Ruth Isserlin

Integration of biological networks and gene expression data using Cytoscape

Cytoscape is a free software package for visualizing, modeling and analyzing molecular and genetic interaction networks. This protocol explains how to use Cytoscape to analyze the results of mRNA expression profiling, and other functional genomics and proteomics experiments, in the context of an interaction network obtained for genes of interest. Five major steps are described: (i) obtaining a gene or protein network, (ii) displaying the network using layout algorithms, (iii) integrating with gene expression and other functional attributes, (iv) identifying putative complexes and functional modules and (v) identifying enriched Gene Ontology annotations in the network. These steps provide a broad sample of the types of analyses performed by Cytoscape.

A draft map of the human proteome

The availability of human genome sequence has transformed biomedical research over the past decade. However, an equivalent map for the human proteome with direct measurements of proteins and peptides does not exist yet. Here we present a draft map of the human proteome using high-resolution Fourier-transform mass spectrometry. In-depth proteomic profiling of 30 histologically normal human samples, including 17 adult tissues, 7 fetal tissues and 6 purified primary haematopoietic cells, resulted in identification of proteins encoded by 17,294 genes accounting for approximately 84% of the total annotated protein-coding genes in humans. A unique and comprehensive strategy for proteogenomic analysis enabled us to discover a number of novel protein-coding regions, which includes translated pseudogenes, non-coding RNAs and upstream open reading frames. This large human proteome catalogue (available as an interactive web-based resource at http://www.humanproteomemap.org) will complement available human genome and transcriptome data to accelerate biomedical research in health and disease.

Enrichment Map: A Network-Based Method for Gene-Set Enrichment Visualization and Interpretation

Background Gene-set enrichment analysis is a useful technique to help functionally characterize large gene lists, such as the results of gene expression experiments. This technique finds functionally coherent gene-sets, such as pathways, that are statistically over-represented in a given gene list. Ideally, the number of resulting sets is smaller than the number of genes in the list, thus simplifying interpretation. However, the increasing number and redundancy of gene-sets used by many current enrichment analysis software works against this ideal. Principal Findings To overcome gene-set redundancy and help in the interpretation of large gene lists, we developed “Enrichment Map”, a network-based visualization method for gene-set enrichment results. Gene-sets are organized in a network, where each set is a node and edges represent gene overlap between sets. Automated network layout groups related gene-sets into network clusters, enabling the user to quickly identify the major enriched functional themes and more easily interpret the enrichment results. Conclusions Enrichment Map is a significant advance in the interpretation of enrichment analysis. Any research project that generates a list of genes can take advantage of this visualization framework. Enrichment Map is implemented as a freely available and user friendly plug-in for the Cytoscape network visualization software (http://baderlab.org/Software/EnrichmentMap/).

The Biomolecular Interaction Network Database and related tools 2005 update

The Biomolecular Interaction Network Database (BIND) (http://bind.ca) archives biomolecular interaction, reaction, complex and pathway information. Our aim is to curate the details about molecular interactions that arise from published experimental research and to provide this information, as well as tools to enable data analysis, freely to researchers worldwide. BIND data are curated into a comprehensive machine-readable archive of computable information and provides users with methods to discover interactions and molecular mechanisms. BIND has worked to develop new methods for visualization that amplify the underlying annotation of genes and proteins to facilitate the study of molecular interaction networks. BIND has maintained an open database policy since its inception in 1999. Data growth has proceeded at a tremendous rate, approaching over 100 000 records. New services provided include a new BIND Query and Submission interface, a Standard Object Access Protocol service and the Small Molecule Interaction Database (http://smid.blueprint.org) that allows users to determine probable small molecule binding sites of new sequences and examine conserved binding residues.

Delineation of two clinically and molecularly distinct subgroups of posterior fossa ependymoma.

Despite the histological similarity of ependymomas from throughout the neuroaxis, the disease likely comprises multiple independent entities, each with a distinct molecular pathogenesis. Transcriptional profiling of two large independent cohorts of ependymoma reveals the existence of two demographically, transcriptionally, genetically, and clinically distinct groups of posterior fossa (PF) ependymomas. Group A patients are younger, have laterally located tumors with a balanced genome, and are much more likely to exhibit recurrence, metastasis at recurrence, and death compared with Group B patients. Identification and optimization of immunohistochemical (IHC) markers for PF ependymoma subgroups allowed validation of our findings on a third independent cohort, using a human ependymoma tissue microarray, and provides a tool for prospective prognostication and stratification of PF ependymoma patients.

