A new procedure to analyze RNA non-branching structures

RNA structure prediction and structural motifs analysis are challenging tasks in the investigation of RNA function. We propose a novel procedure to detect structural motifs shared between two RNAs (a reference and a target). In particular, we developed two core modules: (i) nbRSSP_extractor, to assign a unique structure to the reference RNA encoded by a set of non-branching structures; (ii) SSD_finder, to detect structural motifs that the target RNA shares with the reference, by means of a new score function that rewards the relative distance of the target non-branching structures compared to the reference ones. We integrated these algorithms with already existing software to reach a coherent pipeline able to perform the following two main tasks: prediction of RNA structures (integration of RNALfold and nbRSSP_extractor) and search for chains of matches (integration of Structator and SSD_finder)

Spontaneous Embedding of DNA Mismatches Within the RNA:DNA Hybrid of CRISPR-Cas9.

CRISPR-Cas9 is the forefront technology for editing the genome. In this system, the Cas9 protein is programmed with guide RNAs to process DNA sequences that match the guide RNA forming an RNA:DNA hybrid structure. However, the binding of DNA sequences that do not fully match the guide RNA can limit the applicability of CRISPR-Cas9 for genome editing, resulting in the so-called off-target effects. Here, molecular dynamics is used to probe the effect of DNA base pair mismatches within the RNA:DNA hybrid in CRISPR-Cas9. Molecular simulations revealed that the presence of mismatched pairs in the DNA at distal sites with respect to the Protospacer Adjacent Motif (PAM) recognition sequence induces an extended opening of the RNA:DNA hybrid, leading to novel interactions established by the unwound nucleic acids and the protein counterpart. On the contrary, mismatched pairs upstream of the RNA:DNA hybrid are rapidly incorporated within the heteroduplex, with minor effect on the protein-nucleic acid interactions. As a result, mismatched pairs at PAM distal ends interfere with the activation of the catalytic HNH domain, while mismatches fully embedded in the RNA:DNA do not affect the HNH dynamics and enable its activation to cleave the DNA. These findings provide a mechanistic understanding to the intriguing experimental evidence that PAM distal mismatches hamper a proper function of HNH, explaining also why mismatches within the heteroduplex are much more tolerated. This constitutes a step forward in understanding off-target effects in CRISPR-Cas9, which encourages novel structure-based engineering efforts aimed at preventing the onset of off-target effects

Change of cystine/glutamate antiporter expression in ethanol-dependent rats

Background: Some drugs of abuse down regulate the expression of cystine/glutamate (xCT) antiporter in the nucleus accumbens (Acb) after extinction or withdrawal. The altered level of xCT exchanger in Acb, a structure involved in ethanol reinforcement, may contribute to the pathological glutamatergic signalling, linked to addiction. We hypothesised that the expression of xCT may be changed in Acb and whole brain also in non-dependent (occasional drinkers), ethanol-dependent rats, as well as, during ethanol withdrawal.Methods: Wistar rats were made ethanol-dependent by chronic exposure to an alcoholic milk beverage (from 2.4 to 7.2% v/v ethanol). Ethanol non-dependent rats were exposed to a similar, but non-alcoholic liquid diet and self-administered ethanol (10%) twice a week. Withdrawal in ethanol-dependent rats was studied at 12 hours after the last ethanol-enriched diet exposure. Immediately after the measurement of somatic signs of withdrawal, Western blot analysis with a polyclonal antibody against xCT was carried out in a naïve control group, non-dependent and ethanol-dependent rats as well as withdrawal rats, in order to study the level of xCT expression in Acb and whole brain. Results. Non-dependent rats self-administered an average dose of 1.21±0.02 g/kg per session (30 min). Daily ethanol consumption during chronic exposure to the alcoholic beverage ranged from 6.30±0.16 to 13.99±0.66 g/kg. Ethanol dependent rats after suspension of the ethanol-enriched diet have shown significant somatic signs of withdrawal. Western blotting analysis of Acb lysates revealed that xCT was over expressed in ethanol-dependent rats whereas in whole brain preparations xCT was over expressed in both non-dependent and ethanol-dependent rats compared to control group. On the contrary, xCT expression during withdrawal was down regulated in Acb and restored to control level in whole brain preparations. Conclusions: The changes of xCT expression in both Acb and whole brain followi

Pharmacokinetics and antinociceptive effects of tramadol and its metabolite O-desmethyltramadol following intravenous administration in sheep

Although sheep are widely used as an experimental model for various surgical procedures there is a paucity of data on the pharmacokinetics and efficacy of analgesic drugs in this species. The aims of this study were to investigate the pharmacokinetics of intravenously (IV) administered tramadol and its active metabolite O-desmethyltramadol (M1) and to assess the mechanical antinociceptive effects in sheep. In a prospective, randomized, blinded study, six healthy adult sheep were given 4 and 6\u2009mg/kg tramadol and saline IV in a cross-over design with a 2-week wash-out period. At predetermined time points blood samples were collected and physiological parameters and mechanical nociceptive threshold (MNT) values were recorded. The analytical determination of tramadol and M1 was performed using high performance liquid chromatography. Pharmacokinetic parameters fitted a two- and a non-compartmental model for tramadol and M1, respectively. Normally distributed data were analysed by a repeated mixed linear model. Plasma concentration vs. time profiles of tramadol and M1 were similar after the two doses. Tramadol and M1 plasma levels decreased rapidly in the systemic circulation, with both undetectable after 6\u2009h following drug administration. Physiological parameters did not differ between groups; MNT values were not statistically significant between groups at any time point. It was concluded that although tramadol and M1 concentrations in plasma were above the human minimum analgesic concentration after both treatments, no mechanical antinociceptive effects of tramadol were reported. Further studies are warranted to assess the analgesic efficacy of tramadol in sheep

