The target of rapamycin (TOR) proteins

Rapamycin potently inhibits downstream signaling from the target of rapamycin (TOR) proteins. These evolutionarily conserved protein kinases coordinate the balance between protein synthesis and protein degradation in response to nutrient quality and quantity. The TOR proteins regulate (i) the initiation and elongation phases of translation, (ii) ribosome biosynthesis, (iii) amino acid import, (iv) the transcription of numerous enzymes involved in multiple metabolic pathways, and (v) autophagy. Intriguingly, recent studies have also suggested that TOR signaling plays a critical role in brain development, learning, and memory formation.

Visualization of biochemical networks in living cells

Functional annotation of novel genes can be achieved by detection of interactions of their encoded proteins with known proteins followed by assays to validate that the gene participates in a specific cellular function. We report an experimental strategy that allows for detection of protein interactions and functional assays with a single reporter system. Interactions among biochemical network component proteins are detected and probed with stimulators and inhibitors of the network. In addition, the cellular location of the interacting proteins is determined. We used this strategy to map a signal transduction network that controls initiation of translation in eukaryotes. We analyzed 35 different pairs of full-length proteins and identified 14 interactions, of which five have not been observed previously, suggesting that the organization of the pathway is more ramified and integrated than previously shown. Our results demonstrate the feasibility of using this strategy in efforts of genomewide functional annotation.

Mammalian orthologues of a yeast regulator of nonsense transcript stability.

All eukaryotes that have been studied to date possess the ability to detect and degrade transcripts that contain a premature signal for the termination of translation. This process of nonsense-mediated RNA decay has been most comprehensively studied in the yeast Saccharomyces cerevisiae where at least three trans-acting factors (Upf1p through Upf3P) are required. We have cloned cDNAs encoding human and murine homologues of Upf1p, termed rent1 (regulator of nonsense transcripts). Rent1 is the first identified mammalian protein that contains all of the putative functional elements in Upf1p including zinc finger-like and NTPase domains, as well as all motifs common to members of helicase superfamily I. Moreover, expression of a chimeric protein, N and C termini of Upf1p, complements the Upf1p-deficient phenotype in yeast. Thus, despite apparent differences between yeast and mammalian nonsense-mediated RNA decay, these data suggest that the two pathways use functionally related machinery.

Poly(rC) binding protein 2 binds to stem-loop IV of the poliovirus RNA 5' noncoding region: identification by automated liquid chromatography-tandem mass spectrometry.

The 5' noncoding region of poliovirus RNA contains an internal ribosome entry site (IRES) for cap-independent initiation of translation. Utilization of the IRES requires the participation of one or more cellular proteins that mediate events in the translation initiation reaction, but whose biochemical roles have not been defined. In this report, we identify a cellular RNA binding protein isolated from the ribosomal salt wash of uninfected HeLa cells that specifically binds to stem-loop IV, a domain located in the central part of the poliovirus IRES. The protein was isolated by specific RNA affinity chromatography, and 55% of its sequence was determined by automated liquid chromatography-tandem mass spectrometry. The sequence obtained matched that of poly(rC) binding protein 2 (PCBP2), previously identified as an RNA binding protein from human cells. PCBP2, as well as a related protein, PCBP1, was over-expressed in Escherichia coli after cloning the cDNAs into an expression plasmid to produce a histidine-tagged fusion protein. Specific interaction between recombinant PCBP2 and poliovirus stem-loop IV was demonstrated by RNA mobility shift analysis. The closely related PCBP1 showed no stable interaction with the RNA. Stem-loop IV RNA containing a three nucleotide insertion that abrogates translation activity and virus viability was unable to bind PCBP2.

Presence of mRNA for glutamic acid decarboxylase in both excitatory and inhibitory neurons.

Neurons in very low density hippocampal cultures that are physiologically identified as either GABAergic inhibitory or glutamatergic excitatory all contain mRNA for the gamma-aminobutyric acid (GABA) synthetic enzyme, glutamic acid decarboxylase (GAD), as detected by single cell mRNA amplification and PCR. However, consistent with the physiology, immunocytochemistry revealed that only a subset of the neurons stain for either GAD protein or GABA. A similar fraction hybridize with RNA probes for GAD65 and GAD67. Hippocampal CA1 pyramidal neurons in slice preparations, which are traditionally thought to be excitatory, also contain mRNA for GAD65 and GAD67. Hippocampal neurons in culture did not contain mRNA for two other neurotransmitter synthesizing enzymes, tyrosine hydroxylase, and choline acetyl transferase. These data suggest that in some neurons, presumably the excitatory neurons, GAD mRNA is selectively regulated at the level of translation. We propose that neurotransmitter phenotype may be posttranscriptionally regulated and neurons may exhibit transient phenotypic plasticity in response to environmental influences.

