Translational recoding as a feedback controller : systems approaches reveal polyamine-specific effects on the antizyme ribosomal frameshift

Peer reviewedPublisher PD

FSscan: a mechanism-based program to identify +1 ribosomal frameshift hotspots

In +1 programmed ribosomal frameshifting (PRF), ribosomes skip one nucleotide toward the 3′-end during translation. Most of the genes known to demonstrate +1 PRF have been discovered by chance or by searching homologous genes. Here, a bioinformatic framework called FSscan is developed to perform a systematic search for potential +1 frameshift sites in the Escherichia coli genome. Based on a current state of the art understanding of the mechanism of +1 PRF, FSscan calculates scores for a 16-nt window along a gene sequence according to different effects of the stimulatory signals, and ribosome E-, P- and A-site interactions. FSscan successfully identified the +1 PRF site in prfB and predicted yehP, pepP, nuoE and cheA as +1 frameshift candidates in the E. coli genome. Empirical results demonstrated that potential +1 frameshift sequences identified promoted significant levels of +1 frameshifting in vivo. Mass spectrometry analysis confirmed the presence of the frameshifted proteins expressed from a yehP-egfp fusion construct. FSscan allows a genome-wide and systematic search for +1 frameshift sites in E. coli. The results have implications for bioinformatic identification of novel frameshift proteins, ribosomal frameshifting, coding sequence detection and the application of mass spectrometry on studying frameshift proteins

A new kinetic model reveals the synergistic effect of E-, P- and A-sites on +1 ribosomal frameshifting

Programmed ribosomal frameshifting (PRF) is a process by which ribosomes produce two different polypeptides from the same mRNA. In this study, we propose three different kinetic models of +1 PRF, incorporating the effects of the ribosomal E-, P- and A-sites toward promoting efficient +1 frameshifting in Escherichia coli. Specifically, the timing of E-site tRNA dissociation is discussed within the context of the kinetic proofreading mechanism of aminoacylated tRNA (aa-tRNA) selection. Mathematical modeling using previously determined kinetic rate constants reveals that destabilization of deacylated tRNA in the E-site, rearrangement of peptidyl-tRNA in the P-site, and availability of cognate aa-tRNA corresponding to the A-site act synergistically to promote efficient +1 PRF. The effect of E-site codon:anticodon interactions on +1 PRF was also experimentally examined with a dual fluorescence reporter construct. The combination of predictive modeling and empirical testing allowed the rate constant for P-site tRNA slippage (ks) to be estimated as ks ≈1.9 s−1 for the release factor 2 (RF2) frameshifting sequence. These analyses suggest that P-site tRNA slippage is the driving force for +1 ribosomal frameshifting while the presence of a ‘hungry codon’ in the A-site and destabilization in the E-site further enhance +1 PRF in E. coli

A Prospective Open Labelled Non-Randomized Phase-II Clinical Trial on Pavattai Kudineer in the management of Thandaga Vatham

The disease THANDAGA VATHAM can be correlated with lumbar spondylosis is a degenerative changes in disc and lumbar spine. Disc degenerate is age related and starts in 4rd decade. Low back pain (LBP) affects approximately 60–85% in adult age. Fortunately, for the large majority of individuals, symptoms are mild and transient, with 90% subsiding within 6 weeks. Chronic low back pain, defined as pain symptoms persisting more than 3 months. Despite the high prevalence of low back pain within the general population, the diagnostic approach and therapeutic options are diverse and often inconsistent, resulting in rising costs and variability in management throughout the country. In part, this is due to the difficulty establishing a clear etiology for most patients, with known non receptive pain generators identified throughout the axial spine. Lumbar Spondylosis is defined as degenerative changes, without an associated disruption or defect in the vertebral disc. The clinical symptoms of degenerative spondylosis, or with neurogenic claudication and/or radicular pain, with or without axial back pain. In India Lumbar spondylosis affects 60-85% of the adults during some point in their lives. 84% of men and 74% of women are suffering from Lumbar spondylosis in the age group of 45-64 years of population is prone to develop disorders of vertebral column, like lumbar spondylosis, Prolapsed Intervertebral Disc (PID), osteoporosis and the degenerative disease of spine. Majority of them are suffering from PID or lumbar spondylosis. The principle line of treatment is to use of steroids, physiotherapy, NSAID. Which are having severe side effects on hepatioand renal systems. The treatment such as surgical intervention may inherit drawbacks like disability. Thandaga vatham is a characterized by severe pain in which the body is rendered like a log of wood, unable to stretch the limbs and pass urine and stools. In Thandaga vatham, according to Yugimuni and T.V. Sambasivampillai, can be clinically correlated in modern medicine is lumbar spondylosis. It is one among the 80 types of vatha diseases. This research will deal elaborately discussed about etiology, pathology, clinical features and complications of Thandaga vatham and Prognosis after using Siddha treatment the Pavattaiilai kudieer (Internal). So, in this situation I have been choosen Pavattai ilai kudieer for the management in Lumbar spondylosis. Conventional medicines and surgical treatments end up with side effects and leave with defect and detriment like recurrence of pain, disability & nerve root damage. This study is concerned with A Prospective open labelled non randomised Phase-II Clinical trial to assess the therapeutic efficacy of the siddha formulation PAVATTAI ILAI KUDINEER for the treatment of THANDAGA VATHAM (LUMBAR SPONDYLOSIS), described in Gunapadam Mooligai Vaguppu Part-I text book to detect whether any significant improvements and relief can be done. Totally 40 patients with Thandaga Vatham (30 In patients, 30 Out patients of both sex) were selected randomly from OPD of PothuMaruthuvam, Govt. Siddha Medical College, Tirunelveli, Tamil Nadu. They were given PavattaiIlai Kudineer 30ml twice a day for 45 days. After the course of treatment majority of cases showed good response which is statistically significant