Scalar one-loop integrals for QCD

We construct a basis set of infra-red and/or collinearly divergent scalar one-loop integrals and give analytic formulas, for tadpole, bubble, triangle and box integrals, regulating the divergences (ultra-violet, infra-red or collinear) by regularization in $D=4-2\epsilon$ dimensions. For scalar triangle integrals we give results for our basis set containing 6 divergent integrals. For scalar box integrals we give results for our basis set containing 16 divergent integrals. We provide analytic results for the 5 divergent box integrals in the basis set which are missing in the literature. Building on the work of van Oldenborgh, a general, publicly available code has been constructed, which calculates both finite and divergent one-loop integrals. The code returns the coefficients of $1/\epsilon^2,1/\epsilon^1$ and $1/\epsilon^0$ as complex numbers for an arbitrary tadpole, bubble, triangle or box integral.Comment: 27 pages, 5 figures, associated fortran code available at http://qcdloop.fnal.gov/. New version corrects typographical error in Eq. 5.

Acetaldehyde-Reinforcing Effects: A Study on Oral Self-Administration Behavior

Acetaldehyde (ACD) is the first metabolite of ethanol. Although, the role of ACD in ethanol addiction has been controversial, there are data showing a relationship. The objective of the current study was to further test the hypothesis that ACD itself is reinforcing. For this reason, we carried out a study on operant oral ACD self-administration. Wistar rats were trained to self-administer tap water or ACD by nose-poking in daily 30 min sessions for 15 consecutive days. Response on active nose-poke caused delivery of ACD solution or tap water, whereas responses on inactive nose-poke had no consequences. The results show that ACD maintains oral self-administration behavior and rates of active nose-pokes significantly higher than tap water. The dose–response plot for oral ACD self-administration is a “bell-shaped” curve suggesting reinforcing properties only in a limited range of doses. Furthermore, rats self-administering ACD show a deprivation effect upon ACD removal and gradually reinstated active nose-poke response when ACD was reintroduced. Overall, this study shows that ACD is orally self-administered and further supports the hypothesis that ACD possesses reinforcing properties, which suggests that some of the pharmacological effects attributed to ethanol may result from its biotransformation into ACD, thereby supporting an active involvement of ACD in ethanol addiction

Can morphological features of coccolithophores serve as a reliable proxy to reconstruct environmental conditions of the past?

Morphological changes in coccoliths, tiny calcite platelets covering the outer surface of coccolithophores, can be induced by physiological responses to environmental changes. Coccoliths recovered from sedimentary successions may therefore provide information on paleo-environmental conditions prevailing at the time when the coccolithophores were alive. To calibrate the biomineralization responses of ancient coccolithophore to environmental changes, studies often compared the biological responses of living coccolithophore species with paleo-data from calcareous nannofossils. However, there is uncertainty whether the morphological responses of living coccolithophores are representative of those of the fossilized ancestors. To investigate this, we exposed four living coccolithophore species (Emiliania huxleyi, Gephyrocapsa oceanica, Coccolithus pelagicus subsp. braarudii, and Pleurochrysis carterae) that have been evolutionarily distinct for hundreds of thousands to millions of years, to a range of environmental conditions (i.e., changing light intensity, Mg∕Ca ratio, nutrient availability, temperature, and carbonate chemistry) and evaluated their responses in coccolith morphology (i.e., size, length, width, malformation). The motivation for this study was to test if there is a consistent morphological response of the four species to changes in any of the tested abiotic environmental factors. If this was the case, then this could suggest that coccolith morphology can serve as a paleo-proxy for that specific factor because this response is conserved across species that have been evolutionary distinct over geological timescales. However, we found that the four species responded differently to changing light intensity, Mg∕Ca ratio, nutrient availability, and temperature in terms of coccolith morphology. The lack of a common response reveals the difficulties in using coccolith morphology as a paleo-proxy for these environmental drivers. However, a common response was observed under changing seawater carbonate chemistry (i.e., rising CO2), which consistently induced malformations. This commonality provides some confidence that malformations found in the sedimentary record could be indicative of adverse carbonate chemistry conditions

Squeezing out predictions with leptogenesis from SO(10)

We consider the see-saw mechanism within a non-supersymmetric SO(10) model. By assuming the SO(10) quark-lepton symmetry, and after imposing suitable conditions that ensure that the right-handed (RH) neutrino masses are at most mildly hierarchical (compact RH spectrum) we obtain a surprisingly predictive scenario. The absolute neutrino mass scale, the Dirac and the two Majorana phases of the neutrino mixing matrix remain determined in terms of the set of already measured low energy observables, modulo a discrete ambiguity in the signs of two neutrino mixing angles and of the Dirac phase. The RH neutrinos mass spectrum is also predicted, as well as the size and sign of the leptogenesis CP asymmetries. We compute the cosmological baryon asymmetry generated through leptogenesis and obtain the correct sign and a size compatible with observations.Comment: 18 pages, 2 figures; minor changes, version accepted for publication in PR