An algorithm and a computer program have been prepared for determining RNA secondary structures within any prescribed increment of the computed global minimum free energy. The mathematical problem of determining how well defined a minimum energy folding is can now be solved. All predicted base pairs that can participate in suboptimal structures may be displayed and analyzed graphically. Representative suboptimal foldings are generated by selecting these base pairs one at a time and computing the best foldings that contain them. A distance criterion that ensures that no two structures are "too close" is used to avoid multiple generation of similar structures. Thermodynamic parameters, including free-energy increments for single-base stacking at the ends of helices and for terminal mismatched pairs in interior and hairpin loops, are incorporated into the underlying folding model of the above algorithm.

BACKGROUND

The current combat operations in Iraq and Afghanistan have involved U.S. military personnel in major ground combat and hazardous security duty. Studies are needed to systematically assess the mental health of members of the armed services who have participated in these operations and to inform policy with regard to the optimal delivery of mental health care to returning veterans.

METHODS

We studied members of four U.S. combat infantry units (three Army units and one Marine Corps unit) using an anonymous survey that was administered to the subjects either before their deployment to Iraq (n=2530) or three to four months after their return from combat duty in Iraq or Afghanistan (n=3671). The outcomes included major depression, generalized anxiety, and post-traumatic stress disorder (PTSD), which were evaluated on the basis of standardized, self-administered screening instruments.

RESULTS

Exposure to combat was significantly greater among those who were deployed to Iraq than among those deployed to Afghanistan. The percentage of study subjects whose responses met the screening criteria for major depression, generalized anxiety, or PTSD was significantly higher after duty in Iraq (15.6 to 17.1 percent) than after duty in Afghanistan (11.2 percent) or before deployment to Iraq (9.3 percent); the largest difference was in the rate of PTSD. Of those whose responses were positive for a mental disorder, only 23 to 40 percent sought mental health care. Those whose responses were positive for a mental disorder were twice as likely as those whose responses were negative to report concern about possible stigmatization and other barriers to seeking mental health care.

CONCLUSIONS

This study provides an initial look at the mental health of members of the Army and the Marine Corps who were involved in combat operations in Iraq and Afghanistan. Our findings indicate that among the study groups there was a significant risk of mental health problems and that the subjects reported important barriers to receiving mental health services, particularly the perception of stigma among those most in need of such care.

The evolution in next-generation sequencing (NGS) technology has led to the development of many different assembly algorithms, but few of them focus on assembling the organelle genomes. These genomes are used in phylogenetic studies, food identification and are the most deposited eukaryotic genomes in GenBank. Producing organelle genome assembly from whole genome sequencing (WGS) data would be the most accurate and least laborious approach, but a tool specifically designed for this task is lacking. We developed a seed-and-extend algorithm that assembles organelle genomes from whole genome sequencing (WGS) data, starting from a related or distant single seed sequence. The algorithm has been tested on several new (Gonioctena intermedia and Avicennia marina) and public (Arabidopsis thaliana and Oryza sativa) whole genome Illumina data sets where it outperforms known assemblers in assembly accuracy and coverage. In our benchmark, NOVOPlasty assembled all tested circular genomes in less than 30 min with a maximum memory requirement of 16 GB and an accuracy over 99.99%. In conclusion, NOVOPlasty is the sole de novo assembler that provides a fast and straightforward extraction of the extranuclear genomes from WGS data in one circular high quality contig. The software is open source and can be downloaded at https://github.com/ndierckx/NOVOPlasty.

The Generic Model Organism System Database Project (GMOD) seeks to develop reusable software components for model organism system databases. In this paper we describe the Generic Genome Browser (GBrowse), a Web-based application for displaying genomic annotations and other features. For the end user, features of the browser include the ability to scroll and zoom through arbitrary regions of a genome, to enter a region of the genome by searching for a landmark or performing a full text search of all features, and the ability to enable and disable tracks and change their relative order and appearance. The user can upload private annotations to view them in the context of the public ones, and publish those annotations to the community. For the data provider, features of the browser software include reliance on readily available open source components, simple installation, flexible configuration, and easy integration with other components of a model organism system Web site. GBrowse is freely available under an open source license. The software, its documentation, and support are available at http://www.gmod.org.

