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NORMAL GENOMIC

• Willkomm S, Restle T
• Conformational Dynamics of Ago-Mediated Silencing Processes.
• Int J Mol Sci. 2015; 16: 14769-85
• Display abstract

• Argonaute (Ago) proteins are key players of nucleic acid-based interference mechanisms. Their domains and structural organization are widely conserved in all three domains of life. However, different Ago proteins display various substrate preferences. While some Ago proteins are able to use several substrates, others are limited to a single one. Thereby, they were demonstrated to act specifically on their preferred substrates. Here, we discuss mechanisms of Ago-mediated silencing in relation to structural and biochemical insights. The combination of biochemical and structural information enables detailed analyses of the complex dynamic interplay between Ago proteins and their substrates. Especially, transient binding data allow precise investigations of structural transitions taking place upon Ago-mediated guide and target binding.

• Diekmann Y, Pereira-Leal JB
• Evolution of intracellular compartmentalization.
• Biochem J. 2013; 449: 319-31
• Display abstract

• Cells compartmentalize their biochemical functions in a variety of ways, notably by creating physical barriers that separate a compartment via membranes or proteins. Eukaryotes have a wide diversity of membrane-based compartments, many that are lineage- or tissue-specific. In recent years, it has become increasingly evident that membrane-based compartmentalization of the cytosolic space is observed in multiple prokaryotic lineages, giving rise to several types of distinct prokaryotic organelles. Endosymbionts, previously believed to be a hallmark of eukaryotes, have been described in several bacteria. Protein-based compartments, frequent in bacteria, are also found in eukaryotes. In the present review, we focus on selected intracellular compartments from each of these three categories, membrane-based, endosymbiotic and protein-based, in both prokaryotes and eukaryotes. We review their diversity and the current theories and controversies regarding the evolutionary origins. Furthermore, we discuss the evolutionary processes acting on the genetic basis of intracellular compartments and how those differ across the domains of life. We conclude that the distinction between eukaryotes and prokaryotes no longer lies in the existence of a compartmentalized cell plan, but rather in its complexity.

• Wilson MZ, Gitai Z
• Beyond the cytoskeleton: mesoscale assemblies and their function in spatial organization.
• Curr Opin Microbiol. 2013; 16: 177-83
• Display abstract

• Recent studies have identified a growing number of mesoscale protein assemblies in both bacterial and eukaryotic cells. Traditionally, these polymeric assemblies are thought to provide structural support for the cell and thus have been classified as the cytoskeleton. However a new class of macromolecular structure is emerging as an organizer of cellular processes that occur on scales hundreds of times larger than a single protein. We propose two types of self-assembling structures, dynamic globules and crystalline scaffolds, and suggest they provide a means to achieve cell-scale order. We discuss general mechanisms for assembly and regulation. Finally, we discuss assemblies that are found to organize metabolism and what possible mechanisms may serve these metabolic enzyme complexes.

• Abe N et al.
• Rolling circle amplification in a prokaryotic translation system using small circular RNA.
• Angew Chem Int Ed Engl. 2013; 52: 7004-8

• Sen GC, Fensterl V
• Crystal structure of IFIT2 (ISG54) predicts functional properties of IFITs.
• Cell Res. 2012; 22: 1407-9
• Display abstract

• Interferon carries out its cellular effects, including its antiviral effects, by inducing the synthesis of many new proteins, amongst which is the IFIT (ISG56) family of proteins. The first crystal structure of an IFIT, reported by Yang et al., revealed several functional properties of the protein that may help us to better understand the biological functions of these proteins.

• Garcia SP, Pinho AJ, Rodrigues JM, Bastos CA, Ferreira PJ
• Minimal absent words in prokaryotic and eukaryotic genomes.
• PLoS One. 2011; 6: 16065-16065
• Display abstract

• Minimal absent words have been computed in genomes of organisms from all domains of life. Here, we explore different sets of minimal absent words in the genomes of 22 organisms (one archaeota, thirteen bacteria and eight eukaryotes). We investigate if the mutational biases that may explain the deficit of the shortest absent words in vertebrates are also pervasive in other absent words, namely in minimal absent words, as well as to other organisms. We find that the compositional biases observed for the shortest absent words in vertebrates are not uniform throughout different sets of minimal absent words. We further investigate the hypothesis of the inheritance of minimal absent words through common ancestry from the similarity in dinucleotide relative abundances of different sets of minimal absent words, and find that this inheritance may be exclusive to vertebrates.

