The role of specific regions of ribosomal RNAs in translation termination

The ribosome is a molecular machine that produces proteins in a cell. It consists of RNAs (rRNAs) and proteins. The rRNAs have been implicated in various aspects of protein biosynthesis supporting the idea that they function directly in translation. In this study the direct involvement of rRNA in translation termination was hypothesized and both genetic and biochemical strategies were designed to test this hypothesis. As a result, several regions of rRNAs from both ribosomal subunits were implicated in termination. More specifically, the mutation G1093A in an RNA of the large subunit (23S rRNA) and the mutation C1054A in the small subunit RNA (16S rRNA) of the Escherichia coli ribosome, were shown to affect the binding of the proteins that drive termination, RF1 and RF2. These mutations also caused defects in catalysis of peptidyl-tRNA hydrolysis, the last step of termination. Furthermore, the mutations affected the function of RF2 to a greater extent than that of RF1, a striking result considering the similarity of the RFs. The major defect in RF2 function was consistent with in vivo characteristics of the mutants and can be explained by the inability of the mutant rRNA sites to activate the hydrolytic center, that is the catalytic site for peptidyl-tRNA hydrolysis. Consistent with this explanation is the possibility of a direct interaction between the G1093-region (domain II of 23S rRNA) and the hydrolytic center (most likely domains IV–VI of 23S rRNA). To test that interaction hypothesis selections were performed for mutations in domains IV–VI that compensated for the growth defects caused by G1093A. Several compensatory mutations were isolated which not only restored growth in the presence of G1093A but also appeared to compensate for the termination defects caused by the G1093A. Therefore these results provided genetic evidence for an intramolecular interaction that might lead to peptidyl-tRNA hydrolysis. Finally, a new approach to the study of rRNA involvement in termination was designed. By screening a library of rRNA fragments, a fragment of the 23S rRNA (nt 74-136) was identified that caused readthrough of UGA. The antisense RNA fragment produced a similar effect. The data implicated the corresponding segment of intact 23S rRNA in termination. ^

THE TRANSLATION PRODUCTS OF MOLONEY MURINE SARCOMA VIRUS 124 GENOMIC RNA

Murine sarcoma viruses constitute a class of replication-defective retroviruses. Cellular transformation may be induced by these viruses in vitro; whereas, fibrosarcomas may result in animals infected with them in vivo (Tooze, 1973; Bishop, 1978). Hybridization studies suggest that murine sarcoma viruses arose by recombination between nondefective murine leukemia virus sequences and certain cellular sequences present in uninfected mouse cells (Hu et al., 1977). A specific gene product, however, has not been implicated in murine sarcoma virus transformation.^ One line of murine sarcoma virus-producing cells, Mo-MuSV-clone 124, (Ball et al., 1973), was studied biochemically because it mainly produces the sarcoma virus as a pseudotype packaged with helper murine leukemia virus proteins. The sarcoma viral RNA was translated in a sophisticated cell-free protein synthesizing system (Murphy and Arlinghaus, 1978). The translation products were analyzed by a number of techniques, including electrophoresis in denaturing gels of SDS polyacrylamide, immunoprecipitation, and peptide mapping. The major products of the total RNA purified from the virus preparation were shown to have molecular weights of about 63,000 (P63('gag)), 42,000 (P42), 40,000 (P40), 38,000 (P38), and 23,000 (P23). The size class of mRNA coding for each of the cell-free products was estimated using a poly(A) selection technique and sucrose gradient fractionation. These analyses were used to localize the coding information related to each of the in vitro synthesized cell-free products within the sarcoma virus genome.^ The major findings of these studies were: (1) the 5' half of the sarcoma viral RNA codes for the 63,000 dalton polypeptide and 42,000 - 38,000 dalton polypeptides derived from the "gag" gene; and (2) the 3' half of the sarcoma viral RNA codes for a 38,000 dalton polypeptide and possibly derived from the cellular acquired sequences. ^

Traktaty pedagogiczne Honoraty z Wiśniowskich Zapovej i Klementyny z Tańskich Hoffmanowej – inspiracja czy przeróbka?