The crucial role played by the myofibroblast in wound healing and pathological organ remodeling is well established; the general mechanisms of extracellular matrix synthesis and of tension production by this cell have been amply clarified. This review discusses the pattern of myofibroblast accumulation and fibrosis evolution during lung and liver fibrosis as well as during atheromatous plaque formation. Special attention is paid to the specific features characterizing each of these processes, including the spectrum of different myofibroblast precursors and the distinct pathways involved in the formation of differentiated myofibroblasts in each lesion. Thus, whereas in lung fibrosis it seems that most myofibroblasts derive from resident fibroblasts, hepatic stellate cells are the main contributor for liver fibrosis and media smooth muscle cells are the main contributor for the atheromatous plaque. A better knowledge of the molecular mechanisms conducive to the appearance of differentiated myofibroblasts in each pathological situation will be useful for the understanding of fibrosis development in different organs and for the planning of strategies aiming at their prevention and therapy.

Resting state studies of spontaneous fluctuations in the functional MRI (fMRI) blood oxygen level dependent (BOLD) signal have shown great promise in mapping the brain's intrinsic, large-scale functional architecture. An important data preprocessing step used to enhance the quality of these observations has been removal of spontaneous BOLD fluctuations common to the whole brain (the so-called global signal). One reproducible consequence of global signal removal has been the finding that spontaneous BOLD fluctuations in the default mode network and an extended dorsal attention system are consistently anticorrelated, a relationship that these two systems exhibit during task performance. The dependence of these resting-state anticorrelations on global signal removal has raised important questions regarding the nature of the global signal, the validity of global signal removal, and the appropriate interpretation of observed anticorrelated brain networks. In this study, we investigate several properties of the global signal and find that it is, indeed, global, not residing preferentially in systems exhibiting anticorrelations. We detail the influence of global signal removal on resting state correlation maps both mathematically and empirically, showing an enhancement in detection of system-specific correlations and improvement in the correspondence between resting-state correlations and anatomy. Finally, we show that several characteristics of anticorrelated networks including their spatial distribution, cross-subject consistency, presence with modified whole brain masks, and existence before global regression are not attributable to global signal removal and therefore suggest a biological basis.

A novel method for differentiating between correctly and incorrectly determined regions of protein structures based on characteristic atomic interaction is described. Different types of atoms are distributed nonrandomly with respect to each other in proteins. Errors in model building lead to more randomized distributions of the different atom types, which can be distinguished from correct distributions by statistical methods. Atoms are classified in one of three categories: carbon (C), nitrogen (N), and oxygen (O). This leads to six different combinations of pairwise noncovalently bonded interactions (CC, CN, CO, NN, NO, and OO). A quadratic error function is used to characterize the set of pairwise interactions from nine-residue sliding windows in a database of 96 reliable protein structures. Regions of candidate protein structures that are mistraced or misregistered can then be identified by analysis of the pattern of nonbonded interactions from each window.

A new algorithm is reported which builds an alignment between two protein structures. The algorithm involves a combinatorial extension (CE) of an alignment path defined by aligned fragment pairs (AFPs) rather than the more conventional techniques using dynamic programming and Monte Carlo optimization. AFPs, as the name suggests, are pairs of fragments, one from each protein, which confer structure similarity. AFPs are based on local geometry, rather than global features such as orientation of secondary structures and overall topology. Combinations of AFPs that represent possible continuous alignment paths are selectively extended or discarded thereby leading to a single optimal alignment. The algorithm is fast and accurate in finding an optimal structure alignment and hence suitable for database scanning and detailed analysis of large protein families. The method has been tested and compared with results from Dali and VAST using a representative sample of similar structures. Several new structural similarities not detected by these other methods are reported. Specific one-on-one alignments and searches against all structures as found in the Protein Data Bank (PDB) can be performed via the Web at http://cl.sdsc.edu/ce.html.