Too many roads not taken

Most protein research focuses on those known before the human genome was mapped. Work on the slew discovered since, urge Aled M. Edwards and his colleagues.

PSICQUIC and PSISCORE: accessing and scoring molecular interactions

To study proteins in the context of a cellular system, it is essential that the molecules with which a protein interacts are identified and the functional consequence of each interaction is understood. A plethora of resources now exist to capture molecular interaction data from the many laboratories generating…

A Call for Systematic Research on Solute Carriers

Solute carrier (SLC) membrane transport proteins control essential physiological functions, including nutrient uptake, ion transport, and waste removal. SLCs interact with several important drugs, and a quarter of the more than 400 SLC genes are associated with human diseases. Yet, compared to other gene families of similar stature, SLCs are relatively understudied. The time is right for a systematic attack on SLC structure, specificity, and function, taking into account kinship and expression, as well as the dependencies that arise from the common metabolic space.

Comparative Proteomics Profiling of a Phospholamban Mutant Mouse Model of Dilated Cardiomyopathy Reveals Progressive Intracellular Stress Responses

Defective mobilization of Ca2+ by cardiomyocytes can lead to cardiac insufficiency, but the causative mechanisms leading to congestive heart failure (HF) remain unclear. In the present study we performed exhaustive global proteomics surveys of cardiac ventricle isolated from a mouse model of cardiomyopathy overexpressing a phospholamban mutant, R9C (PLN-R9C), and exhibiting impaired Ca2+ handling and death at 24 weeks and compared them with normal control littermates. The relative expression patterns of 6190 high confidence proteins were monitored by shotgun tandem mass spectrometry at 8, 16, and 24 weeks of disease progression. Significant differential abundance of 593 proteins was detected. These proteins mapped to select biological pathways such as endoplasmic reticulum stress response, cytoskeletal remodeling, and apoptosis and included known biomarkers of HF (e.g. brain natriuretic peptide/atrial natriuretic factor and angiotensin-converting enzyme) and other indicators of presymptomatic functional impairment. These altered proteomic profiles were concordant with cognate mRNA patterns recorded in parallel using high density mRNA microarrays, and top candidates were validated by RT-PCR and Western blotting. Mapping of our highest ranked proteins against a human diseased explant and to available data sets indicated that many of these proteins could serve as markers of disease. Indeed we showed that several of these proteins are detectable in mouse and human plasma and display differential abundance in the plasma of diseased mice and affected patients. These results offer a systems-wide perspective of the dynamic maladaptions associated with impaired Ca2+ homeostasis that perturb myocyte function and ultimately converge to cause HF.

A Lentiviral Functional Proteomics Approach Identifies Chromatin Remodeling Complexes Important for the Induction of Pluripotency

Protein complexes and protein-protein interactions are essential for almost all cellular processes. Here, we establish a mammalian affinity purification and lentiviral expression (MAPLE) system for characterizing the subunit compositions of protein complexes. The system is flexible (i.e. multiple N- and C-terminal tags and multiple promoters), is compatible with GatewayTM cloning, and incorporates a reference peptide. Its major advantage is that it permits efficient and stable delivery of affinity-tagged open reading frames into most mammalian cell types. We benchmarked MAPLE with a number of human protein complexes involved in transcription, including the RNA polymerase II-associated factor, negative elongation factor, positive transcription elongation factor b, SWI/SNF, and mixed lineage leukemia complexes. In addition, MAPLE was used to identify an interaction between the reprogramming factor Klf4 and the Swi/Snf chromatin remodeling complex in mouse embryonic stem cells. We show that the SWI/SNF catalytic subunit Smarca2/Brm is up-regulated during the process of induced pluripotency and demonstrate a role for the catalytic subunits of the SWI/SNF complex during somatic cell reprogramming. Our data suggest that the transcription factor Klf4 facilitates chromatin remodeling during reprogramming.