As is well know, the long-lasting influence in Poland of Hoffmanowa’s conservative pedagogical treatise « Keepsake of a Good Mother » (1819) was criticized by more progressive Polish women writers of the 19th century, such as Narcyza Żmichowska and Eliza Orzeszkowa. But the unexpected success of that book in other Slav countries later in the 19th century, especially in Bohemia, is less well known. Honorata z Wiśniowskich Zapová (1825-1856), a member of the Galician Polish szlachta who moved to Prague after her marriage in 1841 to the Czech writer K. V. Zap, has been recently exhumed as a Czech writer by Czech and Polish literary criticism: apart from her essays in Czech journals on her native country, much emphasis has been laid on the value of her most important work, Nezabudky, dar našim pannám (Forget-me-not: A Gift to Our Young Ladies, published in 1859), which was the first book of pedagogy dedicated to the instruction of young women in Bohemia. In her preface to this book, Zapová mentions Hoffmanowa as being one of her many sources of inspiration. Wishing to know more about what exactly Zapová owes to Hoffmanowa, in this article I compare Hoffmanowa’s book of pedagogy with Zapová’s. I am astonished to discover that Zapová’s book is in fact an almost literal translation from Polish to Czech of „Keepsake of a Good Mother” – in other words, that it is a plagiarism of Hoffmanowa’s book. The main question I ask here is : why did no one until now, either in 19th-century criticism or in our contemporary criticism, mention this literary fraud before?

Hillel und Schamai, oder die conservative Reform und der stabile Conservatismus : eine Friedensstimme an die Gemeinden Israels und ihre Führer

verf. und hrsg. von L. Adler

Minimal intervention dentistry: part 5. Ultra-conservative approach to the treatment of erosive and abrasive lesions.

The therapeutic management of tooth wear lesions does not require the removal of diseased tissue. Nevertheless, diverse etiological factors may be associated with the condition and they could be difficult to eliminate; this has to be considered when planning therapy. Interceptive procedures should be reserved for such situations while regular monitoring is recommended for other cases, in accordance with advice provided for using the Basic Erosive Wear Examination (BEWE). Direct and indirect adhesive procedures with composite resins allow treatment of most clinical situations, including even extensive restorations. The possibility of managing subsequent interventions should be considered when planning the initial therapeutic approach.

tRNA-derived fragments target the small ribosomal subunit to fine-tune translation

Post-transcriptional cleavage of RNA molecules to generate smaller fragments is a widespread mechanism that enlarges the structural and functional complexity of cellular RNomes. In particular, fragments deriving from both precursor and mature tRNAs represent one of the rapidly growing classes of post-transcriptional RNA pieces. Importantly, these tRNA-derived fragments (tRFs) possess distinct expression patterns, abundance, cellular localizations, or biological roles compared with their parental tRNA molecules (1). Here we present evidence that tRFs from the archaeon Haloferax volcanii directly bind to ribosomes. In a previous genomic screen for ribosome-associated small RNAs we have identified a 26 residue long fragment originating from the 5’ part of valine tRNA (Val-tRF) to be by far the most abundant tRF in H. volcanii (2). The Val-tRF is processed in a stress- dependent manner and was found to primarily target the small ribosomal subunit in vitro and in vivo. Translational activity was markedly reduced in the presence of Val-tRF, while control RNA fragments of similar length did not show inhibition of protein biosynthesis. Crosslinking experiments and subsequent primer extension analyses revealed the Val-tRF interaction site to surround the mRNA path in the 30S subunit. In support of this, binding experiments demonstrated that Val-tRF does compete with mRNAs for ribosome binding. Therefore this tRF represents a ribosome-bound non-protein-coding RNA (ncRNA) capable of regulating gene expression in H. volcanii under environmental stress conditions probably by fine-tuning the rate of protein production (1). (1) Gebetsberger J. and Polacek N. (2013), RNA Biol. 10:1798-1808 (2) Gebetsberger J. et. al. (2012), Archaea, Article ID 260909

Strange Relations. Cultural Translation of Noh Theatre in Ezra Pound’s Dance Poems and W. B. Yeats’s At the Hawk’s Well

Drawing on the reception of Noh drama by Ezra Pound and William Butler Yeats, the article analyses both the literary and cultural ‘translations’ of this form of Japanese theatre in their works, focusing on Yeats’s play At the Hawk’s Well (1917). I conceptualize ‘cultural translation’ as the staging of relations that mark a residual cultural difference. Referred to as ‘foreignizing’ in translation theory, this method enables what Erika Fischer-Lichte has termed a ‘liminal experience’ for the audience –– an effect Yeats intended for the performance of his play. It evokes situations in which opposites collapse and new ways of acting or new combinations of symbols can be tried out. Yeats’s play will be used to sketch how an analysis of relations could serve as a general model for the study of cultural transfer as cultural translation in general. Keywords: cultural translation, translation theory, performance, William Butler Yeats, Itō Michio, Ezra Pound, At the Hawk’s Well

ACL reconstruction with physiological graft tension by intraoperative adjustment of the anteroposterior translation to the uninjured contralateral knee.

