variations in CircumAntarctic Fe-Mn deposits with the present-day ocean bottom water circulation in this region (Abouchami and Goldstein, 1995). Although Pb itself is a nonconservative element, binary mixing between water masses or sources will still result in straight linear arrays when plotted in Pb isotope space. However, resolution of such mixing lines depends on the accuracy with which Pb isotope ratios can be measured. In recent studies, palaeoseawater records of Pb and Nd isotopes were obtained using dated depth-profiles from hydrogenous Fe-Mn crusts, and have shown that long-term Pb and Nd isotopic changes in the Pacific and Atlantic appear to be related to deep water circulation changes (Abouchami et al., 1997; Burton et al., 1997; Christensen et al., 1997). So far, because of limitations in analytical precision, only variations in Z~176 ratios have been resolvable. We report high precision Pb isotopic data (2~ext. ~< 100 ppm, a factor of 10 better than conventional analyses) obtained using a Pb triple spike (Galer and Abouchami, This volume), together with Nd isotopic composition on depth profiles from two Fe-Mn crusts from the eastern Atlantic basin. Crust 121DK was dredged from Tropic Seamount which lies off West Africa (24~ 21~ 2000 m), while crust 65GTV comes from the Lion Seamount, west of the Straits of Gibraltar (35~ 15~ 1500 m). These crusts grew in the cores of NADW and MOW at rates of 3 miniMa and 4.5 mm/Ma, respectively, as infen'ed from 1~ data (Koschinsky et al., 1996). The profiles provide a 12 Ma record of changes in eastern North Atlantic Deep Water (ENADW), and over the past 8 Ma for Mediterranean Outflow Water (MOW). Max-Planck Institut far Chemic, Postfach 3060, 55020 Mainz, Germany

New loop-loop tertiary interactions in self-splicing introns of subgroup IC and ID: a complete 3D model of the Tetrahymena thermophila ribozyme.

Group I introns self-splice via two consecutive trans-esterification reactions in the presence of guanosine cofactor and magnesium ions. Comparative sequence analysis has established that a catalytic core of about 120 nucleotides is conserved in all known group I introns. This core is generally not sufficient for activity, however, and most self-splicing group I introns require non-conserved peripheral elements to stabilize the complete three-dimensional (3D) structure. The physico-chemical properties of group I introns make them excellent systems for unraveling the structural basis of the RNA-RNA interactions responsible for promoting the self-assembly of complex RNAs. We present phylogenetic and experimental evidence for the existence of three additional tertiary base pairings between hairpin loops within peripheral components of subgroup IC1 and ID introns. Each of these new long range interactions, called P13, P14 and P16, involves a terminal loop located in domain 2. Although domains 2 of IC and ID introns share very strong sequence similarity, their terminal loops interact with domains 5 and 9 (subgroup IC1) and domain 6 (subgroup ID). Based on these tertiary contacts, comparative sequence analysis, and published experimental results such as Fe(II)-EDTA protection patterns, we propose 3D models for two entire group I introns, the subgroup IC1 intron in the large ribosomal precursor RNA of Tetrahymena thermophila and the SdCob.1 subgroup ID intron found in the cytochrome b gene of Saccharomyces douglasii. Three-dimensional models of group I introns belonging to four different subgroups are now available. They all emphasize the modular and hierarchical organization of the architecture of group I introns and the widespread use of base-pairings between terminal hairpin loops for stabilizing the folded and active structures of large and complex RNA molecules.

Transfer of European coastal pollution to the arctic: Radioactive tracers

Controlled discharges of man-made radionuclides from the nuclear reprocessing facilities at Sellafield (UK) and La Hague (France) have been utilized to trace the transport of European coastal contaminants to the Barents Sea, the Kara Sea, the Arctic Ocean and the East Greenland Current. The transfer is quantified by a transfer factor (TF), calculated as the quotient between observed concentrations in the environment and an average discharge rate t years earlier, where t is the transport time. A TF value of 1–2 ng m −3/t yr −1 has been found for Sellafield discharges in East Greenland Current Polar Water—a water mass reflecting contaminant levels in Arctic Ocean surface water. Such transfer factors are also valid for other conservatively-behaving pollutants discharged to the European coastal zone. Even non-conservative particle-reactive elements (e.g. plutonium) have now been traced from European coastal waters to the Arctic.

