A proteome-scale map of the human interactome network.

Just as reference genome sequences revolutionized human genetics, reference maps of interactome networks will be critical to fully understand genotype-phenotype relationships. Here, we describe a systematic map of ?14,000 high-quality human binary protein-protein interactions. At equal quality, this map is ?30% larger than what is available from small-scale studies published in the literature in the last few decades. While currently available information is highly biased and only covers a relatively small portion of the proteome, our systematic map appears strikingly more homogeneous, revealing a "broader" human interactome network than currently appreciated. The map also uncovers significant interconnectivity between known and candidate cancer gene products, providing unbiased evidence for an expanded functional cancer landscape, while demonstrating how high-quality interactome models will help "connect the dots" of the genomic revolution.

Truncating variant burden in high-functioning autism and pleiotropic effects of LRP1 across psychiatric phenotypes

Background: Previous research has implicated de novo and inherited truncating mutations in autism-spectrum disorder. We aim to investigate whether the load of inherited truncating mutations contributes similarly to high-functioning autism, and to characterize genes that harbour de novo variants in high-functioning autism. Methods: We performed whole-exome sequencing in 20 high-functioning autism families (average IQ = 100). Results: We observed no difference in the number of transmitted versus nontransmitted truncating alleles for high-functioning autism (117 v. 130, p = 0.78). Transmitted truncating and de novo variants in high-functioning autism were not enriched in gene ontology (GO) or Kyoto Encyclopedia of Genes and Genomes (KEGG) categories, or in autism-related gene sets. However, in a patient with high-functioning autism we identified a de novo variant in a canonical splice site of LRP1, a postsynaptic density gene that is a target for fragile X mental retardation protein (FRMP). This de novo variant leads to in-frame skipping of exon 29, removing 2 of 6 blades of the ß-propeller domain 4 of LRP1, with putative functional consequences. Large data sets implicate LRP1 across a number of psychiatric disorders: de novo variants are associated with autism-spectrum disorder (p = 0.039) and schizophrenia (p = 0.008) from combined sequencing projects; common variants using genome-wide association study data sets from the Psychiatric Genomics Consortium show gene-based association in schizophrenia (p = 6.6 × E−07) and in a meta-analysis across 7 psychiatric disorders (p = 2.3 × E−03); and the burden of ultra-rare pathogenic variants has been shown to be higher in autism-spectrum disorder (p = 1.2 × E−05), using whole-exome sequencing from 6135 patients with schizophrenia, 1778 patients with autism-spectrum disorder and 7875 controls. Limitations: We had a limited sample of patients with high-functioning autism, related to difficulty in recruiting probands with high cognitive performance and no family history of psychiatric disorders. Conclusion: Previous studies and ours suggest an effect of truncating mutations restricted to severe autism-spectrum disorder phenotypes that are associated with intellectual disability. We provide evidence for pleiotropic effects of common and rare variants in the LRP1 gene across psychiatric phenotypes.

A PanorOmic view of personal cancer genomes.

The massive molecular profiling of thousands of cancer patients has led to the identification of many tumor type specific driver genes. However, only a few (or none) of them are present in each individual tumor and, to enable precision oncology, we need to interpret the alterations found in a single patient. Cancer PanorOmics (http://panoromics.irbbarcelona.org) is a web-based resource to contextualize genomic variations detected in a personal cancer genome within the body of clinical and scientific evidence available for 26 tumor types, offering complementary cohort- and patient-centric views. Additionally, it explores the cellular environment of mutations by mapping them on the human interactome and providing quasi-atomic structural details, whenever available. This 'PanorOmic' molecular view of individual tumors, together with the appropriate genetic counselling and medical advice, should contribute to the identification of actionable alterations ultimately guiding the clinical decision-making process.

Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods.

An inter-species protein-protein interaction network across vast evolutionary distance.