MicroRNAs (miRNAs) are a class of noncoding RNAs found in organisms as evolutionarily distant as plants and mammals, yet most of the mRNAs they regulate are unknown. Here we show that the ability of an miRNA to translationally repress a target mRNA is largely dictated by the free energy of binding of the first eight nucleotides in the 5' region of the miRNA. However, G:U wobble base-pairing in this region interferes with activity beyond that predicted on the basis of thermodynamic stability. Furthermore, an mRNA can be simultaneously repressed by more than one miRNA species. The level of repression achieved is dependent on both the amount of mRNA and the amount of available miRNA complexes. Thus, predicted miRNA:mRNA interactions must be viewed in the context of other potential interactions and cellular conditions.

Reliability, the consistency of a test or measurement, is frequently quantified in the movement sciences literature. A common metric is the intraclass correlation coefficient (ICC). In addition, the SEM, which can be calculated from the ICC, is also frequently reported in reliability studies. However, there are several versions of the ICC, and confusion exists in the movement sciences regarding which ICC to use. Further, the utility of the SEM is not fully appreciated. In this review, the basics of classic reliability theory are addressed in the context of choosing and interpreting an ICC. The primary distinction between ICC equations is argued to be one concerning the inclusion (equations 2,1 and 2,k) or exclusion (equations 3,1 and 3,k) of systematic error in the denominator of the ICC equation. Inferential tests of mean differences, which are performed in the process of deriving the necessary variance components for the calculation of ICC values, are useful to determine if systematic error is present. If so, the measurement schedule should be modified (removing trials where learning and/or fatigue effects are present) to remove systematic error, and ICC equations that only consider random error may be safely used. The use of ICC values is discussed in the context of estimating the effects of measurement error on sample size, statistical power, and correlation attenuation. Finally, calculation and application of the SEM are discussed. It is shown how the SEM and its variants can be used to construct confidence intervals for individual scores and to determine the minimal difference needed to be exhibited for one to be confident that a true change in performance of an individual has occurred.

One of the most prominent neuropsychologic theories of attention-deficit/hyperactivity disorder (ADHD) suggests that its symptoms arise from a primary deficit in executive functions (EF), defined as neurocognitive processes that maintain an appropriate problem-solving set to attain a later goal. To examine the validity of the EF theory, we conducted a meta-analysis of 83 studies that administered EF measures to groups with ADHD (total N = 3734) and without ADHD (N = 2969). Groups with ADHD exhibited significant impairment on all EF tasks. Effect sizes for all measures fell in the medium range (.46-.69), but the strongest and most consistent effects were obtained on measures of response inhibition, vigilance, working memory, and planning. Weaknesses in EF were significant in both clinic-referred and community samples and were not explained by group differences in intelligence, academic achievement, or symptoms of other disorders. ADHD is associated with significant weaknesses in several key EF domains. However, moderate effect sizes and lack of universality of EF deficits among individuals with ADHD suggest that EF weaknesses are neither necessary nor sufficient to cause all cases of ADHD. Difficulties with EF appear to be one important component of the complex neuropsychology of ADHD.

FoxP3 is a key transcription factor for the development and function of natural CD4(+) regulatory T cells (Treg cells). Here we show that human FoxP3(+)CD4(+) T cells were composed of three phenotypically and functionally distinct subpopulations: CD45RA(+)FoxP3(lo) resting Treg cells (rTreg cells) and CD45RA(-)FoxP3(hi) activated Treg cells (aTreg cells), both of which were suppressive in vitro, and cytokine-secreting CD45RA(-)FoxP3(lo) nonsuppressive T cells. The proportion of the three subpopulations differed between cord blood, aged individuals, and patients with immunological diseases. Terminally differentiated aTreg cells rapidly died whereas rTreg cells proliferated and converted into aTreg cells in vitro and in vivo. This was shown by the transfer of rTreg cells into NOD-scid-common gamma-chain-deficient mice and by TCR sequence-based T cell clonotype tracing in peripheral blood in a normal individual. Taken together, the dissection of FoxP3(+) cells into subsets enables one to analyze Treg cell differentiation dynamics and interactions in normal and disease states, and to control immune responses through manipulating particular FoxP3(+) subpopulations.

