Present-day HTA is broad and has numerous stakeholders, with none so important as the patient. By asking patient-oriented questions in HTA and better involving patients throughout the entire process, we can easily promote patient empowerment, and as such make patients more capable to play a more active role in healthcare decision making.

Mitochondrial tRNAs (mtRNAs) often lack domains and posttranscriptional modifications that are found in cytoplasmic tRNAs. These structural and chemical elements normally stabilize the folding of cytoplasmic tRNAs into canonical structures that are competent for aminoacylation and translation. For example, the dihydrouridine (D) stem and loop domain is involved in the tertiary structure of cytoplasmic tRNAs through hydrogen bonds and a Mg 2+ bridge to the ribothymidine (T) stem and loop domain. These interactions are often absent in mtRNA because the D-domain is truncated or missing. Using gel mobility shift analyses, UV, circular dichroism and NMR spectroscopies and aminoacylation assays, we have investigated the functional folding interactions of chemically synthesized and site-specifically modified mitochondrial and cytoplasmic tRNAs. We found that Mg 2+ is critical for folding of the truncated D-domain of bovine mtRNA Met with the tRNA's T-domain. Contrary to the expectation that Mg 2+ stabilizes RNA folding, the mtRNA Met Ddomain structure was unfolded and relaxed, rather than stabilized in the presence of Mg 2+ . Because the D-domain is transcribed prior to the T-domain, we conclude that Mg 2+ prevents misfolding of the 5′-half of bovine mtRNA Met facilitating its correct interaction with the T-domain. The interaction of the mtRNA Met D-domain with the T-domain was enhanced by a pseudouridine located in either the D-or T-domains compared to that of the unmodified RNAs (K d = 25.3, 24.6 and 44.4 μM, respectively). Mg 2+ also affected the folding interaction of a yeast mtRNA Leu1 , but had minimal effect on the folding of an E. coli cytoplasmic tRNA Leu . The D-domain modification, dihydrouridine, facilitated mtRNA Leu folding. These data indicate that conserved modifications assist and stabilize the formation of the functional mtRNA tertiary structure.

Pacman/Xrn1 is a highly conserved exoribonuclease known to play a critical role in gene regulatory events such as control of mRNA stability, RNA interference and regulation via miRNAs. Although Pacman has been well studied in Drosophila tissue culture cells, the biologically relevant cellular pathways controlled by Pacman in natural tissues are unknown. This study shows that a hypomorphic mutation in pacman (pcm5) results in smaller wing imaginal discs. These tissues, found in the larva, are known to grow and differentiate to form wing and thorax structures in the adult fly. Using microarray analysis, followed by quantitative RT-PCR, we show that eight mRNAs were increased in level by > 2-fold in the pcm5 mutant wing discs compared with the control. The levels of pre-mRNAs were tested for five of these mRNAs; four did not increase in the pcm5 mutant, showing that they are regulated at the post-transcriptional level and, therefore, could be directly affected by Pacman. These transcripts include one that encodes the heat shock protein Hsp67Bc, which is upregulated 11.9-fold at the post-transcriptional level and 2.3-fold at the protein level. One miRNA, miR-277-3p, is 5.6-fold downregulated at the post-transcriptional level in mutant discs, suggesting that Pacman affects its processing in this tissue. Together, these data show that a relatively small number of mRNAs and miRNAs substantially change in abundance in pacman mutant wing imaginal discs. Since Hsp67Bc is known to regulate autophagy and protein synthesis, it is possible that Pacman may control the growth of wing imaginal discs by regulating these processes.

BackgroundOne year of antipsychotic treatment from symptom remission is recommended following a first episode of psychosis (FEP).AimsTo investigate the effectiveness of commonly used antipsychotic medications in FEP.MethodA retrospective cohort study of naturalistic treatment of patients (N=460) accepted by FEP services across seven UK sites. Treatment initiation to all-cause discontinuation determined from case files.ResultsRisk of treatment discontinuation is greatest within 3 months of treatment initiation. Risperidone had longest median survival time. No significant differences were observed in time to discontinuation between commonly used antipsychotics on multivariable Cox regression analysis. Poor adherence and efficacy failure were the most common reasons for discontinuation.ConclusionsEffectiveness differences appear not to be a current reason for antipsychotic choice in FEP. Adherence strategies and weighing up likely adverse effects should be the clinical focus.Declaration of interestR.W., A.T. and S.M. have received research grant, speaker honoraria and conference attendance funding from all companies marketing antipsychotics.Copyright and usage© The Royal College of Psychiatrists 2016. This is an open access article distributed under the terms of the Creative Commons Non-Commercial, No Derivatives (CC BY-NC-ND) license.