In cellular systems, biophysical interactions between macromolecules underlie a complex web of functional interactions. How biophysical and functional networks are coordinated, whether all biophysical interactions correspond to functional interactions, and how such biophysical-versus-functional network coordination is shaped by evolutionary forces are all largely unanswered questions. Here, we investigate these questions using an "inter-interactome" approach. We systematically probed the yeast and human proteomes for interactions between proteins from these two species and functionally characterized the resulting inter-interactome network. After a billion years of evolutionary divergence, the yeast and human proteomes are still capable of forming a biophysical network with properties that resemble those of intra-species networks. Although substantially reduced relative to intra-species networks, the levels of functional overlap in the yeast-human inter-interactome network uncover significant remnants of co-functionality widely preserved in the two proteomes beyond human-yeast homologs. Our data support evolutionary selection against biophysical interactions between proteins with little or no co-functionality. Such non-functional interactions, however, represent a reservoir from which nascent functional interactions may arise.

Interactome3D: adding structural details to protein networks.

Network-centered approaches are increasingly used to understand the fundamentals of biology. However, the molecular details contained in the interaction networks, often necessary to understand cellular processes, are very limited, and the experimental difficulties surrounding the determination of protein complex structures make computational modeling techniques paramount. Here we present Interactome3D, a resource for the structural annotation and modeling of protein-protein interactions. Through the integration of interaction data from the main pathway repositories, we provide structural details at atomic resolution for over 12,000 protein-protein interactions in eight model organisms. Unlike static databases, Interactome3D also allows biologists to upload newly discovered interactions and pathways in any species, select the best combination of structural templates and build three-dimensional models in a fully automated manner. Finally, we illustrate the value of Interactome3D through the structural annotation of the complement cascade pathway, rationalizing a potential common mechanism of action suggested for several disease-causing mutations.

A second-generation protein-protein interaction network of Helicobacter pylori.

Helicobacter pylori infections cause gastric ulcers and play a major role in the development of gastric cancer. In 2001, the first protein interactome was published for this species, revealing over 1500 binary protein interactions resulting from 261 yeast two-hybrid screens. Here we roughly double the number of previously published interactions using an ORFeome-based, proteome-wide yeast two-hybrid screening strategy. We identified a total of 1515 protein-protein interactions, of which 1461 are new. The integration of all the interactions reported in H. pylori results in 3004 unique interactions that connect about 70% of its proteome. Excluding interactions of promiscuous proteins we derived from our new data a core network consisting of 908 interactions. We compared our data set to several other bacterial interactomes and experimentally benchmarked the conservation of interactions using 365 protein pairs (interologs) of E. coli of which one third turned out to be conserved in both species.

IVF cycle cost estimation using Activity Based Costing and Monte Carlo simulation.

The Authors present a new methodological approach in stochastic regime to determine the actual costs of an healthcare process. The paper specifically shows the application of the methodology for the determination of the cost of an Assisted reproductive technology (ART) treatment in Italy. The reason of this research comes from the fact that deterministic regime is inadequate to implement an accurate estimate of the cost of this particular treatment. In fact the durations of the different activities involved are unfixed and described by means of frequency distributions. Hence the need to determine in addition to the mean value of the cost, the interval within which it is intended to vary with a known confidence level. Consequently the cost obtained for each type of cycle investigated (in vitro fertilization and embryo transfer with or without intracytoplasmic sperm injection), shows tolerance intervals around the mean value sufficiently restricted as to make the data obtained statistically robust and therefore usable also as reference for any benchmark with other Countries. It should be noted that under a methodological point of view the approach was rigorous. In fact it was used both the technique of Activity Based Costing for determining the cost of individual activities of the process both the Monte Carlo simulation, with control of experimental error, for the construction of the tolerance intervals on the final result.

3did: a catalog of domain-based interactions of known three-dimensional structure.

The database of 3D interacting domains (3did, available online for browsing and bulk download at http://3did.irbbarcelona.org) is a catalog of protein-protein interactions for which a high-resolution 3D structure is known. 3did collects and classifies all structural templates of domain-domain interactions in the Protein Data Bank, providing molecular details for such interactions. The current version also includes a pipeline for the discovery and annotation of novel domain-motif interactions. For every interaction, 3did identifies and groups different binding modes by clustering similar interfaces into 'interaction topologies'. By maintaining a constantly updated collection of domain-based structural interaction templates, 3did is a reference source of information for the structural characterization of protein interaction networks. 3did is updated every 6 months.

Modeling, Fabrication and Testing of a Customizable Micromachined Hotplate for Sensor Applications.

