Miracle-dz/UniFAR-FTUs huggingface dataset:
The Reduction Potential of the Active Site Disulfides of Human Protein Disulfide Isomerase Limits Oxidation of the Enzyme by Ero1α*

Disulfide formation in newly synthesized proteins entering the mammalian endoplasmic reticulum is catalyzed by protein disulfide isomerase (PDI), which is itself thought to be directly oxidized by Ero1α. The activity of Ero1α is tightly regulated by the formation of noncatalytic disulfides, which need to be broken to activate the enzyme. Here, we have developed a novel PDI oxidation assay, which is able to simultaneously determine the redox status of the individual active sites of PDI. We have used this assay to confirm that when PDI is incubated with Ero1α, only one of the active sites of PDI becomes directly oxidized with a slow turnover rate. In contrast, a deregulated mutant of Ero1α was able to oxidize both PDI active sites at an equivalent rate to the wild type enzyme. When the active sites of PDI were mutated to decrease their reduction potential, both were now oxidized by wild type Ero1α with a 12-fold increase in activity. These results demonstrate that the specificity of Ero1α toward the active sites of PDI requires the presence of the regulatory disulfides. In addition, the rate of PDI oxidation is limited by the reduction potential of the PDI active site disulfide. The inability of Ero1α to oxidize PDI efficiently likely reflects the requirement for PDI to act as both an oxidase and an isomerase during the formation of native disulfides in proteins entering the secretory pathway.
ERK1/2 Phosphorylate Raptor to Promote Ras-dependent Activation of mTOR Complex 1 (mTORC1)*

The Ras/mitogen-activated protein kinase (MAPK) pathway regulates a variety of cellular processes by activating specific transcriptional and translational programs. Ras/MAPK signaling promotes mRNA translation and protein synthesis, but the exact molecular mechanisms underlying this regulation remain poorly understood. Increasing evidence suggests that the mammalian target of rapamycin (mTOR) plays an essential role in this process. Here, we show that Raptor, an essential scaffolding protein of the mTOR complex 1 (mTORC1), becomes phosphorylated on proline-directed sites following activation of the Ras/MAPK pathway. We found that ERK1 and ERK2 interact with Raptor in cells and mediate its phosphorylation in vivo and in vitro. Using mass spectrometry and phosphospecific antibodies, we found three proline-directed residues within Raptor, Ser8, Ser696, and Ser863, which are directly phosphorylated by ERK1/2. Expression of phosphorylation-deficient alleles of Raptor revealed that phosphorylation of these sites by ERK1/2 normally promotes mTORC1 activity and signaling to downstream substrates, such as 4E-BP1. Our data provide a novel regulatory mechanism by which mitogenic and oncogenic activation of the Ras/MAPK pathway promotes mTOR signaling.
Revisiting Iodination Sites in Thyroglobulin with an Organ-oriented Shotgun Strategy

Thyroglobulin (Tg) is secreted by thyroid epithelial cells. It is essential for thyroid hormonogenesis and iodine storage. Although studied for many years, only indirect and partial surveys of its post-translational modifications were reported. Here, we present a direct proteomic approach, used to study the degree of iodination of mouse Tg without any preliminary purification. A comprehensive coverage of Tg was obtained using a combination of different proteases, MS/MS fragmentation procedures with inclusion lists and a hybrid mass high-resolution LTQ-Orbitrap XL mass spectrometer. Although only 16 iodinated sites are currently known for human Tg, we uncovered 37 iodinated tyrosine residues, most of them being mono- or diiodinated. We report the specific isotopic pattern of thyroxine modification, not recognized as a normal peptide pattern. Four hormonogenic sites were detected. Two donor sites were identified through the detection of a pyruvic acid residue in place of the initial tyrosine. Evidence for polypeptide cleavages sites due to the action of cathepsins and dipeptidyl proteases in the thyroid were also detected. This work shows that semi-quantitation of Tg iodination states is feasible for human biopsies and should be of significant medical interest for further characterization of human thyroid pathologies.
Mitochondrial Dysfunction and Oxidative Stress Contribute to the Pathogenesis of Spinocerebellar Ataxia Type 12 (SCA12)*