The idea of reserve against brain damage stems from the repeated observation that there does not appear to be a direct relationship between the degree of brain pathology or brain damage and the clinical manifestation of that damage. This paper attempts to develop a coherent theoretical account of reserve. One convenient subdivision of reserve models revolves around whether they envision reserve as a passive process, such as in brain reserve or threshold, or see the brain as actively attempting to cope with or compensate for pathology, as in cognitive reserve. Cognitive reserve may be based on more efficient utilization of brain networks or of enhanced ability to recruit alternate brain networks as needed. A distinction is suggested between reserve, the ability to optimize or maximize normal performance, and compensation, an attempt to maximize performance in the face of brain damage by using brain structures or networks not engaged when the brain is not damaged. Epidemiologic and imaging data that help to develop and support the concept of reserve are presented.

The coronavirus disease 2019 (COVID-19) pandemic is having a profound effect on all aspects of society, including mental health and physical health. We explore the psychological, social, and neuroscientific effects of COVID-19 and set out the immediate priorities and longer-term strategies for mental health science research. These priorities were informed by surveys of the public and an expert panel convened by the UK Academy of Medical Sciences and the mental health research charity, MQ: Transforming Mental Health, in the first weeks of the pandemic in the UK in March, 2020. We urge UK research funding agencies to work with researchers, people with lived experience, and others to establish a high level coordination group to ensure that these research priorities are addressed, and to allow new ones to be identified over time. The need to maintain high-quality research standards is imperative. International collaboration and a global perspective will be beneficial. An immediate priority is collecting high-quality data on the mental health effects of the COVID-19 pandemic across the whole population and vulnerable groups, and on brain function, cognition, and mental health of patients with COVID-19. There is an urgent need for research to address how mental health consequences for vulnerable groups can be mitigated under pandemic conditions, and on the impact of repeated media consumption and health messaging around COVID-19. Discovery, evaluation, and refinement of mechanistically driven interventions to address the psychological, social, and neuroscientific aspects of the pandemic are required. Rising to this challenge will require integration across disciplines and sectors, and should be done together with people with lived experience. New funding will be required to meet these priorities, and it can be efficiently leveraged by the UK's world-leading infrastructure. This Position Paper provides a strategy that may be both adapted for, and integrated with, research efforts in other countries.

We show that c-myc is an inducible gene that is regulated by specific growth signals in a cell-cycle-dependent manner. Specifically, agents that initiate the first phase of a proliferative response in lymphocytes (lipopolysaccharide or Concanavalin A) and fibroblasts (platelet-derived growth factor) induce c-myc mRNA. Within one to three hr after the addition of these mitogens to the appropriate cells, c-myc mRNA concentration is increased between 10- and 40-fold. This induction of c-myc mRNA occurs in the presence of cycloheximide and, therefore, does not require the synthesis of new protein species. Consequently, the induction of c-myc mRNA is not secondary to growth. In addition, c-myc mRNA is "superinduced" by the combination of cycloheximide and mitogen, a finding consistent with a model that a labile protein may regulate c-myc levels in these cells. Further, this work suggests a regulatory linkage between the function of two oncogenes--c-myc and c-sis--the latter being the putative structural gene for PDGF.