• Tan CS
• Sequence, structure, and network evolution of protein phosphorylation.
• Sci Signal. 2011; 4: 6-6
• Display abstract

• With the increasing amount of information about the phosphoproteomes of diverse organisms, it is now possible to begin to evaluate this information in the context of evolution. Work described at the inaugural Keystone Symposium on "The Evolution of Protein Phosphorylation" covered a wide range of eukaryotic and prokaryotic organisms, revealing insights into the evolution of protein phosphorylation at the sequence, network, and structural levels.

• Suter DM, Molina N, Naef F, Schibler U
• Origins and consequences of transcriptional discontinuity.
• Curr Opin Cell Biol. 2011; 23: 657-62
• Display abstract

• In both prokaryotes and eukaryotes, transcription has been described as being temporally discontinuous, most genes being active mainly during short activity windows interspersed by silent periods. In mammalian cells, recent studies performed at the single cell level have revealed that transcriptional kinetics are highly gene-specific and constrained by the presence of refractory periods of inactivity before a gene can be turned on again. While the underlying mechanisms generating gene-specific kinetic characteristics remain unclear, various biological consequences of transcriptional discontinuity have been unravelled during the past few years. Here we review recent advances on understanding transcriptional kinetics of individual genes at the single cell level and discuss its possible origins and consequences.

• Gnad F, Forner F, Zielinska DF, Birney E, Gunawardena J, Mann M
• Evolutionary constraints of phosphorylation in eukaryotes, prokaryotes, and mitochondria.
• Mol Cell Proteomics. 2010; 9: 2642-53
• Display abstract

• High accuracy mass spectrometry has proven to be a powerful technology for the large scale identification of serine/threonine/tyrosine phosphorylation in the living cell. However, despite many described phosphoproteomes, there has been no comparative study of the extent of phosphorylation and its evolutionary conservation in all domains of life. Here we analyze the results of phosphoproteomics studies performed with the same technology in a diverse set of organisms. For the most ancient organisms, the prokaryotes, only a few hundred proteins have been found to be phosphorylated. Applying the same technology to eukaryotic species resulted in the detection of thousands of phosphorylation events. Evolutionary analysis shows that prokaryotic phosphoproteins are preferentially conserved in all living organisms, whereas-site specific phosphorylation is not. Eukaryotic phosphosites are generally more conserved than their non-phosphorylated counterparts (with similar structural constraints) throughout the eukaryotic domain. Yeast and Caenorhabditis elegans are two exceptions, indicating that the majority of phosphorylation events evolved after the divergence of higher eukaryotes from yeast and reflecting the unusually large number of nematode-specific kinases. Mitochondria present an interesting intermediate link between the prokaryotic and eukaryotic domains. Applying the same technology to this organelle yielded 174 phosphorylation sites mapped to 74 proteins. Thus, the mitochondrial phosphoproteome is similarly sparse as the prokaryotic phosphoproteomes. As expected from the endosymbiotic theory, phosphorylated as well as non-phosphorylated mitochondrial proteins are significantly conserved in prokaryotes. However, mitochondrial phosphorylation sites are not conserved throughout prokaryotes, consistent with the notion that serine/threonine phosphorylation in prokaryotes occurred relatively recently in evolution. Thus, the phosphoproteome reflects major events in the evolution of life.

• Makarova KS, Wolf YI, van der Oost J, Koonin EV
• Prokaryotic homologs of Argonaute proteins are predicted to function as key components of a novel system of defense against mobile genetic elements.
• Biol Direct. 2009; 4: 29-29
• Display abstract

