Plant and animal microRNAs : similarities and differences

Plant and animal microRNAs (miRNAs) are evolutionarily ancient small RNAs, ∼19-24 nucleotides in length, that are generated by cleavage from larger highly structured precursor molecules. In both plants and animals, miRNAs posttranscriptionally regulate gene expression through interactions with their target mRNAs, and these targets are often genes involved with regulating key developmental events. Despite these similarities, plant and animal miRNAs exert their control in fundamentally different ways. Generally, animal miRNAs repress gene expression by mediating translational attenuation through (multiple) miRNA-binding sites located within the 3′ untranslated region of the target gene. In contrast, almost all plant miRNAs regulate their targets by directing mRNA cleavage at single sites in the coding regions. These and other differences suggest that the two systems may have originated independently, possibly as a prerequisite to the development of complex body plans. © Springer-Verlag 2005.

Association of sporadic chondrocalcinosis with a -4-basepair G-to-A transition in the 5′-untranslated region of ANKH that promotes enhanced expression of ANKH protein and excess generation of extracellular inorganic pyrophosphate

Objective Certain mutations in ANKH, which encodes a multiple-pass transmembrane protein that regulates inorganic pyrophosphate (PPi) transport, are linked to autosomal-dominant familial chondrocalcinosis. This study investigated the potential for ANKH sequence variants to promote sporadic chondrocalcinosis. Methods ANKH variants identified by genomic sequencing were screened for association with chondrocalcinosis in 128 patients with severe sporadic chondrocalcinosis or pseudogout and in ethnically matched healthy controls. The effects of specific variants on expression of common markers were evaluated by in vitro transcription/translation. The function of these variants was studied in transfected human immortalized CH-8 articular chondrocytes. Results Sporadic chondrocalcinosis was associated with a G-to-A transition in the ANKH 5′-untranslated region (5′-UTR) at 4 bp upstream of the start codon (in homozygotes of the minor allele, genotype relative risk 6.0, P = 0.0006; overall genotype association P = 0.02). This -4-bp transition, as well as 2 mutations previously linked with familial and sporadic chondrocalcinosis (+14 bp C-to-T and C-terminal GAG deletion, respectively), but not the French familial chondrocalcinosis kindred 143-bp T-to-C mutation, increased reticulocyte ANKH transcription/ANKH translation in vitro. Transfection of complementary DNA for both the wild-type ANKH and the -4-bp ANKH protein variant promoted increased extracellular PPi in CH-8 cells, but unexpectedly, these ANKH mutants had divergent effects on the expression of extracellular PPi and the chondrocyte hypertrophy marker, type X collagen. Conclusion A subset of sporadic chondrocalcinosis appears to be heritable via a -4-bp G-to-A ANKH 5′-UTR transition that up-regulates expression of ANKH and extracellular PPi in chondrocyte cells. Distinct ANKH mutations associated with heritable chondrocalcinosis may promote disease by divergent effects on extracellular PPi and chondrocyte hypertrophy, which is likely to mediate differences in the clinical phenotypes and severity of the disease.

The adenine-rich tract in the 5ʹ-end of the hepatitis C virus ORF encodes a peptide regulating the binding of the C protein to RNA

Hepatitis C virus (HCV ) core (C) protein is thought to bind to viral RNA before it undergoes oligomerization leading to RNA encapsidation. Details of these events are so far unknown. The 5ʹ-terminal C protein coding sequence that includes an adenine (A)-rich tract is a part of an internal ribosome entry site(IRES). This nucleotide sequence but not the corresponding protein sequence is needed for proper initiation of translation of viral RNA by an IRES-dependent mechanism. In this study, we examined the importance of this sequence for the ability of the C protein to bind to viral RNA. Serially truncated C proteins with deletions from 10 up to 45 N-terminal amino acids were expressed in Escherichia coli, purified and tested for binding to viral RNA by a gel shift assay. The results showed that truncation of the C protein from its N-terminus by more than 10 amino acids abolished almost completely its expression in E. coli. The latter could be restored by adding a tag to the N-terminus of the protein. The tagged proteins truncated by 15 or more amino acids showed an anomalous migration in SDS-PAGE. Truncation by more than 20 amino acids resulted in a complete loss of ability of tagged C protein to bind to viral RNA. These results provide clues to the early events in the C protein - RNA interactions leading to C protein oligomerization, RNA encapsidation and virion assembly.