Expression of a recoded nuclear gene inserted into yeast mitochondrial DNA is limited by mRNA-specific translational activation.

Genetic code differences prevent expression of nuclear genes within Saccharomyces cerevisiae mitochondria. To bridge this gap a synthetic gene, ARG8m, designed to specify an arginine biosynthetic enzyme when expressed inside mitochondria, has been inserted into yeast mtDNA in place of the COX3 structural gene. This mitochondrial cox3::ARG8m gene fully complements a nuclear arg8 deletion at the level of cell growth, and it is dependent for expression upon nuclear genes that encode subunits of the COX3 mRNA-specific translational activator. Thus, cox3::ARG8m serves as a mitochondrial reporter gene. Measurement of cox3::ARG8m expression at the levels of steady-state protein and enzymatic activity reveals that glucose repression operates within mitochondria. The levels of this reporter vary among strains whose nuclear genotypes lead to under- and overexpression of translational activator subunits, in particular Pet494p, indicating that mRNA-specific translational activation is a rate-limiting step in this organellar system. Whereas the steady-state level of cox3::ARG8m mRNA was also glucose repressed in an otherwise wild-type strain, absence of translational activation led to essentially repressed mRNA levels even under derepressing growth conditions. Thus, the mRNA is stabilized by translational activation, and variation in its level may be largely due to modulation of translation.

The 5' coding region of Paramecium surface antigen genes controls mutually exclusive transcription.

Paramecium tetraurelia stock 51 can express at least 11 different types of surface antigens, yet only a single type is expressed on the surface of an individual cell at any one time. The differential expression of stock 51 type A and B surface antigen genes (51A and 51B) is regulated at the level of transcription. Previously, we reported that nucleotide sequences upstream of position -26 (relative to the start of translation) in the 51A and 51B surface antigen genes are necessary for transcriptional activity but are not sufficient to direct differential transcriptional control. In this report we demonstrate that at least some of the critical elements necessary for differential transcription of the 51A and 51B genes lie within the 5' coding region. A hybrid gene that contains 51B upstream sequences (-475 to +1) attached to the ATG start codon of 51A is not cotranscribed with the 51B gene. In contrast, further substitution with 51B sequences (-1647 to +885) allows the chimeric gene to be coexpressed with 51B. A different hybrid gene containing a substitution of 51B sequence from -26 to +885 in the 51A gene is also coexpressed with 51B, revealing that the critical elements within the coding region of 51B do not require 51B upstream sequences for their effect. Coinjection of the 51A gene with the chimeric gene that contains 51B up to +885 showed that the same sequences that allow coexpression with 51B prevent cotranscription with 51A. Together, these results demonstrate that a region downstream of the transcriptional start site between nucleotide positions +1 and +885 (relative to translational start) is necessary to control differential transcriptional activity.

In vitro activation of the interferon-induced, double-stranded RNA-dependent protein kinase PKR by RNA from the 3' untranslated regions of human alpha-tropomyosin.

The cellular kinase known as PKR (protein kinase RNA-activated) is induced by interferon and activated by RNA. PKR is known to have antiviral properties due to its role in translational control. Active PKR phosphorylates eukaryotic initiation factor 2 alpha and leads to inhibition of translation, including viral translation. PKR is also known to function as a tumor suppressor, presumably by limiting the rate of tumor-cell translation and growth. Recent research has shown that RNA from the 3' untranslated region (3'UTR) of human alpha-tropomyosin has tumor-suppressor properties in vivo [Rastinejad, F., Conboy, M. J., Rando, T. A. & Blau, H. M. (1993) Cell 75, 1107-1117]. Here we report that purified RNA from the 3'UTR of human alpha-tropomyosin can inhibit in vitro translation in a manner consistent with activation of PKR. Inhibition of translation by tropomyosin 3'UTR RNA was observed in a rabbit reticulocyte lysate system, which is known to contain endogenous PKR but was not seen in wheat germ lysate, which is not responsive to a known activator of PKR. A control RNA purified in the same manner as the 3'UTR RNA did not inhibit translation in either system. The inhibition of translation observed in reticulocyte lysates was prevented by the addition of adenovirus virus-associated RNA1 (VA RNAI), an inhibitor of PKR activation. Tropomyosin 3'UTR RNA was bound by immunoprecipitated PKR and activated the enzyme in an in vitro kinase assay. These data suggest that activation of PKR could be the mechanism by which tropomyosin 3'UTR RNA exerts its tumor-suppression activity in vivo.