• BACKGROUND: In eukaryotes, RNA interference (RNAi) is a major mechanism of defense against viruses and transposable elements as well of regulating translation of endogenous mRNAs. The RNAi systems recognize the target RNA molecules via small guide RNAs that are completely or partially complementary to a region of the target. Key components of the RNAi systems are proteins of the Argonaute-PIWI family some of which function as slicers, the nucleases that cleave the target RNA that is base-paired to a guide RNA. Numerous prokaryotes possess the CRISPR-associated system (CASS) of defense against phages and plasmids that is, in part, mechanistically analogous but not homologous to eukaryotic RNAi systems. Many prokaryotes also encode homologs of Argonaute-PIWI proteins but their functions remain unknown. RESULTS: We present a detailed analysis of Argonaute-PIWI protein sequences and the genomic neighborhoods of the respective genes in prokaryotes. Whereas eukaryotic Ago/PIWI proteins always contain PAZ (oligonucleotide binding) and PIWI (active or inactivated nuclease) domains, the prokaryotic Argonaute homologs (pAgos) fall into two major groups in which the PAZ domain is either present or absent. The monophyly of each group is supported by a phylogenetic analysis of the conserved PIWI-domains. Almost all pAgos that lack a PAZ domain appear to be inactivated, and the respective genes are associated with a variety of predicted nucleases in putative operons. An additional, uncharacterized domain that is fused to various nucleases appears to be a unique signature of operons encoding the short (lacking PAZ) pAgo form. By contrast, almost all PAZ-domain containing pAgos are predicted to be active nucleases. Some proteins of this group (e.g., that from Aquifex aeolicus) have been experimentally shown to possess nuclease activity, and are not typically associated with genes for other (putative) nucleases. Given these observations, the apparent extensive horizontal transfer of pAgo genes, and their common, statistically significant over-representation in genomic neighborhoods enriched in genes encoding proteins involved in the defense against phages and/or plasmids, we hypothesize that pAgos are key components of a novel class of defense systems. The PAZ-domain containing pAgos are predicted to directly destroy virus or plasmid nucleic acids via their nuclease activity, whereas the apparently inactivated, PAZ-lacking pAgos could be structural subunits of protein complexes that contain, as active moieties, the putative nucleases that we predict to be co-expressed with these pAgos. All these nucleases are predicted to be DNA endonucleases, so it seems most probable that the putative novel phage/plasmid-defense system targets phage DNA rather than mRNAs. Given that in eukaryotic RNAi systems, the PAZ domain binds a guide RNA and positions it on the complementary region of the target, we further speculate that pAgos function on a similar principle (the guide being either DNA or RNA), and that the uncharacterized domain found in putative operons with the short forms of pAgos is a functional substitute for the PAZ domain. CONCLUSION: The hypothesis that pAgos are key components of a novel prokaryotic immune system that employs guide RNA or DNA molecules to degrade nucleic acids of invading mobile elements implies a functional analogy with the prokaryotic CASS and a direct evolutionary connection with eukaryotic RNAi. The predictions of the hypothesis including both the activities of pAgos and those of the associated endonucleases are readily amenable to experimental tests.

• Borgese N, Driessen AJ, Rapaport D, Robinson C
• The quest for a better resolution of protein-translocation processes. Conference on the Control, Co-ordination and Regulation of Protein Targeting and Translocation.
• EMBO Rep. 2009; 10: 337-42

• Su H, Trombly MI, Chen J, Wang X
• Essential and overlapping functions for mammalian Argonautes in microRNA silencing.
• Genes Dev. 2009; 23: 304-17
• Display abstract

• MicroRNA (miRNA) silencing fine-tunes protein output and regulates diverse biological processes. Argonaute (Ago) proteins are the core effectors of the miRNA pathway. In lower organisms, multiple Agos have evolved specialized functions for distinct RNA silencing pathways. However, the roles of mammalian Agos have not been well characterized. Here we show that mouse embryonic stem (ES) cells deficient for Ago1-4 are completely defective in miRNA silencing and undergo apoptosis. In miRNA silencing-defective ES cells, the proapoptotic protein Bim, a miRNA target, is increased, and up-regulation of Bim is sufficient to induce ES cell apoptosis. Expression of activated Akt inhibits Bim expression and partially rescues the growth defect in Ago-deficient ES cells. Furthermore, reintroduction of any single Ago into Ago-deficient cells is able to rescue the endogenous miRNA silencing defect and apoptosis. Consistent with this, each Ago is functionally equivalent with bulged miRNA duplexes for translational repression, whereas Ago1 and Ago2 appear to be more effective at utilizing perfectly matched siRNAs. Thus, our results demonstrate that mammalian Agos all contribute to miRNA silencing, and individual Agos have largely overlapping functions in this process.