In the sensors field the active sensing material frequently needs a controlled temperature in order to work properly. In microsystems technology, micro-machined hotplates represent a platform consisting of a thin suspended membrane where the sensing material can be deposited, usually integrating electrical stimuli and temperature readout. The micro-hotplate ensures a series of advantages such as miniaturized size, fast response, high sensitivity, low power consumption and selectivity for chemical sensing. This work compares the coplanar and the buried approach for the micro-hotplate heaters design with the aim to optimize the fabrication process and to propose a guideline for the choice of the suitable design with respect to the applications. In particular, robust Finite Element Method (FEM) models are set up in order to predict the electrical and thermal behavior of the micro-hotplates. The multiphysics approach used for the simulation allows to match as close as possible the actual device to the predictive model: geometries, materials, physics have been carefully linked to the fabricated devices to obtain the best possible accuracy. The materials involved in the fabrication process are accurately selected in order to improve the yield of the process and the performance of the devices. The fabricated micro-hotplates are able to warm the active region up to 400 °C (with a corresponding power consumption equal to 250 mW @ 400 °C) with a uniform temperature distribution in the buried micro-hotplate and a controlled temperature gradient in the coplanar one. A response time of about 70 ms was obtained on the virtual model, which perfectly agrees with the one measured on the fabricated device. Besides morphological, electrical and thermal characterizations, this work includes reliability tests in static and dynamic modes.

Yeast mitochondrial protein-protein interactions reveal diverse complexes and disease-relevant functional relationships.

Although detailed, focused, and mechanistic analyses of associations among mitochondrial proteins (MPs) have identified their importance in varied biological processes, a systematic understanding of how MPs function in concert both with one another and with extra-mitochondrial proteins remains incomplete. Consequently, many questions regarding the role of mitochondrial dysfunction in the development of human disease remain unanswered. To address this, we compiled all existing mitochondrial physical interaction data for over 1200 experimentally defined yeast MPs and, through bioinformatic analysis, identified hundreds of heteromeric MP complexes having extensive associations both within and outside the mitochondria. We provide support for these complexes through structure prediction analysis, morphological comparisons of deletion strains, and protein co-immunoprecipitation. The integration of these MP complexes with reported genetic interaction data reveals substantial crosstalk between MPs and non-MPs and identifies novel factors in endoplasmic reticulum-mitochondrial organization, membrane structure, and mitochondrial lipid homeostasis. More than one-third of these MP complexes are conserved in humans, with many containing members linked to clinical pathologies, enabling us to identify genes with putative disease function through guilt-by-association. Although still remaining incomplete, existing mitochondrial interaction data suggests that the relevant molecular machinery is modular, yet highly integrated with non-mitochondrial processes.

Liposome sensing and monitoring by organic electrochemical transistors integrated in microfluidics.

BACKGROUND: Organic electrochemical transistors (OECTs), which are becoming more and more promising devices for applications in bioelectronics and nanomedicine, are proposed here as ideally suitable for sensing and real time monitoring of liposome-based structures. This is quite relevant since, currently, the techniques used to investigate liposomal structures, their stability in different environments as well as drug loading and delivery mechanisms, operate basically off-line and/or with pre-prepared sampling. METHODS: OECTs, based on the PEDOT:PSS conductive polymer, have been employed as sensors of liposome-based nanoparticles in electrolyte solutions to assess sensitivity and monitoring capabilities based on ion-to-electron amplified transduction. RESULTS: We demonstrate that OECTs are very efficient, reliable and sensitive devices for detecting liposome-based nanoparticles on a wide dynamic range down to 10(-5)mg/ml (with a lowest detection limit, assessed in real-time monitoring, of 10(-7)mg/ml), thus matching the needs of typical drug loading/drug delivery conditions. They are hence particularly well suited for real-time monitoring of liposomes in solution. Furthermore, OECTs are shown to sense and discriminate successive injection of different liposomes, so that they could be good candidates in quality-control assays or in the pharmaceutical industry. GENERAL SIGNIFICANCE: Drug loading and delivery by liposome-based structures is a fast growing and very promising field that will strongly benefit from real-time, highly sensitive and low cost monitoring of their dynamics in different pharma and biomedical environments, with a particular reference to the pharmaceutical and production processes, where a major issue is monitoring and measuring the formation and concentration of liposomes and the relative drug load. The demonstrated ability to sense and monitor complex bio-structures, such as liposomes, paves the way for very promising developments in biosensing and nanomedicine. This article is part of a Special Issue entitled Organic Bioelectronics-Novel Applications in Biomedicine.