Geochemical Evidence for Magma Generation above Subduction Zones

Evaluation of geochemical data from the world's arc-basin systems indicates that the mantle wedge contains two types of element: conservative elements, which owe their concentration predominantly to the nature of the mantle in the wedge; and non-conservative elements, which are added to the mantle during subduction. The most conservative elements are usually high field strength trace elements such as Nb, Ta, HI', Zr, Ti, Y, HREE, V, Sc, Ga. Most major elements are also effectively conservative because they are buffered by their high concentrations in the mantle wedge. Nonconservative elements comprise elements of low to moderate ionic potential such as the alkali and alkaline earth elements, monoand divalent transition metals, some lanthanides (L-MREE) and actinides (Th and U), together with elements of very high ionic potential such as P and S. Conservative and non-conservative elements can be distinguished using plots such as M/Yb v Nb/Yb, where M is the element in question and Nb is treated as the most conservative element. If M is conservative, the data form a trend within the MORB array; if M is non-conservative, the data plot above the MORB array. These plots emphasise that the high field strength elements are mobile only when hot crust in snbducted, in which case partitioning into the residues from slab melting may be more important than ionic potential in determining the composition of the slab component. Investigation of the conservative elements shows that their behaviour largely depends on incompatibility during melting. Thus, Nb, Ta, Hf and Zr (D< 0.05) can be termed very highly i n c o m p a t i b l e ( V H I ) , T i , Y, H R E E (D=0.05-0.15) can be termed highly incompatible (HI), and V, Ga, Sc (D-0.15-0.5) can be termed moderately incompatible (MI). When normalized to fertile MORB mantle (FMM) and plotted in order of incompatibility, the conservative elements form patterns that vary systematically according to the degree of enrichment or depletion of the mantle and the degree of melting (Pearce and Parkinson, 1993).

Gross primary production and nutrient behaviour in a shallow coastal environment

The estimation of the gross primary production (GPP) through measurements of a conserved element, such us phosphorus (P) in macroalgal biomass, surface sediment and settled particulate material (SPM) is proposed. The central Venice lagoon is covered by a biomass ranging from 1 to 20 kg m −2, wet wt, with macroalgae accounting for almost the whole production. The total phosphorus GPP (P-GPP) was more than sixfold the P involved in the net primary production (P-NPP). The GPP also provided the fluxes of nutrients at the water-sediment interface and allowed the estimation of losses of non-conserved elements, such as nitrogen (N) and carbon (C), by denitrification and respiration-decomposition processes, respectively. In the Venice lagoon the N and C losses were evaluated to be 51% and 63%, respectively, of the nutrients involved in the total GPP. From a comparison of these results with the total nutrient loadings entering the lagoon through external inputs, it was suggested that the sea strongly supported the GPP occurring in the lagoon. The release of N from the lagoon to the atmosphere was equivalent to about 4.3 times the N introduced yearly from external inputs.

RNA recognition: towards identifying determinants of specificity

Members of a family of proteins containing a conserved ∼80-amino acid RNA recognition motif (RRM) bind specifically to a wide variety of RNA molecules. Structural studies, in combination with sequence alignments, indicate the structural context of both conserved and non-conserved elements in the motif. These analyses suggest that all RRM proteins share a common fold and a similar protein-RNA interface, and that non-conserved residues contribute additional contacts for sequence-specific RNA recognition.

Force measurement using an ac atomic force microscope

Complete amplitude-displacement-frequency spectra for a lever of an atomic force microscope have been measured and used to determine surface forces in air. Two ac techniques were used to measure the shift in resonant frequency of the Ni lever, as a function of separation from a mica surface. A strong, short-range force was observed for freshly prepared surfaces, implying a van der Waals interaction. A weak, long-range force was observed after exposure of the surfaces for some hours, suggestive of a capillary interaction. The long-range force appeared to contain a nonconservative element which increased on approach to the surface. The results have implications for surface-force determinations by single tip displacement scans, and for imaging with force microscopes.

Groundwater's dynamic role in regulating acidity and chemistry in a precipitation-dominated lake

The role of groundwater in regulating the hydrologic and chemical regime ofan “acid-susceptible” lake situated in a sandy silicate aquifer in northern Wisconsin was determined by direct groundwater measurements using a network of 68 piezometers. Groundwater inputs were compared with precipitation and dryfall components of the hydrologic and chemical budgets for the lake. Groundwater contributed only 10% of the water to Crystal Lake in 1982 but was the major source of alkalinity, calcium, magnesium, sodium, potassium, silicon, and chloride. Precipitation contributed 90% of the water and was the major source of hydrogen and sulfate ions. Continuing analyses suggest that groundwater consistently supplies the bulk of the inoorganic constituents to the lake on annual basis. The flux of groundwater and dissolved solids to the lake is highly seasonal, with large pulses of groundwater inflow occuring after spring snowmelt and in the fall. For certain nonconservative elements such as silicon, the periodic influx of chemically concentrated groundwater can trigger significant changes in the chemical and biological balance in the lake. Despite the fact that the aquifer contains no carbonate minerals and the area experiences acid precipitation (pH = 4.6), the alkalinity of the small amount of groundwater entering Crystal Lake is more than enough to neutralize all of the acidity contributed by precipitation. In-lake sulfate reduction may generate an equal amount of alkalinity. This buffering of pH helps to regulate aluminum concentrations at low levels, thus minimizing potential aluminum toxicity in the lake. Most lakes in the region probably receive even larger amounts of alkalinity-rich groundwater than Crystal Lake, because most lie further downgradient in the regional flow system. This may help to explain why most lakes in this “acid-susceptible” area are no more acidic today than they were 50 years ago.

