Real-time dynamic single-molecule protein sequencing on an integrated semiconductor device.

Studies of the proteome would benefit greatly from methods to directly sequence and digitally quantify proteins and detect posttranslational modifications with single-molecule sensitivity. Here, we demonstrate single-molecule protein sequencing using a dynamic approach in which single peptides are probed in real time by a mixture of dye-labeled N-terminal amino acid recognizers and simultaneously cleaved by aminopeptidases. We annotate amino acids and identify the peptide sequence by measuring fluorescence intensity, lifetime, and binding kinetics on an integrated semiconductor chip. Our results demonstrate the kinetic principles that allow recognizers to identify multiple amino acids in an information-rich manner that enables discrimination of single amino acid substitutions and posttranslational modifications. With further development, we anticipate that this approach will offer a sensitive, scalable, and accessible platform for single-molecule proteomic studies and applications.

Learning curves for transradial access versus transfemoral access in diagnostic cerebral angiography: a case series.

BACKGROUND: The perception of a steep learning curve associated with transradial access has resulted in its limited adoption in neurointervention despite the demonstrated benefits, including decreased access-site complications. OBJECTIVE: To compare learning curves of transradial versus transfemoral diagnostic cerebral angiograms obtained by five neurovascular fellows as primary operator. METHODS: The first 100-150 consecutive transradial and transfemoral angiographic scans performed by each fellow between July 2017 and March 2020 were identified. Mean fluoroscopy time per artery injected (angiographic efficiency) was calculated as a marker of technical proficiency and compared for every 25 consecutive procedures performed (eg, 1-25, 26-50, 51-75). RESULTS: We identified 1242 diagnostic angiograms, 607 transradial and 635 transfemoral. The radial cohort was older (64.3 years vs 62.3 years, p=0.01) and demonstrated better angiographic efficiency (3.4 min/vessel vs 3.7 min/vessel, p=0.03). For three fellows without previous endovascular experience, proficiency was obtained between 25 and 50 transfemoral angiograms. One fellow achieved proficiency after performing 25-50 transradial angiograms; and the two other fellows, in <25 transradial angiograms. The two fellows with previous experience had flattened learning curves for both access types. Two patients experienced transient neurologic symptoms postprocedure. Transradial angiograms were associated with significantly fewer access-site complications (3/607, 0.5% vs 22/635, 3.5%, p<0.01). Radial-to-femoral conversion occurred in 1.2% (7/607); femoral-to-radial conversion occurred in 0.3% (2/635). Over time, the proportion of transradial angiographic procedures increased. CONCLUSION: Technical proficiency improved significantly over time for both access types, typically requiring between 25 and 50 diagnostic angiograms to achieve asymptomatic improvement in efficiency. Reduced access-site complications and decreased fluoroscopy time were benefits associated with transradial angiography.

Sovereign immunity and its implications for neurosurgery.

The proportion of neurosurgeons facing a malpractice suit each year is highest among all medical and surgical specialties. It is critical for neurosurgeons to understand local malpractice laws because they vary among states. Sovereign immunity, as described in the 11th constitutional amendment, provides absolute immunity to states from being sued by their residents and by other states. A state may waive its sovereign immunity, however, and substitute itself as the defendant in place of a state-employed physician in the court of law. This means that a physician working for a state-funded hospital may not be liable to a malpractice suit. Further provisions of the law allow the state not to pay indemnity beyond a certain limit, which discourages plaintiff attorneys from pursuing indemnity charges against physicians working for state-funded institutions. In this review, the authors describe the concept of sovereign immunity and its implications for the practice of neurosurgery.

A massively parallel barcoded sequencing pipeline enables generation of the first ORFeome and interactome map for rice.

Systematic mappings of protein interactome networks have provided invaluable functional information for numerous model organisms. Here we develop PCR-mediated Linkage of barcoded Adapters To nucleic acid Elements for sequencing (PLATE-seq) that serves as a general tool to rapidly sequence thousands of DNA elements. We validate its utility by generating the ORFeome for Oryza sativa covering 2,300 genes and constructing a high-quality protein-protein interactome map consisting of 322 interactions between 289 proteins, expanding the known interactions in rice by roughly 50%. Our work paves the way for high-throughput profiling of protein-protein interactions in a wide range of organisms.

