An alternative method for the calculation of joint probability distributions. Application to the expectation of the triplet invariant.

This paper presents a completely new method for the calculation of expectations (and thus joint probability distributions) of structure factors or phase invariants. As an example, a first approximation of the expectation of the triplet invariant (up to a constant) is given and a complex number is obtained. Instead of considering the atomic vector positions or reciprocal vectors as the fundamental random variables, the method samples over all functions (distributions) with a given number of atoms and given Patterson function. The aim of this paper was to explore the feasibility of the method, so the easiest problem was chosen: the calculation of the expectation value of the triplet invariant in P1. Calculation of the joint probability distribution of the triplet is not performed here but will be done in the future.

Acknowledging conservation trade-offs and embracing complexity.

There is a growing recognition that conservation often entails trade-offs. A focus on trade-offs can open the way to more complete consideration of the variety of positive and negative effects associated with conservation initiatives. In analyzing and working through conservation trade-offs, however, it is important to embrace the complexities inherent in the social context of conservation. In particular, it is important to recognize that the consequences of conservation activities are experienced, perceived, and understood differently from different perspectives, and that these perspectives are embedded in social systems and preexisting power relations. We illustrate the role of trade-offs in conservation and the complexities involved in understanding them with recent debates surrounding REDD (Reducing Emissions from Deforestation and Degradation), a global conservation policy designed to create incentives to reduce tropical deforestation. Often portrayed in terms of the multiple benefits it may provide: poverty alleviation, biodiversity conservation, and climate-change mitigation; REDD may involve substantial trade-offs. The gains of REDD may be associated with a reduction in incentives for industrialized countries to decrease carbon emissions; relocation of deforestation to places unaffected by REDD; increased inequality in places where people who make their livelihood from forests have insecure land tenure; loss of biological and cultural diversity that does not directly align with REDD measurement schemes; and erosion of community-based means of protecting forests. We believe it is important to acknowledge the potential trade-offs involved in conservation initiatives such as REDD and to examine these trade-offs in an open and integrative way that includes a variety of tools, methods, and points of view.

Seeing (and doing) conservation through cultural lenses.

In this paper, we first discuss various vantage points gained through the authors' experience of approaching conservation through a "cultural lens." We then draw out more general concerns that many anthropologists hold with respect to conservation, summarizing and commenting on the work of the Conservation and Community Working Group within the Anthropology and Environment Section of the American Anthropological Association. Here we focus on both critiques and contributions the discipline of anthropology makes with regard to conservation, and show how anthropologists are moving beyond conservation critiques to engage actively with conservation practice and policy. We conclude with reflections on the possibilities for enhancing transdisciplinary dialogue and practice through reflexive questioning, the adoption of disciplinary humility, and the realization that "cross-border" collaboration among conservation scholars and practitioners can strengthen the political will necessary to stem the growing commoditization and ensuing degradation of the earth's ecosystems.

The triplet invariant revisited.

It is shown that the formula for the positivity of the triplet invariant in P 1bar changes drastically if one uses a different statistical method by imposing acceptable and unbiased additional structural information. We obtain a much lower probability for the strength (almost (1/2)) of the triplet formula than the classical one.

A statistical interpretation of the triplet and quartet invariant in P1. A theoretical discussion.

We present a method that we call symbolic asymptotic development (SAD) to obtain joint probability distributions (j.p.d.'s) of phases of structure factors for general even densities of the atomic position vectors. The formula for the triplet and quartet invariant that we obtain in this way reduces to the well known classical formula for the case of a uniform density of the atomic position vectors. For the case of complete knowledge of the atomic vectors it reduces to first order to the exact probability density of the triplet (quartet) phase invariant. Applying this formula to the most general j.p.d. of the atomic vectors we obtain a statistical interpretation of Hauptman's algebraic B(3,0) and B(4,0) formulas. We also give a heuristic derivation of the SAD method. Another method that we shall discuss uses a method called linearization of the invariants that also produces formulas for the triplet phase invariant. This method is based on previous work and is also more laborious to calculate with than the SAD method. It can also give a statistical interpretation of the B(3,0) formula. We show that the formula obtained for the triplet resembles the formula obtained with SAD.