The regulation of professional advertising practices in Spain, 1988–2013

From the perspective of the sociology of professions, every professional activity should have its own clearly circumscribed and regulated sphere of action. Such an articulation facilitates the regulation of the production of a given profession as well as the way in which it is practiced. The purpose of the research reported here was to provide a comprehensive review and evaluation of the regulatory framework governing the advertising sector in Spain. To this end, the authors analysed external regulatory legislation and self-regulatory codes extracted from the data base of the Asociación para la Autoregulación de la Comunicación Comercial (Autocontrol) that had been enacted or adopted between 1988, the year that Law 11/1998 on General Telecommunications entered into force, and 2003 as well as other relevant documents retrieved from the Boletin Oficial del Estado (BOE) pertaining to the same period. Findings indicate that although there has been a groundswell of legislation governing advertising practices in Spain since 1988, especially at the regional level, lawmakers have focused on the content of advertising messages and shown very little interest in regulating the professions of advertising and public relations. Furthermore, Spanish legislation enacted in 2003 and EU policies appear to have encouraged the adoption of voluntary codes of ethics. Sectors traditionally subject to mandatory advertising regulation, either due to the vulnerability of their target audiences or the potential impact of their commercial messages on public health or the environment, are more likely to develop self-regulatory codes of conduct than others

Forgiven, not Forgotten: Communicative Translation Activities in Second Language Teaching

Although frequently discarded and despised in the 20th century, translation now seems to find wider acceptance within the Second Language Teaching (SLT) field. However, it still has a long way to go before recovering its due place in the L2 classroom. The aim of this paper is to suggest a number of translation (and interpreting)-based activities covering the different competence levels, thus showing that communicative content and translation can perfectly go hand in hand so that old, unjustified prejudices can be superseded once and for all.

How Peripheral Is the Periphery? Translating Portugal Back and Forth. Essays in Honour of João Ferreira Duarte

This volume is a result of the need to reflect upon Portugal’s position from the viewpoint of the literary assets imported and exported through translation. It brings together a number of scholars working in the field of Translation Studies directly concerned with the Portuguese cultural system in order to analyse this question from various theoretical perspectives and from case studies of translation flows and movements in Portuguese culture. By Translating Portugal Back and Forth, the articles discuss issues such as: how can one draw the borderline between a peripheral and a semi-peripheral system? Is this borderline useful or necessary? How peripheral is the Portuguese cultural system as far as translation transfers are concerned? How stable or pacific has this positioning been? Does the economic and historical perception of Portugal as peripheral entail that, from the viewpoint of translation, it would behave similarly? By addressing some of these questions, and as shown by the (second) subtitle – Essays in Honour of João Ferreira Duarte –, the volume pays homage to one of the most prominent Translation Studies scholars in Portugal, who has extensively reflected on the binary discourse on translation, its metaphors and images.

On the pragmatics of social theory. The case of Elias's "On the Process of Civilization"

This paper proposes a new approach to the study of sociological classics. This approach is pragmatic in character. It draws upon the social pragmatism of G.H. Mead and the sociology of texts of D.F. McKenzie. Our object of study is Norbert Elias’s On the Process of Civilization. The pragmatic genealogy of this book reveals the importance of taking materiality seriously. By documenting the successive entanglements between human agency and non-human factors, we discuss the origins of the book in the 1930s, how it was forgotten for thirty years, and how in the mid-1970s it became a sociological classic. We explain canonization as a matter of fusion between book’s material form and its content, in the context of the paperback revolution of the 1960s, the events of May 1968, and the demise of Parsons’ structural functionalism, and how this provided Elias with an opportunity to advance his model of sociology.

Zhurnal analiticheskoĭ khimii = Journal of analytical chemistry /

Latest issue consulted: T. 42, vyp. 12 (dek. 1987).

Muscularity, the habitus and the social construction of gender: Towards a gender-relevant physical education

This paper begins to develop the concept of gender-relevant physical education, combining the work of Pierre Bourdieu and his notion of the habitus and feminist philosopher Iris Marion Young's analysis of feminine motility. It draws on data generated from a study of young people's articulation of the relationships between muscularity, physicality and gender. The social construction of the body has been of central importance to the construction of femininities and masculinities, and has formed an enduring meta-theme through much of the research on physical education and gender. We build on the young people's insights to argue that Bourdieu's notions of the habitus and the exchange of physical capital provide a useful means of conceptualizing issues of embodiment and gender in school physical education and sport. We conclude by sketching an outline of gender-relevant physical education as a process of interrupting the habitus.