The role of structural disorder in the rewiring of protein interactions through evolution.

Structurally disordered regions play a key role in protein-protein interaction networks and the evolution of highly connected proteins, enabling the molecular mechanisms for multiple binding. However, the role of protein disorder in the evolution of interaction networks has only been investigated through the analysis of individual proteins, making it impossible to distinguish its specific impact in the (re)shaping of their interaction environments. Now, the availability of large interactomes for several model organisms permits exploration of the role of disorder in protein interaction networks not only at the level of the interacting proteins but of the interactions themselves. By comparing the interactomes of human, fly, and yeast, we discovered that, despite being much more abundant, disordered interactions are significantly less conserved than their ordered counterparts. Furthermore, our analyses provide evidence that this happens not only because disordered proteins are less conserved but also because they display a higher capacity to rewire their interaction neighborhood through evolution. Overall, our results support the hypothesis that conservation of disorder gives a clear evolutionary advantage, facilitating the change of interaction partners during evolution. Moreover, this mechanism is not exclusive of a few anecdotal cases but a global feature present in the interactome networks of entire organisms.

Low temperature sensing properties of a nano hybrid material based on ZnO nanotetrapods and titanyl phthalocyanine.

ZnO nanotetrapods have recently been exploited for the realization of high-sensitivity gas sensors, but they are affected by the typical drawbacks of metal-oxides, i.e., poor selectivity and a relatively high working temperature. On the other hand, it has been also demonstrated that the combined use of nanostructured metal oxides and organic molecules can improve the gas sensing performance sensitivity or selectivity, even at lower temperatures. A gas sensor device, based on films of interconnected ZnO nanotetrapods properly functionalized by titanyl phthalocyanine (TiOPc), has been realized in order to combine the high surface to volume ratio and structural stability of the crystalline ZnO nanostructures with the enhanced sensitivity of the semiconducting TiOPc molecule, especially at low temperature. The electronic properties of the resulting nanohybrid material are different from those of each single component. The response of the hybrid nanostructure towards different gases has been compared with that of ZnO nanotetrapod without functionalization in order to highlight the peculiar properties of the hybrid interaction(s). The dynamic response in time has been studied for different gases and temperatures; in particular, an increase in the response to NO2 has been observed, even at room temperature. The formation of localized p-n heterojunctions and the possibility of exchanging charge carriers at the hybrid interface is shown to be crucial for the sensing mechanism.

Buckling critical pressures in collapsible tubes relevant for biomedical flows.

The behaviour of collapsed or stenotic vessels in the human body can be studied by means of simplified geometries like a collapsible tube. The objective of this work is to determine the value of the buckling critical pressure of a collapsible tube by employing Landau's theory of phase transition. The methodology is based on the implementation of an experimentally validated 3D numerical model of a collapsible tube. The buckling critical pressure is estimated for different values of geometric parameters of the system by treating the relation between the intramural pressure and the area of the central cross-section as the order parameter function of the system. The results show the dependence of the buckling critical pressures on the geometric parameters of a collapsible tube. General non-dimensional equations for the buckling critical pressures are derived. The advantage of this method is that it does not require any geometric assumption, but it is solely based on the observation that the buckling of a collapsible tube can be treated as a second-order phase transition. The investigated geometric and elastic parameters are sensible for biomedical application, with particular interest to the study of the bronchial tree under pathophysiological conditions like asthma.

The influence of catalysis on mad2 activation dynamics.