The quartet revisited.

Using an unbiased and very general joint density of the atomic position vectors we are able to calculate different probabilities for the sign of the quartet given its second neighborhood. One already knows that additional chemical information alters the joint probability distribution (j.p.d.) of structure factors. That is, they can and will give different j.p.d.'s for the quartet invariant given its second neighborhood. In this paper we show that even without additional chemical information the j.p.d.'s of structure factors can be strongly different from the classical ones if we impose a general j.p.d. for the atomic vectors based on the fact that the real distribution of the atomic position vectors is a sum of delta functions.

Modern genomes with retro-look: retrotransposed elements, retroposition and the origin of new genes.

A fascinating evolutionary facet of retroposition is its ability to generate a dynamic reservoir of sequences for the formation of new genes within genomes. Retroelement genes, such as gag from retrotransposons or envelope genes from endogenous retroviruses, have been repeatedly exapted and domesticated during evolution. Such genes fulfill now useful novel functions in diverse aspects of host biology, for example placenta formation in mammals. New protein-coding genes can also be generated through the reverse transcription of mRNA from 'classical' genes by the enzymatic machinery of autonomous retroelements. Many of these retrogenes, which generally show a modified expression pattern compared to their molecular progenitor, have a testis-biased expression and a potential role in spermatogenesis in different animals. New non-protein-coding RNA genes have also been repeatedly generated through retroposition during evolution. A striking evolutionary parallel has been observed between two such RNA genes, the rodent BC1 and the primate BC200 genes. Although both genes are derived from different types of sequences (tRNA and Alu short interspersed element, respectively), they are both expressed almost specifically in neurons, transported into the dendrites and included in ribonucleoprotein complexes containing the poly(A)-binding protein PABP. Both BC1 and BC200 RNA are able to inhibit translation in vitro and are progenitors of new families of short interspersed elements. These genes, which might play a role in animal behavior, provide an astonishing example of evolutionary convergence in two distinct mammalian lineages, which is also observed for placenta genes derived from endogenous retroviruses. Finally, there are indications that genes for small nucleolar RNAs (snoRNAs) and possibly microRNAs (miRNAs) can also be duplicated via retroposition. Taken together, these observations definitely demonstrate the major role of retroposition as mediator of genomic plasticity and contributor to gene novelties. Therefore, the 'retro-look' of genomes is in fact indicative of their modernity.

Echoes from the past--are we still in an RNP world?

Availability of the human genome sequence and those of other species is unmeasured in their value for a comprehensive understanding of the architecture, function and evolution of genomes and cells. Various mechanisms keep genomes in flux and generate intra- and interspecies variation. The conversion of RNA modules into DNA and their more or less random integration into chromosomes (retroposition) is in many lineages including our own the most pervasive and perhaps the most enigmatic. The proclivity of such events in extant multicellular eukaryotes, even in more recent evolutionary times, gives the impression that the transition period from the RNP (ribonucleoprotein) world to the emergence of modern cells, where DNA became the predominant carrier of genetic information, has lasted billions of years and is an endlessly drawn-out process rather than the punctuated event one might expect. Apart from the impact of such RNA-mediated processes as retroposition, the role of RNA in a wide variety of cellular functions has only recently become more widely appreciated.

An unusual primate locus that attracted two independent Alu insertions and facilitates their transcription.