Oligoribonucleotide map and protein of CMII: detection of conserved and nonconserved genetic elements in avian acute leukemia viruses CMII, MC29, and MH2

RNA and protein of the defective avian acute leukemia virus CMII, which causes myelocytomas in chickens, and of CMII-associated helper virus (CMIIAV) were investigated. The RNA of CMII measured 6 kilobases (kb) and that of CMIIAV measured 8.5 kb. By comparing more than 20 mapped oligonucleotides of CMII RNA with mapped and nonmapped oligonucleotides of acute leukemia viruses MC29 and MH2 and with mapped oligonucleotides of CMIIAV and other nondefective avian tumor viruses, three segments were distinguished in the oligonucleotide map of CMII RNA: (i) a 5' group-specific segment of 1.5 kb which was conserved among CMII, MC29, and MH2 and also homologous with gag-related oligonucleotides of CMIIAV and other helper viruses (hence, group specific); (ii) an internal segment of 2 kb which was conserved specifically among CMII, MC29, and MH2 and whose presence in CMII lends new support to the view that this class of genetic elements is essential for oncogenicity, because it was absent from an otherwise isogenic, nontransforming helper, CMIIAV; and (iii) a 3' group-specific segment of 2.5 kb which shared 13 of 14 oligonucleotides with CMIIAV and included env oligonucleotides of other nondefective viruses of the avian tumor virus group (hence, group specific). This segment and analogous map segments of MC29 and MH2 were not conserved at the level of shared oligonucleotides. CMII-transformed cells contained a nonstructural, gag gene-related protein of 90,000 daltons, distinguished by its size from 110,000-daltom MC29 and 100,000-dalton MH2 counterparts. The gag relatedness and similarity to the 110,000-dalton MC29 counterpart indicated that the 90,000-dalton CMII protein is translated from the 5' and internal segments of CMII RNA. The existence of conserved 5' and internal RNA segments and conserved nonstructural protein products in CMII, MC29, and MH2 indicates that these viruses belong to a related group, termed here the MC29 group. Viruses of the MC29 group differ from one another mainly in their 3' RNA segments and in minor variations of their conserved RNA segments as well as by strain-specific size markers of their gag-related proteins. Because (i) the conserved 5' gag-related and internal RNA segments and their gag-related, nonvirion protein products correlate with the conserved oncogenic spectra of the MC29 group of viruses and because (ii) the internal RNA sequences and nonvirion proteins are not found in nondefective viruses, we propose that the conserved RNA and protein elements are necessary for oncogenicity and probably are the onc gene products of the MC29 group of viruses.

Lagrange’s Equations for Complex Bond Graph Systems

The standard means of imposing causality to extract state equations for bond graph models of physical systems can be inconvenient when algebraic loops and derivative causality in combination with nonlinear constraints are present. This paper presents an alternative means of writing system differential equations using energy and coenergy state functions and Lagrange’s equations. For certain types of systems, particularly mechanical and electromechanical systems, this indirect means of finding state equations turns out to be very convenient. In this technique, causality is used in a new way to establish generalized coordinates and generalized efforts for nonconservative elements. Finally, it is shown that in some cases in which a Lagrangian can be written by inspection for a complex mechanism, a detailed bond graph for this component is unnecessary and yet the equations of the mechanism can be easily coupled to the bond graph equations for the remainder of the system.

A phosphorus residence time model: Theory and application

concentration in the lake is identical to the inputconcentration, and the losses from the lake occur onlythrough the outlet. Neither of these conditions apply tomany lakes. Mean annual concentrations in lakes areoften much lower than mean annual inputconcentrations, and the major loss of phosphorusfrequently results from deposition to the sediments, notfrom discharge through the outlet.For non-conservative elements, the validity of themodel is improved if internal losses as well as outwashlosses are taken into account in the mass balance.Thus, equation (1) must be modified to account for thereactivity of the element in the lake. Studies haveshown that the sediments of lakes act as sinks or trapsfor phosphorus. Although there is some release ofphosphorus from sediments, the net flux of phosphorusover an annual cycle is to the sediments. This followsfrom the fact that a significant portion of the organismsthat settle onto the bottom are refractory so thatregeneration of the phosphorus upon mineralization isless than 100 percent (Jewell,