De Novo Transcriptome Sequence Assembly and Analysis of RNA Silencing Genes of Nicotiana benthamiana

Background: Nicotiana benthamiana has been widely used for transient gene expression assays and as a model plant in the study of plant-microbe interactions, lipid engineering and RNA silencing pathways. Assembling the sequence of its transcriptome provides information that, in conjunction with the genome sequence, will facilitate gaining insight into the plant's capacity for high-level transient transgene expression, generation of mobile gene silencing signals, and hyper-susceptibility to viral infection. Methodology/Results: RNA-seq libraries from 9 different tissues were deep sequenced and assembled, de novo, into a representation of the transcriptome. The assembly, of16GB of sequence, yielded 237,340 contigs, clustering into 119,014 transcripts (unigenes). Between 80 and 85% of reads from all tissues could be mapped back to the full transcriptome. Approximately 63% of the unigenes exhibited a match to the Solgenomics tomato predicted proteins database. Approximately 94% of the Solgenomics N. benthamiana unigene set (16,024 sequences) matched our unigene set (119,014 sequences). Using homology searches we identified 31 homologues that are involved in RNAi-associated pathways in Arabidopsis thaliana, and show that they possess the domains characteristic of these proteins. Of these genes, the RNA dependent RNA polymerase gene, Rdr1, is transcribed but has a 72 nt insertion in exon1 that would cause premature termination of translation. Dicer-like 3 (DCL3) appears to lack both the DEAD helicase motif and second dsRNA binding motif, and DCL2 and AGO4b have unexpectedly high levels of transcription. Conclusions: The assembled and annotated representation of the transcriptome and list of RNAi-associated sequences are accessible at www.benthgenome.com alongside a draft genome assembly. These genomic resources will be very useful for further study of the developmental, metabolic and defense pathways of N. benthamiana and in understanding the mechanisms behind the features which have made it such a well-used model plant. © 2013 Nakasugi et al.

Rice ragged stunt oryzavirus genome segment S4 could encode an RNA dependent RNA polymerase and a second protein of unknown function

The complete nucleotide sequence of genome segment S4 of rice ragged stunt oryzavirus (RRSV, Thai-isolate) was determined. The 3823 bp sequence contains two large open reading frames (ORFs). ORF1, spanning nucleotides 12 to 3776, is capable of encoding a protein of M(r) 141,380 (P4a). The P4a amino acid sequence predicted from the nucleotide sequence contains sequence motifs conserved in RNA-dependent RNA polymerases (RDRPs). When compared for evolutionary relationships with RDRPs of other reoviruses using the amino acid sequences around the conserved GDD motif, P4a was shown to be more related to Nilaparvata lugens reovirus and reovirus serotype 3 than to rice dwarf phytoreovirus, bovine rotavirus or bluetongue virus. The ORF2, spanning nucleotides 491 to 1468, is out of frame with ORF1 and is capable of encoding a protein of 36, 920 (P4b). Coupled in vitro transcription-translation from cloned ORF2 in wheat germ extract confirmed the existence of ORF2 but in vivo production and possible function of P4b is yet to be determined.

Translation of addictions science into practice : update and future directions.

Recent advances in the understanding of the genetic, neurochemical, behavioral and cultural underpinnings of addiction have led to rapid advances in the understanding of addiction as a disease. In fact, advances in basic science and the development of new pharmacological and behavioral therapies associated with them are appearing faster than can be assimilated not only by clinical researchers but practitioners and policy makers as well. Translation of science-based addictions knowledge into improved prevention, assessment and treatment, and communication of these changes to researchers and practitioners are significant challenges to the field. The general aim of this book is to summarize current and potential linkages between advances in addiction science and innovations in clinical practice. Whilst this book is primarily focused on translation, it also encompasses some scientific advances that are relevant to dissemination, and the book is itself a tool for disseminating innovative thinking. The goal is to generate interest in application opportunities from both recent research and theoretical advances.