CT-DNA interaction, Fluorescence binding and Microbial activity of Schiff base metal complexes derived from 4-bromobenzaldehyde and 2-amino-6-methylbenzothiazole

A novel bidentate Schiff base ligand was prepared by the condensation of 4-bromobenzaldehyde and 2-amino-6-methylbenzothiazole. The ligand has been characterized by microanalytical data such as IR, UV-Vis, 1H-NMR, EPR and Mass spectra. Its complexes of Cu (II), Ni (II), Co (II), and Zn (II) were synthesized and characterized by spectral and analytical techniques. This fact shows the composition of the type ML2 where, M= Cu (II), Ni (II), Co (II), and Zn (II). The UVV is spectral data implies that the complexes have octahedral geometry. They show non electrical conductance which reveals that these chelates are electrolytes. Their magnetic susceptibility values afford evidence for the monomeric nature. Absorption and gel electrophorese experiments have been accepted out on the interaction of the complexes with CT-DNA and results suggest that all the complexes of DNA and Fluorescence binding. The Antimicrobial activities of the compounds are tested in vitro against four bacteria

Effect of jerky movement of ring rail on quality of ring yarn

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Polyamine sensing by nascent ornithine decarboxylase antizyme stimulates decoding of its mRNA

Polyamines are essential organic polycations with multiple cellular functions relevant for cell division, cancer and ageing(1-3). Regulation of polyamine synthesis is mainly achieved by controlling the activity of ornithine decarboxylase (ODC) through an unusual mechanism involving ODC antizyme(1,4), the binding of which disrupts homodimeric ODC and targets it for ubiquitin-independent degradation by the 26S proteasome(5). Whereas mammals express several antizyme genes(6), we have identified a single orthologue, termed OAZ1, in Saccharomyces cerevisiae(7). Similar to its mammalian counterparts, OAZ1 synthesis is induced with rising intracellular polyamine concentrations, which also inhibit ubiquitin-dependent degradation of the OAZ1 protein(7). Together, these mechanisms contribute to a homeostatic feedback regulation of polyamines(1,7,8). Antizyme synthesis involves a conserved +1 ribosomal frameshifting (RFS) event at an internal STOP codon during decoding of its messenger RNA(6-10). Here we used S. cerevisiae OAZ1 to dissect the enigmatic mechanism underlying polyamine regulation of RFS. In contrast with previous assumptions, we report here that the nascent antizyme polypeptide is the relevant polyamine sensor that operates in cis to negatively regulate upstream RFS on the polysomes, where its own mRNA is being translated. At low polyamine levels, the emerging antizyme polypeptide inhibits completion of its synthesis causing a ribosome pile-up on antizyme mRNA, whereas polyamine binding to nascent antizyme promotes completion of its synthesis. Thus, our study reveals a novel autoregulatory mechanism, in which binding of a small metabolite to a nascent sensor protein stimulates the latter's synthesis co-translationally

Polyamines directly promote antizyme-mediated degradation of ornithine decarboxylase by the proteasome

Ornithine decarboxylase (ODC), a ubiquitin-independent substrate of the proteasome, is a homodimeric protein with a rate-limiting function in polyamine biosynthesis. Polyamines regulate ODC levels by a feedback mechanism mediated by ODC antizyme (OAZ). Higher cellular polyamine levels trigger the synthesis of OAZ and also inhibit its ubiquitin-dependent proteasomal degradation. OAZ binds ODC monomers and targets them to the proteasome. Here, we report that polyamines, aside from their role in the control of OAZ synthesis and stability, directly enhance OAZ-mediated ODC degradation by the proteasome. Using a stable mutant of OAZ, we show that polyamines promote ODC degradation in Saccharomyces cerevisiae cells even when OAZ levels are not changed. Furthermore, polyamines stimulated the in vitro degradation of ODC by the proteasome in a reconstituted system using purified components. In these assays, spermine shows a greater effect than spermidine. By contrast, polyamines do not have any stimulatory effect on the degradation of ubiquitin-dependent substrates