Mad2 is a key component of the spindle assembly checkpoint, a safety device ensuring faithful sister chromatid separation in mitosis. The target of Mad2 is Cdc20, an activator of the anaphase-promoting complex/cyclosome (APC/C). Mad2 binding to Cdc20 is a complex reaction that entails the conformational conversion of Mad2 from an open (O-Mad2) to a closed (C-Mad2) conformer. Previously, it has been hypothesized that the conversion of O-Mad2 is accelerated by its conformational dimerization with C-Mad2. This hypothesis, known as the Mad2-template hypothesis, is based on the unproven assumption that the natural conversion of O-Mad2 required to bind Cdc20 is slow. Here, we provide evidence for this fundamental assumption and demonstrate that conformational dimerization of Mad2 accelerates the rate of Mad2 binding to Cdc20. On the basis of our measurements, we developed a set of rate equations that deliver excellent predictions of experimental binding curves under a variety of different conditions. Our results strongly suggest that the interaction of Mad2 with Cdc20 is rate limiting for activation of the spindle checkpoint. Conformational dimerization of Mad2 is essential to accelerate Cdc20 binding, but it does not modify the equilibrium of the Mad2:Cdc20 interaction, i.e., it is purely catalytic. These results surpass previously formulated objections to the Mad2-template model and predict that the release of Mad2 from Cdc20 is an energy-driven process.

Does early extubation after cardiac surgery lead to a reduction in intensive care unit length of stay?

A best evidence topic in cardiac surgery was written according to a structured protocol. The question addressed was whether early extubation (EE) after cardiac surgery leads to a reduction in intensive care unit (ICU) length of stay (LOS)? A total of 564 papers were found using the reported search, of which 4 were randomized trials and hence represented the best evidence to answer the clinical question. The authors, journal, date and country of publication, patient group studied, study type, relevant outcomes and results of these papers are tabulated. EE was defined as extubation in theatre (n = 2), within 6 h of surgery (n = 1) and within 8 h of surgery (n = 1). EE was associated with significantly reduced ICU LOS in all studies. Despite the Society of Thoracic Surgeons using extubation <6 h after surgery as a measure of quality, this study has demonstrated that no standardized definition for EE currently exists. The body of evidence identified in this work has demonstrated that for appropriately selected patients (avoiding patients with multiple comorbidities, advanced age and undergoing complex non-elective surgery) early tracheal extubation is associated with a reduction in ICU LOS without an increase in the rate of postoperative complications.

Pushing structural information into the yeast interactome by high-throughput protein docking experiments.

The last several years have seen the consolidation of high-throughput proteomics initiatives to identify and characterize protein interactions and macromolecular complexes in model organisms. In particular, more that 10,000 high-confidence protein-protein interactions have been described between the roughly 6,000 proteins encoded in the budding yeast genome (Saccharomyces cerevisiae). However, unfortunately, high-resolution three-dimensional structures are only available for less than one hundred of these interacting pairs. Here, we expand this structural information on yeast protein interactions by running the first-ever high-throughput docking experiment with some of the best state-of-the-art methodologies, according to our benchmarks. To increase the coverage of the interaction space, we also explore the possibility of using homology models of varying quality in the docking experiments, instead of experimental structures, and assess how it would affect the global performance of the methods. In total, we have applied the docking procedure to 217 experimental structures and 1,023 homology models, providing putative structural models for over 3,000 protein-protein interactions in the yeast interactome. Finally, we analyze in detail the structural models obtained for the interaction between SAM1-anthranilate synthase complex and the MET30-RNA polymerase III to illustrate how our predictions can be straightforwardly used by the scientific community. The results of our experiment will be integrated into the general 3D-Repertoire pipeline, a European initiative to solve the structures of as many as possible protein complexes in yeast at the best possible resolution. All docking results are available at http://gatealoy.pcb.ub.es/HT_docking/.

RAPIDO: a web server for the alignment of protein structures in the presence of conformational changes.

Rapid alignment of proteins in terms of domains (RAPIDO) is a web server for the 3D alignment of crystal structures of different protein molecules in the presence of conformational change. The structural alignment algorithm identifies groups of equivalent atoms whose interatomic distances are constant (within a defined tolerance) in the two structures being compared and considers these groups of atoms as rigid bodies. In addition to the functionalities provided by existing tools, RAPIDO can identify structurally equivalent regions also when these consist of fragments that are distant in terms of sequence and separated by other movable domains. Furthermore, RAPIDO takes the variation in the reliability of atomic coordinates into account in the comparison of distances between equivalent atoms by employing weighting-functions based on the refined B-values. The regions identified as equivalent by RAPIDO furnish reliable sets of residues for the superposition of the two structures for subsequent detailed analysis. The RAPIDO server, with related documentation, is available at http://webapps.embl-hamburg.de/rapido.