BC200 RNA, a neuronal, small non-messenger RNA that originated from a monomeric Alu element is specific to anthropoid primates. Tarsiers lack an insert at the orthologous genomic position, whereas strepsirrhines (Lemuriformes and Lorisiformes) acquired a dimeric Alu element, independently from anthropoids. In Galago moholi, the CpG dinucleotides are conspicuously conserved, while in Eulemur coronatus a large proportion is changed, indicating that the G.moholi Alu is under purifying selection and might be transcribed. Indeed, Northern blot analysis of total brain RNA from G.moholi with a specific probe revealed a prominent signal. In contrast, a corresponding signal was absent from brain RNA from E.coronatus. Isolation and sequence analysis of additional strepsirrhine loci confirmed the differential sequence conservation including CpG patterns of the orthologous dimeric Alu elements in Lorisiformes and Lemuriformes. Interestingly, all examined Alu elements from Lorisiformes were transcribed, while all from Lemuriformes were silent when transiently transfected into HeLa cells. Upstream sequences, especially those between the transcriptional start site and -22 upstream, were important for basal transcriptional activity. Thus, the BC200 RNA gene locus attracted two independent Alu insertions during its evolutionary history and provided upstream promoter elements required for their transcription.

Role of a neuronal small non-messenger RNA: behavioural alterations in BC1 RNA-deleted mice.

BC1 RNA is a small non-messenger RNA common in dendritic microdomains of neurons in rodents. In order to investigate its possible role in learning and behaviour, we compared controls and knockout mice from three independent founder lines established from separate embryonic stem cells. Mutant mice were healthy with normal brain morphology and appeared to have no neurological deficits. A series of tests for exploration and spatial memory was carried out in three different laboratories. The tests were chosen as to ensure that different aspects of spatial memory and exploration could be separated and that possible effects of confounding variables could be minimised. Exploration was studied in a barrier test, in an open-field test, and in an elevated plus-maze test. Spatial memory was investigated in a Barnes maze and in a Morris water maze (memory for a single location), in a multiple T-maze and in a complex alley maze (route learning), and in a radial maze (working memory). In addition to these laboratory tasks, exploratory behaviour and spatial memory were assessed under semi-naturalistic conditions in a large outdoor pen. The combined results indicate that BC1 RNA-deficient animals show behavioural changes best interpreted in terms of reduced exploration and increased anxiety. In contrast, spatial memory was not affected. In the outdoor pen, the survival rates of BC1-depleted mice were lower than in controls. Thus, we conclude that the neuron-specific non-messenger BC1 RNA contributes to the aptive modulation of behaviour.

tRNAs in the spotlight during protein biosynthesis.

High resolution crystal structures of the ribosome provide fascinating insights into perhaps the most sophisticated machine of a cell. Yet, this translational apparatus must have developed from a much more primitive structure. Throughout the evolution of this apparatus, tRNAs have been, and still are, key players in the translation process.

[Study of the binding of the S7 protein with 16S rRNA fragment 926-986/1219-1393 as a key step in the assembly of the small subunit of prokaryotic ribosomes]. / Izuchenie sviazyvanie belka S7 s fragmentom 926-986/1219-1393 16S rRNK--kliuchevogo étapa sborki maloi subchastitsy prokarioticheskikh ribosom.

Both structural and thermodynamic studies are necessary to understand the ribosome assembly. An initial step was made in studying the interaction between a 16S rRNA fragment and S7, a key protein in assembling the prokaryotic ribosome small subunit. The apparent dissociation constant was obtained for complexes of recombinant Escherichia coli and Thermus thermophilus S7 with a fragment of the 3' domain of the E. coli 16S rRNA. Both proteins showed a high rRNA-binding activity, which was not observed earlier. Since RNA and proteins are conformationally labile, their folding must be considered to correctly describe the RNA-protein interactions.

RNomics: an experimental approach that identifies 201 candidates for novel, small, non-messenger RNAs in mouse.