One of the greatest challenges facing human geneticists is the identification and characterization of susceptibility genes for common complex multifactorial human diseases. This challenge is partly due to the limitations of parametric-statistical methods for detection of gene effects that are dependent solely or partially on interactions with other genes and with environmental exposures. We introduce multifactor-dimensionality reduction (MDR) as a method for reducing the dimensionality of multilocus information, to improve the identification of polymorphism combinations associated with disease risk. The MDR method is nonparametric (i.e., no hypothesis about the value of a statistical parameter is made), is model-free (i.e., it assumes no particular inheritance model), and is directly applicable to case-control and discordant-sib-pair studies. Using simulated case-control data, we demonstrate that MDR has reasonable power to identify interactions among two or more loci in relatively small samples. When it was applied to a sporadic breast cancer case-control data set, in the absence of any statistically significant independent main effects, MDR identified a statistically significant high-order interaction among four polymorphisms from three different estrogen-metabolism genes. To our knowledge, this is the first report of a four-locus interaction associated with a common complex multifactorial disease.

Movement disorders associated with basal ganglia dysfunction comprise a spectrum of abnormalities that range from the hypokinetic disorders (of which Parkinson's disease is the best-known example) at one extreme to the hyperkinetic disorders (exemplified by Huntington's disease and hemiballismus) at the other. Both extremes of this movement disorder spectrum can be accounted for by postulating specific disturbances within the basal ganglia-thalamocortical 'motor' circuit. In this paper, Mahlon DeLong describes the changes in neuronal activity in the motor circuit in animal models of hypo- and hyperkinetic disorders.

Two types of cannabinoid receptor have been discovered so far, CB(1) (2.1: CBD:1:CB1:), cloned in 1990, and CB(2) (2.1:CBD:2:CB2:), cloned in 1993. Distinction between these receptors is based on differences in their predicted amino acid sequence, signaling mechanisms, tissue distribution, and sensitivity to certain potent agonists and antagonists that show marked selectivity for one or the other receptor type. Cannabinoid receptors CB(1) and CB(2) exhibit 48% amino acid sequence identity. Both receptor types are coupled through G proteins to adenylyl cyclase and mitogen-activated protein kinase. CB(1) receptors are also coupled through G proteins to several types of calcium and potassium channels. These receptors exist primarily on central and peripheral neurons, one of their functions being to inhibit neurotransmitter release. Indeed, endogenous CB(1) agonists probably serve as retrograde synaptic messengers. CB(2) receptors are present mainly on immune cells. Such cells also express CB(1) receptors, albeit to a lesser extent, with both receptor types exerting a broad spectrum of immune effects that includes modulation of cytokine release. Of several endogenous agonists for cannabinoid receptors identified thus far, the most notable are arachidonoylethanolamide, 2-arachidonoylglycerol, and 2-arachidonylglyceryl ether. It is unclear whether these eicosanoid molecules are the only, or primary, endogenous agonists. Hence, we consider it premature to rename cannabinoid receptors after an endogenous agonist as is recommended by the International Union of Pharmacology Committee on Receptor Nomenclature and Drug Classification. Although pharmacological evidence for the existence of additional types of cannabinoid receptor is emerging, other kinds of supporting evidence are still lacking.

BACKGROUND

The lowering of cholesterol concentrations in individuals at high risk of cardiovascular disease improves outcome. No study, however, has assessed benefits of cholesterol lowering in the primary prevention of coronary heart disease (CHD) in hypertensive patients who are not conventionally deemed dyslipidaemic.

METHODS

Of 19342 hypertensive patients (aged 40-79 years with at least three other cardiovascular risk factors) randomised to one of two antihypertensive regimens in the Anglo-Scandinavian Cardiac Outcomes Trial, 10305 with non-fasting total cholesterol concentrations 6.5 mmol/L or less were randomly assigned additional atorvastatin 10 mg or placebo. These patients formed the lipid-lowering arm of the study. We planned follow-up for an average of 5 years, the primary endpoint being non-fatal myocardial infarction and fatal CHD. Data were analysed by intention to treat.