PURPOSE Fixation of anterior cruciate ligament (ACL) substitutes with non-physiological anteroposterior translation (APT) worsens outcome. The aim was to present a technique for physiological APT adjustment of the transplant in ACL reconstruction and its outcome at midterm. METHODS In a consecutive series of 28 patients (age 32 ± 11 years, 24 male), chronic ACL deficiency was treated by bone-patella-tendon-bone reconstruction. Transplant APT was adjusted to that of the contralateral uninjured ACL, measured 3, 6, and 12 months postoperatively using the Rolimeter. At a median follow-up of 5.3 years (3-8 years), 82% of the patients were re-evaluated with APT measurement and using IKDC-, Tegner-, Lysholm-Scores, conventional radiographs and MRI. RESULTS No differences in APT (mean ± SD) between uninjured and reconstructed knees were observed after adjustment (6 ± 1 versus 6 ± 1 mm, n.s.). Three months postoperatively, a statistically significant increase in APT (7 ± 1 mm) and a further increase at midterm (9 ± 2 mm) were observed. Patients scored "normal" or "nearly normal", respectively, in 79% (IKDC) and 4 (3-9) points (Tegner; median, range) or 89 ± 9 points (Lysholm; mean ± SD). Radiological evaluation showed no, minimal or moderate joint degeneration in 5, 20 and 75% of patients, respectively. MRI confirmed intact ACL transplants in all patients. CONCLUSION ACL reconstruction using the presented technique was considered successful, as patients did not suffer from subjective instability, radiographic analysis did not provide evidence for graft rupture at midterm. However, APT increase and occurrence of degenerative changes in reconstructed knees at the midterm might not be prevented even by restoration of a physiological APT in ACL reconstruction. The Rolimeter can be used for quick and easy intraoperative indirect control of the applied tension to the ACL transplant by measuring the APT to obtain physiological tensioning resulting in a satisfying outcome at midterm. LEVEL OF EVIDENCE IV.

Ribosome-associated ncRNAs: An emerging class of translation regulators

Accumulating recent evidence identified the ribosome as binding target for numerous small and long non-protein-coding RNAs (ncRNAs) in various organisms of all 3 domains of life. Therefore it appears that ribosome-associated ncRNAs (rancRNAs) are a prevalent, yet poorly understood class of cellular transcripts. Since rancRNAs are associated with the arguable most central enzyme of the cell it seems plausible to propose a role in translation control. Indeed first experimental evidence on small rancRNAs has been presented, linking ribosome association with fine-tuning the rate of protein biosynthesis in a stress-dependent manner.

Self-replicating Replicon-RNA Delivery to Dendritic Cells by Chitosan-nanoparticles for Translation In Vitro and In Vivo.

Self-amplifying replicon RNA (RepRNA) possesses high potential for increasing antigen load within dendritic cells (DCs). The major aim of the present work was to define how RepRNA delivered by biodegradable, chitosan-based nanoparticulate delivery vehicles (nanogel-alginate (NGA)) interacts with DCs, and whether this could lead to translation of the RepRNA in the DCs. Although studies employed virus replicon particles (VRPs), there are no reports on biodegradable, nanoparticulate vehicle delivery of RepRNA. VRP studies employed cytopathogenic agents, contrary to DC requirements-slow processing and antigen retention. We employed noncytopathogenic RepRNA with NGA, demonstrating for the first time the efficiency of RepRNA association with nanoparticles, NGA delivery to DCs, and RepRNA internalization by DCs. RepRNA accumulated in vesicular structures, with patterns typifying cytosolic release. This promoted RepRNA translation, in vitro and in vivo. Delivery and translation were RepRNA concentration-dependent, occurring in a kinetic manner. Including cationic lipids with chitosan during nanoparticle formation enhanced delivery and translation kinetics, but was not required for translation of immunogenic levels in vivo. This work describes for the first time the characteristics associated with chitosan-nanoparticle delivery of self-amplifying RepRNA to DCs, leading to translation of encoded foreign genes, namely influenza virus hemagglutinin and nucleoprotein.