Translation and news making : a study of contemporary arabic television

Arabic satellite television has recently attracted tremendous attention in both the academic and professional worlds, with a special interest in Aljazeera as a curious phenomenon in the Arab region. Having made a household name for itself worldwide with the airing of the Bin Laden tapes, Aljazeera has set out to deliberately change the culture of Arabic journalism, as it has been repeatedly stated by its current General Manager Waddah Khanfar, and to shake up the Arab society by raising awareness to issues never discussed on television before and challenging long-established social and cultural values and norms while promoting, as it claims, Arab issues from a presumably Arab perspective. Working within the meta-frame of democracy, this Qatari-based network station has been received with mixed reactions ranging from complete support to utter rejection in both the west and the Arab world. This research examines the social semiotics of Arabic television and the socio-cultural impact of translation-mediated news in Arabic satellite television, with the aim to carry out a qualitative content analysis, informed by framing theory, critical linguistic analysis, social semiotics and translation theory, within a re-mediation framework which rests on the assumption that a medium “appropriates the techniques, forms and social significance of other media and attempts to rival or refashion them in the name of the real" (Bolter and Grusin, 2000: 66). This is a multilayered research into how translation operates at two different yet interwoven levels: translation proper, that is the rendition of discourse from one language into another at the text level, and translation as a broader process of interpretation of social behaviour that is driven by linguistic and cultural forms of another medium resulting in new social signs generated from source meaning reproduced as target meaning that is bound to be different in many respects. The research primarily focuses on the news media, news making and reporting at Arabic satellite television and looks at translation as a reframing process of news stories in terms of content and cultural values. This notion is based on the premise that by its very nature, news reporting is a framing process, which involves a reconstruction of reality into actualities in presenting the news and providing the context for it. In other words, the mediation of perceived reality through a media form, such as television, actually modifies the mind’s ordering and internal representation of the reality that is presented. The research examines the process of reframing through translation news already framed or actualized in another language and argues that in submitting framed news reports to the translation process several alterations take place, driven by the linguistic and cultural constraints and shaped by the context in which the content is presented. These alterations, which involve recontextualizations, may be intentional or unintentional, motivated or unmotivated. Generally, they are the product of lack of awareness of the dynamics and intricacies of turning a message from one language form into another. More specifically, they are the result of a synthesis process that consciously or subconsciously conforms to editorial policy and cultural interpretive frameworks. In either case, the original message is reproduced and the news is reframed. For the case study, this research examines news broadcasts by the now world-renowned Arabic satellite television station Aljazeera, and to a lesser extent the Lebanese Broadcasting Corporation (LBC) and Al- Arabiya where access is feasible, for comparison and crosschecking purposes. As a new phenomenon in the Arab world, Arabic satellite television, especially 24-hour news and current affairs, provides an interesting area worthy of study, not only for its immediate socio-cultural and professional and ethical implications for the Arabic media in particular, but also for news and current affairs production in the western media that rely on foreign language sources and translation mediation for international stories.

Translation of addictions science into practice

Recent advances in the understanding of the genetic, neurochemical, behavioral and cultural underpinnings of addiction have led to rapid advances in the understanding of addiction as a disease. In fact, advances in basic science and the development of new pharmacological and behavioral therapies associated with them are appearing faster than can be assimilated not only by clinical researchers but practitioners and policy makers as well. Translation of science-based addictions knowledge into improved prevention, assessment and treatment, and communication of these changes to researchers and practitioners are significant challenges to the field. The general aim of this book is to summarize current and potential linkages between advances in addiction science and innovations in clinical practice. Whilst this book is primarily focused on translation, it also encompasses some scientific advances that are relevant to dissemination, and the book is itself a tool for disseminating innovative thinking. The goal is to generate interest in application opportunities from both recent research and theoretical advances.

Lineage replacement accompanying duplication and rapid fixation of an RNA element in the nsP3 gene in a species of alphavirus

A sequence of thirty-six nucleotides in the nsP3 gene of Ross River virus (RRV), coding for the amino acid sequence HADTVSLDSTVS, was duplicated some time between 1969 and 1979 coinciding with the appearance of a new lineage of this virus and with a major outbreak of Epidemic Polyarthritis among residents of the Pacific Islands. This lineage of RRV continues to circulate throughout Australia and both earlier lineages, which lacked the duplicated element, now are extinct. Multiple copies of several other elements also were observed in this region of the nsP3 gene in all lineages of RRV. Multiple copies of one of these, coding for the amino acid sequence P*P*PR, were detected in the C-terminal region of the nsP3 protein of all alphaviruses except those of African origin. The fixation of duplications and insertions in 3' region of nsP3 genes from all lineages of alphaviruses, suggests they provide some fitness advantage

Physical and functional interaction of the archaeal single-stranded DNA-binding protein SSB with RNA polymerase

Archaeal transcription utilizes a complex multisubunit RNA polymerase and the basal transcription factors TBP and TF(II)B, closely resembling its eukaryal counterpart. We have uncovered a tight physical and functional interaction between RNA polymerase and the single-stranded DNA-binding protein SSB in Sulfolobus solfataricus. SSB stimulates transcription from promoters in vitro under TBP-limiting conditions and supports transcription in the absence of TBP. SSB also rescues transcription from repression by reconstituted chromatin. We demonstrate the potential for promoter melting by SSB, suggesting a plausible basis for the stimulation of transcription. This stimulation requires both the single-stranded DNA-binding domain and the acidic C-terminal tail of the SSB. The tail forms a stable interaction with RNA polymerase. These data reveal an unexpected role for single-stranded DNA-binding proteins in transcription in archaea.