RESULTS

Treatment was stopped after a median follow-up of 3.3 years. By that time, 100 primary events had occurred in the atorvastatin group compared with 154 events in the placebo group (hazard ratio 0.64 [95% CI 0.50-0.83], p=0.0005). This benefit emerged in the first year of follow-up. There was no significant heterogeneity among prespecified subgroups. Fatal and non-fatal stroke (89 atorvastatin vs 121 placebo, 0.73 [0.56-0.96], p=0.024), total cardiovascular events (389 vs 486, 0.79 [0.69-0.90], p=0.0005), and total coronary events (178 vs 247, 0.71 [0.59-0.86], p=0.0005) were also significantly lowered. There were 185 deaths in the atorvastatin group and 212 in the placebo group (0.87 [0.71-1.06], p=0.16). Atorvastatin lowered total serum cholesterol by about 1.3 mmol/L compared with placebo at 12 months, and by 1.1 mmol/L after 3 years of follow-up.

CONCLUSIONS

The reductions in major cardiovascular events with atorvastatin are large, given the short follow-up time. These findings may have implications for future lipid-lowering guidelines.

One of the most consistent observations in human functional imaging is that a network of brain regions referred to as the "default network" increases its activity during passive states. Here we explored the anatomy and function of the default network across three studies to resolve divergent hypotheses about its contributions to spontaneous cognition and active forms of decision making. Analysis of intrinsic activity revealed the network comprises multiple, dissociated components. A midline core (posterior cingulate and anterior medial prefrontal cortex) is active when people make self-relevant, affective decisions. In contrast, a medial temporal lobe subsystem becomes engaged when decisions involve constructing a mental scene based on memory. During certain experimentally directed and spontaneous acts of future-oriented thought, these dissociated components are simultaneously engaged, presumably to facilitate construction of mental models of personally significant events.

<AbstractText>A cluster of patients with coronavirus disease 2019 (COVID-19) pneumonia caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were successively reported in Wuhan, China. We aimed to describe the CT findings across different timepoints throughout the disease course.</AbstractText><AbstractText>Patients with COVID-19 pneumonia (confirmed by next-generation sequencing or RT-PCR) who were admitted to <em>one</em> of two hospitals in Wuhan and who underwent serial chest CT scans were retrospectively enrolled. Patients were grouped on the basis of the interval between symptom onset and the first CT scan: group 1 (subclinical patients; scans d<em>one</em> before symptom onset), group 2 (scans d<em>one</em> ≤1 week after symptom onset), group 3 (>1 week to 2 weeks), and group 4 (>2 weeks to 3 weeks). Imaging features and their distribution were analysed and compared across the four groups.</AbstractText><AbstractText>81 patients admitted to hospital between Dec 20, 2019, and Jan 23, 2020, were retrospectively enrolled. The cohort included 42 (52%) men and 39 (48%) women, and the mean age was 49·5 years (SD 11·0). The mean number of involved lung segments was 10·5 (SD 6·4) overall, 2·8 (3·3) in group 1, 11·1 (5·4) in group 2, 13·0 (5·7) in group 3, and 12·1 (5·9) in group 4. The predominant pattern of abnormality observed was bilateral (64 [79%] patients), peripheral (44 [54%]), ill-defined (66 [81%]), and ground-glass opacification (53 [65%]), mainly involving the right lower lobes (225 [27%] of 849 affected segments). In group 1 (n=15), the predominant pattern was unilateral (nine [60%]) and multifocal (eight [53%]) ground-glass opacities (14 [93%]). Lesions quickly evolved to bilateral (19 [90%]), diffuse (11 [52%]) ground-glass opacity predominance (17 [81%]) in group 2 (n=21). Thereafter, the prevalence of ground-glass opacities continued to decrease (17 [57%] of 30 patients in group 3, and five [33%] of 15 in group 4), and consolidation and mixed patterns became more frequent (12 [40%] in group 3, eight [53%] in group 4).</AbstractText><AbstractText>COVID-19 pneumonia manifests with chest CT imaging abnormalities, even in asymptomatic patients, with rapid evolution from focal unilateral to diffuse bilateral ground-glass opacities that progressed to or co-existed with consolidations within 1-3 weeks. Combining assessment of imaging features with clinical and laboratory findings could facilitate early diagnosis of COVID-19 pneumonia.</AbstractText><AbstractText>N<em>one</em>.</AbstractText>