Spinal cerebellar ataxia type 12 (SCA12) has been attributed to the elevated expression of ppp2r2b. To better elucidate the pathomechanism of the neuronal disorder and to search for a pharmacological treatment, Drosophila models of SCA12 were generated by overexpression of a human ppp2r2b and its Drosophila homolog tws. Ectopic expression of ppp2r2b or tws caused various pathological features, including neurodegeneration, apoptosis, and shortened life span. More detailed analysis revealed that elevated ppp2r2b and tws induced fission of mitochondria accompanied by increases in cytosolic reactive oxygen species (ROS), cytochrome c, and caspase 3 activity. Transmission electron microscopy revealed that fragmented mitochondria with disrupted cristae were engulfed by autophagosomes in photoreceptor neurons of flies overexpressing tws. Additionally, transgenic flies were more susceptible to oxidative injury induced by paraquat. By contrast, ectopic Drosophila Sod2 expression and antioxidant treatment reduced ROS and caspase 3 activity and extended the life span of the SCA12 fly model. In summary, our study demonstrates that oxidative stress induced by mitochondrial dysfunction plays a causal role in SCA12, and reduction of ROS is a potential therapeutic intervention for this neuropathy.
Constituting corporate Europe: a study of elite social organization

This article explores the emerging shape and form of the European corporate community since 1996. We examine the cohesion of corporate Europe through the network of interlocking corporate directorates and memberships in the European Round Table of Industrialists. We focus on the unequal structure of representation; the interplay of national and transnational aspects of the network; the role of finance capitalists as a signpost of a regime of internationalized finance capital; and the embeddedness of corporate Europe in the global corporate network. Although the transnational European network gained in strength while national networks eroded, expansion of the European network did not negate a structure of representation favoring the northwest. Bankers became less dominant, yet industrialists with financial connections formed the core of the European corporate community, signaling a departure from national corporate communities centered upon banks. At the threshold of the current economic crisis, corporate Europe comprised the most integrated segment of the global corporate elite.
Reconstitution of Proapoptotic BAK Function in Liposomes Reveals a Dual Role for Mitochondrial Lipids in the BAK-driven Membrane Permeabilization Process*

BAK is a key effector of mitochondrial outer membrane permeabilization (MOMP) whose molecular mechanism of action remains to be fully dissected in intact cells, mainly due to the inherent complexity of the intracellular apoptotic machinery. Here we show that the core features of the BAK-driven MOMP pathway can be reproduced in a highly simplified in vitro system consisting of recombinant human BAK lacking the carboxyl-terminal 21 residues (BAKΔC) and tBID in combination with liposomes bearing an appropriate lipid environment. Using this minimalist reconstituted system we established that tBID suffices to trigger BAKΔC membrane insertion, oligomerization, and pore formation. Furthermore, we demonstrate that tBID-activated BAKΔC permeabilizes the membrane by forming structurally dynamic pores rather than a large proteinaceous channel of fixed size. We also identified two distinct roles played by mitochondrial lipids along the molecular pathway of BAKΔC-induced membrane permeabilization. First, using several independent approaches, we showed that cardiolipin directly interacts with BAKΔC, leading to a localized structural rearrangement in the protein that “primes” BAKΔC for interaction with tBID. Second, we provide evidence that selected curvature-inducing lipids present in mitochondrial membranes specifically modulate the energetic expenditure required to create the BAKΔC pore. Collectively, our results support the notion that BAK functions as a direct effector of MOMP akin to BAX and also adds significantly to the growing evidence indicating that mitochondrial membrane lipids are actively implicated in BCL-2 protein family function.
Optimizing BeLPT criteria for beryllium sensitization.

BACKGROUND The beryllium lymphocyte proliferation test (BeLPT) is used to identify persons sensitized to beryllium. ATSDR convened an expert panel of physicians and scientists in April 2006 to discuss this test and to consider what BeLPT test results actually establish beryllium sensitization. The three criteria proposed by panel members were (1)one abnormal result, (2)one abnormal and one borderline result, and (3)two abnormal results. METHODS Complete algorithms were developed for each of the three proposed criteria. Using single-test outcome probabilities developed by Stange et al. [2004. Am J Ind Med 46:453-462], we calculated and compared the sensitivity, specificity, and positive predictive values (PPVs) for each set of criteria. RESULTS The overall sensitivity and specificity of the three criteria were similar. When the criteria required confirmation of an abnormal result the PPV was higher--whether the requirement was satisfied by a borderline result, or only by another abnormal result. Confirmation also reduced the likelihood of false positives. The differences between the three criteria decreased as the prevalence of sensitization increased. CONCLUSIONS A single unconfirmed abnormal is usually insufficient to establish sensitization for an apparently healthy person. When the prevalence of beryllium sensitization in a group is high, however, even a single abnormal BeLPT can be a strong predictor.
Structural Basis of Carbohydrate Recognition by Calreticulin*