Interactome INSIDER: a structural interactome browser for genomic studies.

We present Interactome INSIDER, a tool to link genomic variant information with structural protein-protein interactomes. Underlying this tool is the application of machine learning to predict protein interaction interfaces for 185,957 protein interactions with previously unresolved interfaces in human and seven model organisms, including the entire experimentally determined human binary interactome. Predicted interfaces exhibit functional properties similar to those of known interfaces, including enrichment for disease mutations and recurrent cancer mutations. Through 2,164 de novo mutagenesis experiments, we show that mutations of predicted and known interface residues disrupt interactions at a similar rate and much more frequently than mutations outside of predicted interfaces. To spur functional genomic studies, Interactome INSIDER (http://interactomeinsider.yulab.org) enables users to identify whether variants or disease mutations are enriched in known and predicted interaction interfaces at various resolutions. Users may explore known population variants, disease mutations, and somatic cancer mutations, or they may upload their own set of mutations for this purpose.

mutation3D: Cancer Gene Prediction Through Atomic Clustering of Coding Variants in the Structural Proteome.

A new algorithm and Web server, mutation3D (http://mutation3d.org), proposes driver genes in cancer by identifying clusters of amino acid substitutions within tertiary protein structures. We demonstrate the feasibility of using a 3D clustering approach to implicate proteins in cancer based on explorations of single proteins using the mutation3D Web interface. On a large scale, we show that clustering with mutation3D is able to separate functional from nonfunctional mutations by analyzing a combination of 8,869 known inherited disease mutations and 2,004 SNPs overlaid together upon the same sets of crystal structures and homology models. Further, we present a systematic analysis of whole-genome and whole-exome cancer datasets to demonstrate that mutation3D identifies many known cancer genes as well as previously underexplored target genes. The mutation3D Web interface allows users to analyze their own mutation data in a variety of popular formats and provides seamless access to explore mutation clusters derived from over 975,000 somatic mutations reported by 6,811 cancer sequencing studies. The mutation3D Web interface is freely available with all major browsers supported.

A Proteome-wide Fission Yeast Interactome Reveals Network Evolution Principles from Yeasts to Human.

Here, we present FissionNet, a proteome-wide binary protein interactome for S. pombe, comprising 2,278 high-quality interactions, of which ∼ 50% were previously not reported in any species. FissionNet unravels previously unreported interactions implicated in processes such as gene silencing and pre-mRNA splicing. We developed a rigorous network comparison framework that accounts for assay sensitivity and specificity, revealing extensive species-specific network rewiring between fission yeast, budding yeast, and human. Surprisingly, although genes are better conserved between the yeasts, S. pombe interactions are significantly better conserved in human than in S. cerevisiae. Our framework also reveals that different modes of gene duplication influence the extent to which paralogous proteins are functionally repurposed. Finally, cross-species interactome mapping demonstrates that coevolution of interacting proteins is remarkably prevalent, a result with important implications for studying human disease in model organisms. Overall, FissionNet is a valuable resource for understanding protein functions and their evolution.

BISQUE: locus- and variant-specific conversion of genomic, transcriptomic and proteomic database identifiers.

UNLABELLED: Biological sequence databases are integral to efforts to characterize and understand biological molecules and share biological data. However, when analyzing these data, scientists are often left holding disparate biological currency-molecular identifiers from different databases. For downstream applications that require converting the identifiers themselves, there are many resources available, but analyzing associated loci and variants can be cumbersome if data is not given in a form amenable to particular analyses. Here we present BISQUE, a web server and customizable command-line tool for converting molecular identifiers and their contained loci and variants between different database conventions. BISQUE uses a graph traversal algorithm to generalize the conversion process for residues in the human genome, genes, transcripts and proteins, allowing for conversion across classes of molecules and in all directions through an intuitive web interface and a URL-based web service. AVAILABILITY AND IMPLEMENTATION: BISQUE is freely available via the web using any major web browser (http://bisque.yulab.org/). Source code is available in a public GitHub repository (https://github.com/hyulab/BISQUE). CONTACT: haiyuan.yu@cornell.edu SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Studying Autism in Context.