• Warmuth S, Zimmermann I, Dutzler R
• X-ray structure of the C-terminal domain of a prokaryotic cation-chloride cotransporter.
• Structure. 2009; 17: 538-46
• Display abstract

• The cation-chloride cotransporters (CCCs) mediate the electroneutral transport of chloride in dependence of sodium and potassium. The proteins share a conserved structural scaffold that consists of a transmembrane transport domain followed by a cytoplasmic regulatory domain. We have determined the X-ray structure of the C-terminal domain of the archaea Methanosarcina acetivorans. The structure shows a novel fold of a regulatory domain that is distantly related to universal stress proteins. The protein forms dimers in solution, which is consistent with the proposed dimeric organization of eukaryotic CCC transporters. The dimer interface observed in different crystal forms is unusual because the buried area is relatively small and hydrophilic. By using a biochemical approach we show that this interaction is preserved in solution and in the context of the full-length transporter. Our studies reveal structural insight into the CCC family and establish the oligomeric organization of this important class of transport proteins.

• Ling SH, Cheng Z, Song H
• Structural aspects of RNA helicases in eukaryotic mRNA decay.
• Biosci Rep. 2009; 29: 339-49
• Display abstract

• mRNA decay is critical for the regulation of gene expression and the quality control of mRNA. RNA helicases play a key role in eukaryotic mRNA decay. In general, RNA helicases utilize the energy of ATP hydrolysis to remodel RNA or RNA-protein complexes, resulting in the separation of RNA duplex strand and/or displacement of proteins from the RNA molecule in RNP (ribonucleoprotein) complexes. Recently, high-resolution crystal structures of RNA helicases in mRNA decay have contributed a great deal to our understanding of these key molecules. In the present review, we focus on the structural and mechanistic aspects of three RNA helicases, Dhh1, Upf1 and eIF4AIII, that are involved in eukaryotic mRNA decay.

• Symmons MF, Luisi BF
• Through ancient rings thread programming strings.
• Structure. 2009; 17: 1429-31

• Siomi MC, Kuramochi-Miyagawa S
• RNA silencing in germlines--exquisite collaboration of Argonaute proteins with small RNAs for germline survival.
• Curr Opin Cell Biol. 2009; 21: 426-34
• Display abstract

• As the proper development of germlines is vital for species preservation, elaborative, regulatory systems for gene expression must operate in germlines. One such system is RNA silencing, sequence-specific gene silencing mechanisms mediated by small RNAs of 20-30 nucleotides long. Indeed, recent studies have revealed that various types of small RNAs are expressed germline-specifically. To preserve the germlines, they collaborate with Argonaute proteins, the catalytic engines in RNA silencing, to inhibit injurious, parasitic genes, transcriptionally or post-transcriptionally. This chapter summarizes the exquisite collaboration of Argonaute proteins with small RNAs in the RNA silencing mechanisms necessary for germline survival in Drosophila and mice.

• Jannot G, Boisvert ME, Banville IH, Simard MJ
• Two molecular features contribute to the Argonaute specificity for the microRNA and RNAi pathways in C. elegans.
• RNA. 2008; 14: 829-35
• Display abstract

• In Caenorhabditis elegans, specific Argonaute proteins are dedicated to the RNAi and microRNA pathways. To uncover how the precise Argonaute selection occurs, we designed dsRNA triggers containing both miRNA and siRNA sequences. While dsRNA carrying nucleotides mismatches can only enter the miRNA pathway, a fully complementary dsRNA successfully rescues let-7 miRNA function and initiates silencing by RNAi. We demonstrated that RDE-1 is essential for RNAi induced by the perfectly paired trigger, yet is not required for silencing by the let-7 miRNA. In contrast, ALG-1/ALG-2 are required for the miRNA function, but not for the siRNA-directed gene silencing. Finally, a dsRNA containing a bulged miRNA and a perfectly paired siRNA can enter both pathways suggesting that the sorting of small RNAs occurs after that the dsRNA trigger has been processed by Dicer. Thus, our data suggest that the selection of Argonaute proteins is affected by two molecular features: (1) the structure of the small RNA duplex; and (2) the Argonautes specific characteristics.