The calnexin cycle is a process by which glycosylated proteins are subjected to folding cycles in the endoplasmic reticulum lumen via binding to the membrane protein calnexin (CNX) or to its soluble homolog calreticulin (CRT). CNX and CRT specifically recognize monoglucosylated Glc1Man9GlcNAc2 glycans, but the structural determinants underlying this specificity are unknown. Here, we report a 1.95-Å crystal structure of the CRT lectin domain in complex with the tetrasaccharide α-Glc-(1→3)-α-Man-(1→2)-α-Man-(1→2)-Man. The tetrasaccharide binds to a long channel on CRT formed by a concave β-sheet. All four sugar moieties are engaged in the protein binding via an extensive network of hydrogen bonds and hydrophobic contacts. The structure explains the requirement for glucose at the nonreducing end of the carbohydrate; the oxygen O2 of glucose perfectly fits to a pocket formed by CRT side chains while forming direct hydrogen bonds with the carbonyl of Gly124 and the side chain of Lys111. The structure also explains a requirement for the Cys105–Cys137 disulfide bond in CRT/CNX for efficient carbohydrate binding. The Cys105–Cys137 disulfide bond is involved in intimate contacts with the third and fourth sugar moieties of the Glc1Man3 tetrasaccharide. Finally, the structure rationalizes previous mutagenesis of CRT and lays a structural groundwork for future studies of the role of CNX/CRT in diverse biological pathways.
Epidemiology of back pain in older adults: prevalence and risk factors for back pain onset.

OBJECTIVES To determine the prevalence of disabling and non-disabling back pain across age in older adults, and identify risk factors for back pain onset in this age group. METHODS Participants aged ≥ 75 years answered interviewer-administered questions on back pain as part of a prospective cohort study [Cambridge City over-75s Cohort Study (CC75C)]. Descriptive analyses of data from two surveys, 1988-89 and 1992-93, estimated prevalence and new onset of back pain. Relative risks (RRs) and 95% CIs were estimated using Poisson regression, adjusted for age and gender. RESULTS Prevalence of disabling and non-disabling back pain was 6 and 23%, respectively. While prevalence of non-disabling back pain did not vary significantly across age (χ²trend : 0.90; P = 0.34), the prevalence of disabling back pain increased with age (χ²trend : 4.02; P = 0.04). New-onset disabling and non-disabling back pain at follow-up was 15 and 5%, respectively. Risk factors found to predict back pain onset at follow-up were: poor self-rated health (RR 3.8; 95% CI 1.8, 8.0); depressive symptoms (RR 2.2; 95% CI 1.3, 3.7); use of health or social services (RR 1.7; 95% CI 1.1, 2.7); and previous back pain (RR 2.1; 95% CI 1.2-3.5). From these, poor self-rated health, previous back pain and depressive symptoms were found to be independent predictors of pain onset. Markers of social networks were not associated with the reporting of back pain onset. Conclusion. The risk of disabling back pain rises in older age. Older adults with poor self-rated health, depressive symptoms, increased use of health and social services and a previous episode of back pain are at greater risk of reporting future back pain onset.
Sequence-dependent Prion Protein Misfolding and Neurotoxicity*

Prion diseases are neurodegenerative disorders caused by misfolding of the normal prion protein (PrP) into a pathogenic “scrapie” conformation. To better understand the cellular and molecular mechanisms that govern the conformational changes (conversion) of PrP, we compared the dynamics of PrP from mammals susceptible (hamster and mouse) and resistant (rabbit) to prion diseases in transgenic flies. We recently showed that hamster PrP induces spongiform degeneration and accumulates into highly aggregated, scrapie-like conformers in transgenic flies. We show now that rabbit PrP does not induce spongiform degeneration and does not convert into scrapie-like conformers. Surprisingly, mouse PrP induces weak neurodegeneration and accumulates small amounts of scrapie-like conformers. Thus, the expression of three highly conserved mammalian prion proteins in transgenic flies uncovered prominent differences in their conformational dynamics. How these properties are encoded in the amino acid sequence remains to be elucidated.
Aurora B Interacts with NIR-p53, Leading to p53 Phosphorylation in Its DNA-binding Domain and Subsequent Functional Suppression*