BACKGROUND

Visualisation of genome comparisons is invaluable for helping to determine genotypic differences between closely related prokaryotes. New visualisation and abstraction methods are required in order to improve the validation, interpretation and communication of genome sequence information; especially with the increasing amount of data arising from next-generation sequencing projects. Visualising a prokaryote genome as a circular image has become a powerful means of displaying informative comparisons of one genome to a number of others. Several programs, imaging libraries and internet resources already exist for this purpose, however, most are either limited in the number of comparisons they can show, are unable to adequately utilise draft genome sequence data, or require a knowledge of command-line scripting for implementation. Currently, there is no freely available desktop application that enables users to rapidly visualise comparisons between hundreds of draft or complete genomes in a single image.

RESULTS

BLAST Ring Image Generator (BRIG) can generate images that show multiple prokaryote genome comparisons, without an arbitrary limit on the number of genomes compared. The output image shows similarity between a central reference sequence and other sequences as a set of concentric rings, where BLAST matches are coloured on a sliding scale indicating a defined percentage identity. Images can also include draft genome assembly information to show read coverage, assembly breakpoints and collapsed repeats. In addition, BRIG supports the mapping of unassembled sequencing reads against one or more central reference sequences. Many types of custom data and annotations can be shown using BRIG, making it a versatile approach for visualising a range of genomic comparison data. BRIG is readily accessible to any user, as it assumes no specialist computational knowledge and will perform all required file parsing and BLAST comparisons automatically.

CONCLUSIONS

There is a clear need for a user-friendly program that can produce genome comparisons for a large number of prokaryote genomes with an emphasis on rapidly utilising unfinished or unassembled genome data. Here we present BRIG, a cross-platform application that enables the interactive generation of comparative genomic images via a simple graphical-user interface. BRIG is freely available for all operating systems at http://sourceforge.net/projects/brig/.

A discontinuous electrophoretic system for the isolation of membrane proteins from acrylamide gels has been developed using equipment for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Coomassie dyes were introduced to induce a charge shift on the proteins and aminocaproic acid served to improve solubilization of membrane proteins. Solubilized mitochondria or extracts of heart muscle tissue, lymphoblasts, yeast, and bacteria were applied to the gels. From cells containing mitochondria, all the multiprotein complexes of the oxidative phosphorylation system were separated within one gel. The complexes were resolved into the individual polypeptides by second-dimension Tricine-SDS-PAGE or extracted without SDS for functional studies. The recovery of all respiratory chain complexes was almost quantitative. The percentage recovery of functional activity depended on the respective protein complex studied and was zero for some complexes, but almost quantitative for others. The system is especially useful for small scale purposes, e.g., separation of radioactively labeled membrane proteins, N-terminal protein sequencing, preparation of proteins for immunization, and diagnostic studies of inborn neuromuscular diseases.

Meiotic recombination in S. cerevisiae is initiated by double-strand breaks (DSBs). In certain mutants, breaks accumulate with a covalently attached protein, suggesting that cleavage is catalyzed by the DSB-associated protein via a topoisomerase-like transesterase mechanism. We have purified these protein-DNA complexes and identified the protein as Spo11, one of several proteins required for DSB formation. These findings strongly implicate Spo11 as the catalytic subunit of the meiotic DNA cleavage activity. This is the first identification of a biochemical function for any of the gene products involved in DSB formation. Spo11 defines a protein family with other members in fission yeast, nematodes, and archaebacteria. The S. pombe homolog, rec12p, is also known to be required for meiotic recombination. Thus, these findings provide direct evidence that the mechanism of meiotic recombination initiation is evolutionarily conserved.