• Desset S, Buchon N, Meignin C, Coiffet M, Vaury C
• In Drosophila melanogaster the COM locus directs the somatic silencing of two retrotransposons through both Piwi-dependent and -independent pathways.
• PLoS One. 2008; 3: 1526-1526
• Display abstract

• BACKGROUND: In the Drosophila germ line, repeat-associated small interfering RNAs (rasiRNAs) ensure genomic stability by silencing endogenous transposable elements. This RNA silencing involves small RNAs of 26-30 nucleotides that are mainly produced from the antisense strand and function through the Piwi protein. Piwi belongs to the subclass of the Argonaute family of RNA interference effector proteins, which are expressed in the germline and in surrounding somatic tissues of the reproductive apparatus. In addition to this germ-line expression, Piwi has also been implicated in diverse functions in somatic cells. PRINCIPAL FINDINGS: Here, we show that two LTR retrotransposons from Drosophila melanogaster, ZAM and Idefix, are silenced by an RNA silencing pathway that has characteristics of the rasiRNA pathway and that specifically recognizes and destroys the sense-strand RNAs of the retrotransposons. This silencing depends on Piwi in the follicle cells surrounding the oocyte. Interestingly, this silencing is active in all the somatic tissues examined from embryos to adult flies. In these somatic cells, while the silencing still involves the strict recognition of sense-strand transcripts, it displays the marked difference of being independent of the Piwi protein. Finally, we present evidence that in all the tissues examined, the repression is controlled by the heterochromatic COM locus. CONCLUSION: Our data shed further light on the silencing mechanism that acts to target Drosophila LTR retrotransposons in somatic cells throughout fly development. They demonstrate that different RNA silencing pathways are involved in ovarian versus other somatic tissues, since Piwi is necessary for silencing in the former tissues but is dispensable in the latter. They further demonstrate that these pathways are controlled by the heterochromatic COM locus which ensures the overall protection of Drosophila against the detrimental effects of random retrotransposon mobilization.

• Rudel S, Flatley A, Weinmann L, Kremmer E, Meister G
• A multifunctional human Argonaute2-specific monoclonal antibody.
• RNA. 2008; 14: 1244-53
• Display abstract

• Small regulatory RNAs including small interfering RNAs (siRNAs), microRNAs (miRNAs), or Piwi interacting RNAs (piRNAs) guide regulation of gene expression in many different organisms. The Argonaute (Ago) protein family constitutes the cellular binding partners of such small RNAs and regulates gene expression on the levels of transcription, mRNA stability, or translation. Due to the lack of highly specific and potent monoclonal antibodies directed against the different Ago proteins, biochemical analyses such as Ago complex purification and characterization rely on overexpression of tagged Ago proteins. Here, we report the generation and functional characterization of a highly specific monoclonal anti-Ago2 antibody termed anti-Ago2(11A9). We show that anti-Ago2(11A9) is specific for human Ago2 and detects Ago2 in Western blots as well as in immunoprecipitation experiments. We further demonstrate that Ago2 can be efficiently eluted from our antibody by a competing peptide. Finally, we show that anti-Ago2(11A9) recognizes Ago2 in immunofluorescence experiments, and we find that Ago2 not only localizes to cytoplasmic processing bodies (P-bodies) and the diffuse cytoplasm but also to the nucleus. With the anti-Ago2(11A9) antibody we have generated a potent tool that is useful for many biochemical or cell biological applications.

• Rudel S, Meister G
• Phosphorylation of Argonaute proteins: regulating gene regulators.
• Biochem J. 2008; 413: 79-79
• Display abstract

• Members of the Ago (Argonaute) protein family are the mediators of small RNA-guided gene-silencing pathways including RNAi (RNA interference), translational regulation by miRNAs (microRNAs) and transcriptional silencing. Recent findings by Zeng et al. in this issue of the Biochemical Journal demonstrate that Ago proteins are post-translationally modified by phosphorylation of Ser(387). Mutating Ser(387) to alanine leads to reduced localization of human Ago2 to cytoplasmic P-bodies (processing bodies), cellular sites where RNA turnover and, at least in part, miRNA-guided gene regulation occurs. Zeng et al. further show that a member of the MAPK (mitogen-activated protein kinase) signalling pathway phosphorylates Ago2 at Ser(387), suggesting that Ago2-mediated gene silencing might be linked to distinct signalling pathways.