Studying autism genes in the context of the protein complexes to which they belong illustrates the potential of network-centric approaches for understanding complex genetic disease.

Exploring mechanisms of human disease through structurally resolved protein interactome networks.

The study of the molecular basis of human disease has gained increasing attention over the past decade. With significant improvements in sequencing efficiency and throughput, a wealth of genotypic data has become available. However the translation of this information into concrete advances in diagnostic and clinical setups has proved far more challenging. Two major reasons for this are the lack of functional annotation for genomic variants and the complex nature of genotype-to-phenotype relationships. One fundamental approach to bypass these issues is to examine the effects of genetic variation at the level of proteins as they are directly involved in carrying out biological functions. Within the cell, proteins function by interacting with other proteins as a part of an underlying interactome network. This network can be determined using interactome mapping - a combination of high-throughput experimental toolkits and curation from small-scale studies. Integrating structural information from co-crystals with the network allows generation of a structurally resolved network. Within the context of this network, the structural principles of disease mutations can be examined and used to generate reliable mechanistic hypotheses regarding disease pathogenesis.

INstruct: a database of high-quality 3D structurally resolved protein interactome networks.

UNLABELLED: INstruct is a database of high-quality, 3D, structurally resolved protein interactome networks in human and six model organisms. INstruct combines the scale of available high-quality binary protein interaction data with the specificity of atomic-resolution structural information derived from co-crystal evidence using a tested interaction interface inference method. Its web interface is designed to allow for flexible search based on standard and organism-specific protein and gene-naming conventions, visualization of protein architecture highlighting interaction interfaces and viewing and downloading custom 3D structurally resolved interactome datasets. AVAILABILITY: INstruct is freely available on the web at http://instruct.yulab.org with all major browsers supported.

Confronting deep moral disagreement: the President's Council on Bioethics, moral status, and human embryos.

The report of the President's Council on Bioethics, Human Cloning and Human Dignity, addresses the central ethical, political, and policy issue in human embryonic stem cell research: the moral status of extracorporeal human embryos. The Council members were in sharp disagreement on this issue and essentially failed to adequately engage and respectfully acknowledge each others' deepest moral concerns, despite their stated commitment to do so. This essay provides a detailed critique of the two extreme views on the Council (i.e., embryos have full moral status or they have none at all) and then gives theoretical grounding for our judgment about the intermediate moral status of embryos. It also supplies an account of how to address profound moral disagreements in the public arena, especially by way of constructing a middle ground that deliberately pays sincere respect to the views of those with whom it has deep disagreements.

Digital subtraction radiographic analysis of GTR in human intrabony defects.

The relationship between radiographic crestal alveolar bone mass and changes in clinical periodontal attachment level following guided tissue regeneration (GTR) was evaluated in this retrospective study. A total of 12 intrabony two- or three-walled defects and 12 adjacent nondiseased proximal sites in 10 nonsmoking adult subjects received subgingival debridement and GTR using resorbable (Resolut; five sites) or nonresorbable (Gore-Tex; seven sites) barrier membranes. At a mean of 48.8 months posttreatment, clinical periodontal attachment level alterations were measured, and crestal alveolar bone mass changes on digital subtraction radiographic images derived from serial periapical radiographs were analyzed to correct for between-film geometric and contrast density differences. Intrabony defects exhibited a mean clinical periodontal attachment level gain of 2.3 +/- 0.4 mm, in contrast to a mean loss of 0.5 +/- 0.2 mm in adjacent nondiseased interproximal sites. Digital subtraction radiography revealed an increase in crestal alveolar bone mass at all intrabony sites treated with GTR and a decrease in three of the adjacent nondiseased sites. Site-based analysis yielded an odds ratio of 36 (P < .001) for the association between radiographic increases in crestal alveolar bone mass and clinical periodontal attachment level gains of > or = 2 mm. These results suggest a strong concordance between digital subtraction radiographic assessments of crestal alveolar bone mass and clinical periodontal attachment level in evaluating the long-term effects of GTR at human interproximal intrabony defects.