NIR (novel INHAT repressor) is a transcriptional co-repressor with inhibitor of histone acetyltransferase (INHAT) activity and has previously been shown to physically interact with and suppress p53 transcriptional activity and function. However, the mechanism by which NIR suppresses p53 is not completely understood. Using a proteomic approach, we have identified the Aurora kinase B as a novel binding partner of NIR. We show that Aurora B, NIR and p53 exist in a protein complex in which Aurora B binds to NIR, thus also indirectly associates with p53. Functionally, overexpression of Aurora B or NIR suppresses p53 transcriptional activity, and depletion of Aurora B or NIR causes p53-dependent apoptosis and cell growth arrest, due to the up-regulation of p21 and Bax. We then demonstrate that Aurora B phosphorylates multiple sites in the p53 DNA-binding domain in vitro, and this phosphorylation probably also occurs in cells. Importantly, the Aurora B-mediated phosphorylation on Ser269 or Thr284 significantly compromises p53 transcriptional activity. Taken together, these results provide novel insight into NIR-mediated p53 suppression and also suggest an additional way for p53 regulation.
Risks associated with tendinitis: effects from demographic, socioeconomic, and psychological status among Brazilian workers.

BACKGROUND Self-reported tendinitis/tenosynovitis was evaluated by gender, age group, skin color, family income, and educational and psychological status. METHODS ! The study was carried out in a representative sample of formally contracted Brazilian workers from a household survey. A total of 54,660 participants were included. Occupations were stratified according to estimated prevalences of self-reported injuries. Non-conditional logistic regression was performed, and all variables were analyzed in two occupational groups. RESULTS The overall prevalence rate of tendinitis/tenosynovitis was 3.1%: 5.5% in high-prevalence occupations (n = 10,726); and 2.5% in low-prevalence occupations (n = 43,934). White female workers between the ages of 45 and 64 years and at a higher socioeconomic level were more likely to report tendinitis/tenosynovitis regardless of their occupational category. An adjusted OR = 3.59 [95% CI: 3.15--4.09] was found between tendinitis/tenosynovitis and psychological status. CONCLUSION Among formally contracted Brazilian workers, higher income can imply greater physical and psychological demands that, regardless of occupational stratum, increase the risk of tendinitis/tenosynovitis.
TMEM16A Inhibitors Reveal TMEM16A as a Minor Component of Calcium-activated Chloride Channel Conductance in Airway and Intestinal Epithelial Cells*

TMEM16A (ANO1) functions as a calcium-activated chloride channel (CaCC). We developed pharmacological tools to investigate the contribution of TMEM16A to CaCC conductance in human airway and intestinal epithelial cells. A screen of ∼110,000 compounds revealed four novel chemical classes of small molecule TMEM16A inhibitors that fully blocked TMEM16A chloride current with an IC50 < 10 μm, without interfering with calcium signaling. Following structure-activity analysis, the most potent inhibitor, an aminophenylthiazole (T16Ainh-A01), had an IC50 of ∼1 μm. Two distinct types of inhibitors were identified. Some compounds, such as tannic acid and the arylaminothiophene CaCCinh-A01, fully inhibited CaCC current in human bronchial and intestinal cells. Other compounds, including T16Ainh-A01 and digallic acid, inhibited total CaCC current in these cells poorly, but blocked mainly an initial, agonist-stimulated transient chloride current. TMEM16A RNAi knockdown also inhibited mainly the transient chloride current. In contrast to the airway and intestinal cells, all TMEM16A inhibitors fully blocked CaCC current in salivary gland cells. We conclude that TMEM16A carries nearly all CaCC current in salivary gland epithelium, but is a minor contributor to total CaCC current in airway and intestinal epithelia. The small molecule inhibitors identified here permit pharmacological dissection of TMEM16A/CaCC function and are potential development candidates for drug therapy of hypertension, pain, diarrhea, and excessive mucus production.
Postoperative Libido and Genital Sexual Sensitivity Following Various Forms of Hysterectomy