• Kasap M, Sazci A, Ergul E, Akpinar G
• Molecular phylogenetic analysis of methylenetetrahydrofolate reductase family of proteins.
• Mol Phylogenet Evol. 2007; 42: 838-46
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• Methylenetetrahydrofolate reductase (MTHFR) family of proteins catalyze the conversion of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate. They contain a flavin adenine dinucleotide (FAD) as the cofactor and the enzyme in eukaryotes, except in yeast, is known to be allosterically regulated by S-adenosylmethionine. Some cardiovascular diseases, neural tube defects, neuropsychiatric diseases and certain type of cancers in humans are associated with certain polymorphisms of MTHFR. Here, we analyzed 57 of MTHFR polypeptide sequences by multiple sequence alignment and determined previously unrecognized conserved residues that may have a functional or structural importance. A previously unrecognized ATP synthase motif was found in all of the examined plant MTHFRs, suggesting a different functional capability to the plant MTHFRs in addition to the known function. On a phylogenetic tree built, eukaryotic MTHFR proteins formed a clear cluster separated from prokaryotic and archeal relatives. The sequence identities among the eukaryotic MTHFRs were less divergent than the bacterial MTHFRs.

• Brodersen P, Voinnet O
• The diversity of RNA silencing pathways in plants.
• Trends Genet. 2006; 22: 268-80
• Display abstract

• RNA silencing was discovered in plants as a mechanism whereby invading nucleic acids, such as transgenes and viruses, are silenced through the action of small (20-26 nt) homologous RNA molecules. Our understanding of small RNA biology has significantly improved in recent years, and it is now clear that there are several cellular silencing pathways in addition to those involved in defense. Endogenous silencing pathways have important roles in gene regulation at the transcriptional, RNA stability and translational levels. They share a common core of small RNA generator and effector proteins with multiple paralogs in plant genomes, some of which have acquired highly specialized functions. Here, we review recent developments in the plant RNA silencing field that have identified components of specific silencing pathways and have shed light on the mechanisms and biological roles of RNA silencing in plants.

• Parker JS, Barford D
• Argonaute: A scaffold for the function of short regulatory RNAs.
• Trends Biochem Sci. 2006; 31: 622-30
• Display abstract

• Argonaute is the central protein component of RNA-silencing mechanisms. It provides the platform for target-mRNA recognition by short regulatory guide RNA strands and the Slicer catalytic activity for mRNA cleavage in RNA interference. Multiple Argonaute sub-families can be identified phylogenetically yet, despite this diversity, molecular and sequence analyses show that Argonaute proteins share common molecular properties and the capacity to function through a common mechanism. Recently, the members of the Piwi sub-family have been shown to interact with new classes of short regulatory RNAs, Piwi-interacting RNAs (piRNAs) and repeat-associated small interfering RNAs (rasiRNAs), which has implications for developmental processes and introduces a new dimension to the field of RNA silencing.

• Zhang X et al.
• Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis Argonaute1 cleavage activity to counter plant defense.
• Genes Dev. 2006; 20: 3255-68
• Display abstract

• RNA silencing refers to small regulatory RNA-mediated processes that repress endogenous gene expression and defend hosts from offending viruses. As an anti-host defense mechanism, viruses encode suppressors that can block RNA silencing pathways. Cucumber mosaic virus (CMV)-encoded 2b protein was among the first suppressors identified that could inhibit post-transcriptional gene silencing (PTGS), but with little or no effect on miRNA functions. The mechanisms underlying 2b suppression of RNA silencing are unknown. Here, we demonstrate that the CMV 2b protein also interferes with miRNA pathways, eliciting developmental anomalies partially phenocopying ago1 mutant alleles. In contrast to most characterized suppressors, 2b directly interacts with Argonaute1 (AGO1) in vitro and in vivo, and this interaction occurs primarily on one surface of the PAZ-containing module and part of the PIWI-box of AGO1. Consistent with this interaction, 2b specifically inhibits AGO1 cleavage activity in RISC reconstitution assays. In addition, AGO1 recruits virus-derived small interfering RNAs (siRNAs) in vivo, suggesting that AGO1 is a major factor in defense against CMV infection. We conclude that 2b blocks AGO1 cleavage activity to inhibit miRNA pathways, attenuate RNA silencing, and counter host defense. These findings provide insight on the molecular arms race between host antiviral RNA silencing and virus counterdefense.