The mitochondrial tRNA genes are hot spots for mutations that lead to human disease. A single point mutation (T4409C) in the gene for human mitochondrial tRNA Met (hmtRNA Met ) has been found to cause mitochondrial myopathy. This mutation results in the replacement of U8 in hmtRNA Met with a C8. The hmtRNA Met serves both in translational initiation and elongation in human mitochondria making this tRNA of particular interest in mitochondrial protein synthesis. Here we show that the single 8U3 C mutation leads to a failure of the tRNA to respond conformationally to Mg 2؉ . This mutation results in a drastic disruption of the structure of the hmtRNA Met , which significantly reduces its aminoacylation. The small fraction of hmtRNA Met that can be aminoacylated is not formylated by the mitochondrial Met-tRNA transformylase preventing its function in initiation, and it is unable to form a stable ternary complex with elongation factor EF-Tu preventing any participation in chain elongation. We have used structural probing and molecular reconstitution experiments to examine the structures formed by the normal and mutated tRNAs. In the presence of Mg 2؉ , the normal tRNA displays the structural features expected of a tRNA. However, even in the presence of Mg 2؉ , the mutated tRNA does not form the cloverleaf structure typical of tRNAs. Thus, we believe that this mutation has disrupted a critical Mg 2؉ -binding site on the tRNA required for formation of the biologically active structure. This work establishes a foundation for understanding the physiological consequences of the numerous mitochondrial tRNA mutations that result in disease in humans.Human mitochondria are subcellular organelles that produce more than 90% of the energy required by the cell. The mitochondrial genome encodes 13 proteins necessary for energy production, two rRNAs and all of the 22 tRNAs required for the synthesis of these proteins (1, 2). Mammalian mitochondrial tRNAs have several unusual features that distinguish them from canonical tRNAs. In many cases, they lack a number of the conserved or semi-conserved nucleotides that play important roles in creating the L-shaped tertiary structure of prokaryotic and eukaryotic cytoplasmic tRNAs (3). There is little detailed structural information on these tRNAs. No data are currently available that examine the structure of mammalian mitochondrial tRNAs with single nucleotide resolution. However, chemical and enzymatic probing has lead to the idea that these tRNAs have retained the basic cloverleaf structure of canonical tRNAs but that they lack several conserved tertiary interactions leading to a weaker three-dimensional structure (4 -8). In particular, a number of the long range interactions between the Dand T-arms of the tRNAs appear to be missing.All 22 tRNAs that function in mammalian mitochondria are encoded in the mitochondrial DNA. Considerable interest in mitochondrial tRNAs centers on the occurrence of diseases arising from mutations in their genes that lead to maternally inherited genetic diso...

Background - Clinical factors associated with development of intravascular lead adherence (ILA) are unreliable predictors. Because vascular injury in the superior vena cava - right atrium (SVC-RA) during transvenous lead extraction (TLE) is more likely to occur in segments with higher degrees of ILA, reliable and accurate assessment of ILA is warranted. We hypothesized that intravascular ultrasound (IVUS) could accurately visualize and quantify ILA and degree of ILA correlates with TLE difficulty. Methods - Serial imaging of leads occurred prior to TLE using IVUS. ILA areas were classified as high or low grade. Degree of extraction difficulty was assessed using 2 metrics and correlated with ILA grade. Lead extraction difficulty (LED) was calculated for each patient and compared to IVUS findings. Results - 158 vascular segments in 60 patients were analyzed: 141 (89%) low grade versus 17 (11%) high grade. Median extraction time (low=0 versus high grade=97 seconds, p<0.001) and median laser pulsations delivered (low=0 versus high grade=5852, p<0.001) were significantly higher in high grade segments. Most patients with low LED score had low ILA grades. 86% of patients with high LED score had low IVUS grade and the degree of TLE difficulty was similar to patients with low IVUS grades and LED scores. Conclusions - IVUS is a feasible imaging modality that may be useful in characterizing ILA in the SVC-RA region. An ILA grading system using imaging correlates with extraction difficulty. Most patients with clinical factors associated with higher extraction difficulty may exhibit lower ILA and extraction difficulty based on IVUS imaging.

RNA degradation is a vital post-transcriptional process which ensures that transcripts are maintained at the correct level within the cell. DIS3L2 and XRN1 are conserved exoribonucleases which are critical for the degradation of cytoplasmic RNAs. Although the molecular mechanisms of RNA degradation by DIS3L2 and XRN1 have been well studied, less is known about their specific roles in development of multicellular organisms or human disease. This review focusses on the roles of DIS3L2 and XRN1 in the pathogenesis of human disease, particularly in relation to phenotypes seen in model organisms. The known diseases associated with loss of activity of DIS3L2 and XRN1 are discussed, together with possible mechanisms and cellular pathways leading to these disease conditions.

IntroductionOur project follows community requests for health service incorporation into conservation collaborations in the rainforests of Papua New Guinea (PNG). This protocol is for health needs assessments, our first step in coplanning medical provision in communities with no existing health data.Methods and analysisThe study includes clinical assessments and rapid anthropological assessment procedures (RAP) exploring the health needs and perspectives of partner communities in two areas, conducted over 6 weeks fieldwork. First, in Wanang village (population c.200), which is set in lowland rainforest. Second, in six communities (population c.3000) along an altitudinal transect up the highest mountain in PNG, Mount Wilhelm. Individual primary care assessments incorporate physical examinations and questioning (providing qualitative and quantitative data) while RAP includes focus groups, interviews and field observations (providing qualitative data). Given absence of in-community primary care, treatments are offered alongside research activity but will not form part of the study. Data are collected by a research fellow, primary care clinician and two PNG research technicians. After quantitative and qualitative analyses, we will report: ethnoclassifications of disease, causes, symptoms and perceived appropriate treatment; community rankings of disease importance and service needs; attitudes regarding health service provision; disease burdens and associations with altitudinal-related variables and cultural practices. To aid wider use study tools are in online supplemental file, and paper and ODK versions are available free from the corresponding author.Ethics and disseminationChallenges include supporting informed consent in communities with low literacy and diverse cultures, moral duties to provide treatment alongside research in medically underserved areas while minimising risks of therapeutic misconception and inappropriate inducement, and PNG research capacity building. Brighton and Sussex Medical School (UK), PNG Institute of Medical Research and PNG Medical Research Advisory Committee have approved the study. Dissemination will be via journals, village meetings and plain language summaries.