ABSTRACT The objective of the present study was to evaluate differences in libido and genital sexual sensitivity following established types of hysterectomy in cases of benign disease. These included the conventional vaginal (VH) and abdominal hysterectomy (AH), as well as the more modern, laparoscopy-assisted (LAVH) and pelviscopic (CASH) surgical techniques. The study was performed in a Vienna Hospital for Gynecology and Obstetrics in a prospective randomized manner. A total of 41 premenopausal, sexually active women were hysterectomized using one of the techniques cited and followed up for 1 year (3, 6, and 12 months). Measurement of sex hormones (E2, P, LH, and FSH), ovulation monitoring (basal temperature and vaginosonography), and the Kupperman score verified that the premenopasual status remained stable and constant throughout both the preoperative and postoperative periods. The quality of libido and genital sexual sensitivity was compared preoperatively and postoperatively within four assigned gro.
The Oligomeric State and Arrangement of the Active Bacterial Translocon*

Protein secretion in bacteria is driven through the ubiquitous SecYEG complex by the ATPase SecA. The structure of SecYEG alone or as a complex with SecA in detergent reveal a monomeric heterotrimer enclosing a central protein channel, yet in membranes it is dimeric. We have addressed the functional significance of the oligomeric status of SecYEG in protein translocation using single molecule and ensemble methods. The results show that while monomers are sufficient for the SecA- and ATP-dependent association of SecYEG with pre-protein, active transport requires SecYEG dimers arranged in the back-to-back conformation. Molecular modeling of this dimeric structure, in conjunction with the new functional data, provides a rationale for the presence of both active and passive copies of SecYEG in the functional translocon.
Treatment decisions and related costs differ significantly depending on the choice of a disease activity index in RA, according to 1987 and 2010 classification criteria.

OBJECTIVE To evaluate the therapeutic and economic consequences of various disease activity indices (DAIs) in RA according to 1987 and 2010 criteria. METHODS Data on disease activity states from all sustained visits were assessed from all patients who participate in the treatment in the Rotterdam Early Arthritis Cohort (tREACH) study, a stratified randomized trial to evaluate different treatment strategies in patients with a symptom duration of <1 year. Frequencies of treatment adaptations, based upon exclusive thresholds of various DAIs, were visualized in reclassification tables. The Stuart-Maxwell test was applied to analyse any significant differences between treatment decisions according to the different DAIs. Simulated annual median medication costs were estimated using the tREACH medication protocol with standard national costs. RESULTS DAIs perform similar in RA according to 1987 and 2010 criteria. A total of 1104 DASs per DAI were available from 296 patients. DAIs differ significantly, compared with DASs, in classifying a patient's disease state. Consequently, treatment intensifications occur more frequently with SDAI, CDAI and DAS-28 usage, compared with DAS. Tapering treatment occurs less frequently with SDAI and CDAI and more frequently with DAS-28 usage. Simulated annual median medication costs are significantly higher if DAS-28, SDAI and CDAI are used compared with DAS usage. CONCLUSION Usage of various DAIs in a single patient leads to inconsistent disease state categorizations. Consequently, these inconsistencies significantly influence therapeutic decisions and accompanying costs. As DAI usage is imperative to uphold current European League Against Rheumatology (EULAR) treatment recommendations, physicians should consider these therapeutic and economic consequences before choosing a particular DAI.
Use of O*NET as a job exposure matrix: A literature review.

BACKGROUND O*NET is a publicly available online database that describes occupational features across US job titles and that has been used to estimate workplace physical and psychosocial exposures and organizational characteristics. The aim of this review is to describe and evaluate the use of O*NET as a job exposure matrix. METHODS A review of the peer-reviewed published and gray literature was conducted. Twenty-eight studies were found that used O*NET to estimate work exposures related to health or safety outcomes. Each was systematically evaluated across eight main features. RESULTS Many health outcomes have been studied with O*NET estimates of job exposures. Some studies did not use conceptual definitions of exposure; few studies estimated convergent validity, most used predictive validity. Multilevel analysis was underutilized. CONCLUSION O*NET is worthy of exploration by the occupational health community, although its scientific value is still undetermined. More studies could eventually provide evidence of convergent validity. O*NET has the potential to allow examination of occupational risks that might have otherwise been ignored due to missing data or resource constraints on field data collection of job exposure information.