In mouse brain cDNA libraries generated from small RNA molecules we have identified a total of 201 different expressed RNA sequences potentially encoding novel small non-messenger RNA species (snmRNAs). Based on sequence and structural motifs, 113 of these RNAs can be assigned to the C/D box or H/ACA box subclass of small nucleolar RNAs (snoRNAs), known as guide RNAs for rRNA. While 30 RNAs represent mouse homologues of previously identified human C/D or H/ACA snoRNAs, 83 correspond to entirely novel snoRNAS: Among these, for the first time, we identified four C/D box snoRNAs and four H/ACA box snoRNAs predicted to direct modifications within U2, U4 or U6 small nuclear RNAs (snRNAs). Furthermore, 25 snoRNAs from either class lacked antisense elements for rRNAs or snRNAS: Therefore, additional snoRNA targets have to be considered. Surprisingly, six C/D box snoRNAs and one H/ACA box snoRNA were expressed exclusively in brain. Of the 88 RNAs not belonging to either snoRNA subclass, at least 26 are probably derived from truncated heterogeneous nuclear RNAs (hnRNAs) or mRNAS: Short interspersed repetitive elements (SINEs) are located on five RNA sequences and may represent rare examples of transcribed SINES: The remaining RNA species could not as yet be assigned either to any snmRNA class or to a part of a larger hnRNA/mRNA. It is likely that at least some of the latter will represent novel, unclassified snmRNAS:

Birth of a gene: locus of neuronal BC200 snmRNA in three prosimians and human BC200 pseudogenes as archives of change in the Anthropoidea lineage.

The gene encoding brain-specific dendritic BC200 small non-messenger RNA is limited to the primate order and arose from a monomeric Alu element. It is present and neuronally expressed in all Anthropoidea examined. By comparing the human sequence of about 13.2 kb with each of the prosimian (lemur 14.6 kb, galago 12 kb, and tarsier 13.8 kb) orthologous loci, we could establish that the BC200 RNA gene is absent from the prosimian lineages. In Strepsirhini (lemurs and lorises), a dimeric AluJ-like element integrated very close to the BC200 insertion point, while the corresponding tarsier region is devoid of any repetitive element. Consequently, insertion of the Alu monomer that gave rise to the BC200 RNA gene must have occurred after the anthropoid lineage diverged from the prosimian lineage(s). Shared insertions of other repetitive elements favor proximity of simians and tarsiers in support of their grouping into Haplorhini and the omomyid hypothesis. On the other hand, the nucleotide sequences in the segment that is available for comparison in all four species reveal less exchanges between Strepsirhini (lemur and galago) and human than between tarsier and human. Our data imply that the early activity of dimeric Alu sequences must have been concurrent with the activity of monomeric Alu elements that persisted longer than is usually thought. As BC200 RNA gave rise to more than 200 pseudogenes, we used their consensus sequence variations as a molecular archive recording the BC200 RNA sequence changes in the anthropoid lineage leading to Homo sapiens and timed these alterations over the past 35-55 million years.

Neuronal BC1 RNA structure: evolutionary conversion of a tRNA(Ala) domain into an extended stem-loop structure.

By chemical and enzymatic probing, we have analyzed the secondary structure of rodent BC1 RNA, a small brain-specific non-messenger RNA. BC1 RNA is specifically transported into dendrites of neuronal cells, where it is proposed to play a role in regulation of translation near synapses. In this study we demonstrate that the 5' domain of BC1 RNA, derived from tRNA(Ala), does not fold into the predicted canonical tRNA cloverleaf structure. We present evidence that by changing bases within the tRNA(Ala) domain during the course of evolution, an extended stem-loop structure has been created in BC1 RNA. The new structural domain might function, in part, as a putative binding site for protein(s) involved in dendritic transport of BC1 RNA within neurons. Furthermore, BC1 RNA contains, in addition to the extended stem-loop structure, an internal poly(A)-rich region that is supposedly single stranded, followed by a second smaller stem-loop structure at the 3' end of the RNA. The three distinct structural domains reflect evolutionary legacies of BC1 RNA.

