Partitioning between recoding and termination at a stop codon–selenocysteine insertion sequence

Selenocysteine (Sec) is inserted into proteins by recoding a UGA stop codon followed by a selenocysteine insertion sequence (SECIS). UGA recoding by the Sec machinery is believed to be very inefficient owing to RF2-mediated termination at UGA. Here we show that recoding efficiency in vivo is 30–40% independently of the cell growth rate. Efficient recoding requires sufficient selenium concentrations in the medium. RF2 is an unexpectedly poor competitor of Sec. We recapitulate the major characteristics of SECIS-dependent UGA recoding in vitro using a fragment of fdhF-mRNA encoding a natural bacterial selenoprotein. Only 40% of actively translating ribosomes that reach the UGA codon insert Sec, even in the absence of RF2, suggesting that the capacity to insert Sec into proteins is inherently limited. RF2 does not compete with the Sec incorporation machinery; rather, it terminates translation on those ribosomes that failed to incorporate Sec. The data suggest a model in which early recruitment of Sec-tRNA^Sec–SelB–GTP to the SECIS blocks the access of RF2 to the stop codon, thereby prioritizing recoding over termination at Sec-dedicated stop codons.     

Pre-fibrillar α-synuclein mutants cause Parkinson's disease-like non-motor symptoms in Drosophila

Parkinson's disease (PD) is linked to the formation of insoluble fibrillar aggregates of the presynaptic protein α-Synuclein (αS) in neurons. The appearance of such aggregates coincides with severe motor deficits in human patients. These deficits are often preceded by non-motor symptoms such as sleep-related problems in the patients. PD-like motor deficits can be recapitulated in model organisms such as Drosophila melanogaster when αS is pan-neurally expressed. Interestingly, both these deficits are more severe when αS mutants with reduced aggregation properties are expressed in flies. This indicates that that αS aggregation is not the primary cause of the PD-like motor symptoms. Here we describe a model for PD in Drosophila which utilizes the targeted expression of αS mutants in a subset of dopadecarboxylase expressing serotonergic and dopaminergic (DA) neurons. Our results show that targeted expression of pre-fibrillar αS mutants not only recapitulates PD-like motor symptoms but also the preceding non-motor symptoms such as an abnormal sleep-like behavior, altered locomotor activity and abnormal circadian periodicity. Further, the results suggest that the observed non-motor symptoms in flies are caused by an early impairment of neuronal functions rather than by the loss of neurons due to cell death.     

Brain signal predictions from multi-scale networks using a linearized framework

Simulations of neural activity at different levels of detail are ubiquitous in modern neurosciences, aiding the interpretation of experimental data and underlying neural mechanisms at the level of cells and circuits. Extracellular measurements of brain signals reflecting transmembrane currents throughout the neural tissue remain commonplace. The lower frequencies (≲ 300Hz) of measured signals generally stem from synaptic activity driven by recurrent interactions among neural populations and computational models should also incorporate accurate predictions of such signals. Due to limited computational resources, large-scale neuronal network models (≳ 10⁶ neurons or so) often require reducing the level of biophysical detail and account mainly for times of action potentials (‘spikes’) or spike rates. Corresponding extracellular signal predictions have thus poorly accounted for their biophysical origin.Here we propose a computational framework for predicting spatiotemporal filter kernels for such extracellular signals stemming from synaptic activity, accounting for the biophysics of neurons, populations, and recurrent connections. Signals are obtained by convolving population spike rates by appropriate kernels for each connection pathway and summing the contributions. Our main results are that kernels derived via linearized synapse and membrane dynamics, distributions of cells, conduction delay, and volume conductor model allow for accurately capturing the spatiotemporal dynamics of ground truth extracellular signals from conductance-based multicompartment neuron networks. One particular observation is that changes in the effective membrane time constants caused by persistent synapse activation must be accounted for.The work also constitutes a major advance in computational efficiency of accurate, biophysics-based signal predictions from large-scale spike and rate-based neuron network models drastically reducing signal prediction times compared to biophysically detailed network models. This work also provides insight into how experimentally recorded low-frequency extracellular signals of neuronal activity may be approximately linearly dependent on spiking activity. A new software tool LFPykernels serves as a reference implementation of the framework.     

Comparative analysis of metabolites in contrasting chickpea cultivars

Journal of Plant Biochemistry and Biotechnology - Chickpea (Cicer arietinum L.) is a good source of nutrients for animals and human consumption. In the present study, we analyzed the anthocyanin...     

Stereoselective synthesis of a ketohexofuranose from an aldohexopyranose by a [6+1-1] strategy

Ozonolysis of 2-acetoxymethyl-1,5-anhydro-3,4,6-tri-O-benzyl-2-deoxy-D-arabino-hex-1-enitol gave 1-O-acetyl-3,4,6-tri-O-benzyl-4-O-formyl-D-arabino-hex-2-ulose (5). Subsequent hydrolysis and acetylation of 5 provided 1,2-di-O-acetyl-3,4,6-tri-O-benzyl-D-fructofuranose 6 in excellent yield. This methodology allows specific deuteration at C-1 of a protected D-fructofuranose derivative. This approach therefore could serve as [6+1-1] formulation for hexose series inter-conversion, that is, aldohexopyranose to ketohexofuranose.     

Evidence for p62 aggregate formation: role in cell survival

The polyubiquitin-binding protein p62 has been shown to localize in aggregates common to several types of diseases. Here, we report that p62 forms independent fibrillar aggregates in vitro in a time- and concentration-dependent manner. FTIR spectra and ThT fluorescence assay of p62 reveals increased beta-sheet content as aggregates form compared to the native protein. The fibrillar nature of the aggregates was observed by transmission electron microscopy. Overexpression of p62 in HEK cells results in aggregate formation that may protect cells from apoptosis. Altogether, these results suggest that p62 fibrils may influence cell viability and indicates an important role for p62 in aggresome formation.     

Biosynthesis of dihydrochalcomycin: Characterization of a deoxyallosyltransferase (gerGTI)

Through an inactivation experiment followed by complementation, the gerGTII gene was previously characterized as a chalcosyltransferase gene involved in the biosynthesis of dihydochalcomycin. The glycosyltransferase gerGTI was identified as a deoxyallosyltransferase required for the glycosylation of D-mycinose sugar. This 6-deoxyhexose sugar was converted to mycinose, via bis-O-methylation, following attachment to the polyketide lactone during dihydrochalcomycin biosynthesis. Gene sequence alignment of gerGTI to several glycosyltransferases revealed a consensus sequence motif that appears to be characteristic of the enzymes in this sub-group of the glycosyltransferase family. To characterize its putative function, genetic disruption of gerGTI in the wild-type strain Streptomyces sp. KCTC 0041BP and in the gerGTII-deleted mutant (S. sp. ΔgerGTII), as well as complementation of gerGTII in S. sp. ΔgerGTII-GTI, were carried out, and the products were analyzed by LC/MS. S. sp. ΔgerGTII-GTI mutant produced dihydrochalconolide macrolide. S. sp. ΔgerGTI and S. sp. ΔgerGTII-GTI complementation of gerGTII yielded dihydrochalconolide without the mycinose sugar. The intermediate shows that gerGTI encodes a deoxyallosyltransferase that acts after gerGTII.