• Shi H, Tschudi C, Ullu E
• Functional replacement of Trypanosoma brucei Argonaute by the human slicer Argonaute2.
• RNA. 2006; 12: 943-7
• Display abstract

• RNA interference (RNAi) is widespread throughout the eukaryotic lineage, from protozoa to man. Central to all RNAi phenomena is a member of the Argonaute protein family, and, in the case of dsRNA-triggered mRNA cleavage, the Ago protein functions as the RNAi endonuclease or slicer. However, at present there is no definite experimental evidence that slicer Argonautes can be interchanged between distantly related organisms. Here, we show that the human slicer Argonaute2 (HsAgo2), but not HsAgo1, functions in RNAi in the early divergent protozoan Trypanosoma brucei, thus mimicking the situation in mammalian cells. This finding indicates that the basic features of the RNAi mechanism are conserved from T. brucei to man.

• Waltermann M, Steinbuchel A
• Neutral lipid bodies in prokaryotes: recent insights into structure, formation, and relationship to eukaryotic lipid depots.
• J Bacteriol. 2005; 187: 3607-19

• Mohler PJ, Bennett V
• Defects in ankyrin-based cellular pathways in metazoan physiology.
• Front Biosci. 2005; 10: 2832-40
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• Ankyrins are a ubiquitously expressed family of membrane-adaptor proteins found in most vertebrate tissues. Since the first ankyrin polypeptide was identified over 25 years ago, studies in humans, mice, and lower organisms have implicated critical roles for ankyrins in normal metazoan physiology. This review will provide an overview of the ankyrin family and highlight seminal findings in the field which have linked dysfunction in ankyrin-based pathways with defects in metazoan physiology and human disease.

• Sontheimer EJ
• Assembly and function of RNA silencing complexes.
• Nat Rev Mol Cell Biol. 2005; 6: 127-38
• Display abstract

• In the RNA-interference pathway, double-stranded RNA induces sequence-specific mRNA degradation through the action of the RNA-induced silencing complex (RISC). Recent work has provided our first glimpses of the RISC-assembly pathway and uncovered the biochemical roles of critical RISC components. These advances have taken our mechanistic understanding of RNA interference to a new level and promise to improve our ability to exploit this biological process for use in experimental biology and medicine.

• Cramer P
• Structure and function of RNA polymerase II.
• Adv Protein Chem. 2004; 67: 1-42

• Saakov VS
• Alternative pathways of carotenoid biosynthesis in prokaryotes and eukaryotes.
• Dokl Biochem Biophys. 2003; 392: 294-300

• Schiltz M, Fourme R, Prange T
• Use of noble gases xenon and krypton as heavy atoms in protein structure determination.
• Methods Enzymol. 2003; 374: 83-119

• Miller C
• ClC channels: reading eukaryotic function through prokaryotic spectacles.
• J Gen Physiol. 2003; 122: 129-31

• Mullinax RL, Sorge JA
• Preparing lambda libraries for expression of proteins in prokaryotes or eukaryotes.
• Methods Mol Biol. 2003; 221: 271-87

• Li L, Shakhnovich EI, Mirny LA
• Amino acids determining enzyme-substrate specificity in prokaryotic and eukaryotic protein kinases.
• Proc Natl Acad Sci U S A. 2003; 100: 4463-8
• Display abstract