Neuronal BC1 RNA: co-expression with growth-associated protein-43 messenger RNA.

Brain-specific cytoplasmic RNA 1 (BC1-RNA), a non-coding RNA polymerase III transcript, is a neuronal RNA that is specifically targeted to dendritic domains. It is co-localized with components of the dendritic protein synthetic machinery, and it has been suggested to operate in the regulation of local translation-related processes in postsynaptic microdomains, thus subserving long-term synaptic plasticity in neurons. To probe the relevance of BC1 expression in neuronal plasticity, we have analyzed the expression pattern of BC1 RNA in the rat nervous system. We found that BC1 RNA is expressed by a specific subset of neurons (but not by non-neuronal cells) in the central and peripheral nervous system of the adult rat. The BC1 labeling pattern indicates that the subcellular location of the RNA is typically postsynaptic which, depending on cell type, manifests itself in a predominantly somatic, somatodendritic, or dendritic location. Our results further show that BC1-expressing neurons typically co-express the messenger RNA for growth-associated protein-43 (GAP-43). Such co-expression was observed in diverse brain areas, including the olfactory bulb, neocortex, and hippocampus, among others. While BC1 RNA was in many neuronal cell types detectable in distal dendritic domains, GAP-43 messenger RNA was typically more restricted to neuronal perikarya. In the mature nervous system, expression of GAP-43 has been described as an intrinsic determinant of predominantly presynaptic plasticity, while BC1 RNA has been implicated in postsynaptic plasticity. Co-expression of both RNAs, as reported here, thus identifies a distinct subset of neurons in the rat nervous system that exhibits both types of plasticity.

A key role in catalysis for His89 of adenylosuccinate lyase of Bacillus subtilis.

Adenylosuccinate lyase of Bacillus subtilis is a tetrameric enzyme which catalyzes the cleavage of adenylosuccinate to AMP and fumarate. We have mutated His(89), one of three conserved histidines, to Gln, Ala, Glu, and Arg. The enzymes were expressed in Escherichia coli and purified to homogeneity. As compared to a specific activity of 1. 56 micromol of adenylosuccinate converted/min/mg protein for wild-type enzyme, the mutant enzymes exhibit specific activities of 0.0225, 0.0036, 0.0036, and 0.0009 for H89Q, H89A, H89E, and H89R, respectively. Circular dichroism and FPLC gel filtration reveal that mutant enzymes have a similar conformation and oligomeric state to that of wild-type enzyme. In H89Q, the K(M) for adenylosuccinate increases slightly to 2.5-fold that of wild-type, the K(M) for fumarate is elevated 3.3-fold, and the K(M) for AMP is 13 times higher than that observed in wild-type enzyme. The catalytic efficiency of the H89Q enzyme is compromised, with k(cat)/K(M) reduced 174-fold in the direction of AMP formation. These data suggest that His(89) plays a role in both the binding of the AMP portion of the substrate and in correctly orienting the substrate for catalysis. Incubation of H89Q with inactive H141Q enzyme [Lee, T. T., Worby, C., Bao, Z.-Q., Dixon, J. E., and Colman, R. F. (1999) Biochemistry 38, 22-32] leads to a 30-fold increase in activity. This intersubunit complementation indicates that His(89) and His(141) from different subunits participate in the active site and that both are required for catalysis.

A tRNA pseudogene in the archaeon Methanococcus jannaschii.

While searching the first completely sequenced genome of the archaeon Methanococcus jannaschii for a small RNA gene, we discovered a 5' truncated gene of a transfer RNA (tRNA(Ser-UCR)) at position 334,431-334,486; including the CCA-end that exactly matched the 3' terminal domain of the annotated M. jannaschii tRNA(Ser-UCR) gene located at position 303,992-304,081. This truncated tRNA gene covering 56 nucleotides (about 2/3) of the genuine tRNA represents, to the best of our knowledge, the first described tRNA pseudogene in the archaeal domain.