• The binding between a PK and its target is highly specific, despite the fact that many different PKs exhibit significant sequence and structure homology. There must be, then, specificity-determining residues (SDRs) that enable different PKs to recognize their unique substrate. Here we use and further develop a computational procedure to discover putative SDRs (PSDRs) in protein families, whereby a family of homologous proteins is split into orthologous proteins, which are assumed to have the same specificity, and paralogous proteins, which have different specificities. We reason that PSDRs must be similar among orthologs, whereas they must necessarily be different among paralogs. Our statistical procedure and evolutionary model identifies such residues by discriminating a functional signal from a phylogenetic one. As case studies we investigate the prokaryotic two-component system and the eukaryotic AGC (i.e., cAMP-dependent PK, cGMP-dependent PK, and PKC) PKs. Without using experimental data, we predict PSDRs in prokaryotic and eukaryotic PKs, and suggest precise mutations that may convert the specificity of one PK to another. We compare our predictions with current experimental results and obtain considerable agreement with them. Our analysis unifies much of existing data on PK specificity. Finally, we find PSDRs that are outside the active site. Based on our results, as well as structural and biochemical characterizations of eukaryotic PKs, we propose the testable hypothesis of "specificity via differential activation" as a way for the cell to control kinase specificity.

• Schepers U, Kolter T
• RNA Interference: A New Way to Analyze Protein Function.
• Angew Chem Int Ed Engl. 2001; 40: 2437-2439

• Poole A, Jeffares D, Penny D
• Early evolution: prokaryotes, the new kids on the block.
• Bioessays. 1999; 21: 880-9
• Display abstract

• Prokaryotes are generally assumed to be the oldest existing form of life on earth. This assumption, however, makes it difficult to understand certain aspects of the transition from earlier stages in the origin of life to more complex ones, and it does not account for many apparently ancient features in the eukaryotes. From a model of the RNA world, based on relic RNA species in modern organisms, one can infer that there was an absolute requirement for a high-accuracy RNA replicase even before proteins evolved. In addition, we argue here that the ribosome (together with the RNAs involved in its assembly) is so large that it must have had a prior function before protein synthesis. A model that connects and equates these two requirements (high-accuracy RNA replicase and prior function of the ribosome) can explain many steps in the origin of life while accounting for the observation that eukaryotes have retained more vestiges of the RNA world. The later derivation of prokaryote RNA metabolism and genome structure can be accounted for by the two complementary mechanisms of r-selection and thermoreduction.

• Kiselev LL
• [Termination of protein synthesis in eukaryotes and prokaryotes is significantly different].
• Mol Biol (Mosk). 1999; 33: 1054-62

• Szathmary E
• Origins of life. The first two billion years.
• Nature. 1997; 387: 662-3

• Higgins CF
• Stability and degradation of mRNA.
• Curr Opin Cell Biol. 1991; 3: 1013-8
• Display abstract

• Differential mRNA stability plays an important role in the regulation of gene expression. Several recent advances have helped to define the general pathways by which mRNA is degraded in prokaryotic cells, although many details remain to be elucidated. Much less is known about the pathways of degradation in eukaryotic cells, but recent studies on specific systems have highlighted both differences from and similarities to prokaryotic pathways.

• Selker EU
• DNA methylation and chromatin structure: a view from below.
• Trends Biochem Sci. 1990; 15: 103-7
• Display abstract

• An understanding of the function and control of DNA methylation in eukaryotes has been elusive. Studies of Neurospora crassa have led to a model that accounts for the chromosomal distribution of methylation and suggests a basic function for DNA methylation in eukaryotes.

• Glass RE, Nene V, Hunter MG
• Informational suppression as a tool for the investigation of gene structure and function.
• Biochem J. 1982; 203: 1-13

• Streeck RE
• [Synthesis and recombination of DNA from eukaryotic and prokaryotic cells].
• Fortschr Med. 1977; 95: 2005-2005

• EBEL JP
• [RECENT DATA ON THE RELATIONSHIP BETWEEN THE STRUCTURE AND BIOLOGIC ACTIVITY OF TRANSFER RIBONUCLEIC ACIDS].
• Annee Biol. 1965; 59: 205-29

• FAURES A, ERRERA M
• Metabolism of ribonucleic acid and of proteins in x-irradiated isolated thymus nuclei.
• Int J Radiat Biol. 1962; 4: 477-86

• NATHANS D, von EHRENSTEIN, MONRO R, LIPMANN F
• Protein synthesis from aminoacyl-soluble ribonucleic acid.
• Fed Proc. 1962; 21: 127-33

• BRACHET J, FICQ A
• [Not Available].
• Exp Cell Res. 1956; 11: 135-45

• KEDROVSKII BV
• [Ribonucleic acid and its role in the development and function of the cell].
• Usp Sovrem Biol. 1951; 31: 38-56

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