GProX, a user-friendly platform for bioinformatics analysis and visualization of quantitative proteomics data

Abstract

Recent technological advances have made it possible to identify and quantify thousands of proteins in a single proteomics experiment. As a result of these developments, the analysis of data has become the bottleneck of proteomics experiment. To provide the proteomics community with a user-friendly platform for comprehensive analysis, inspection and visualization of quantitative proteomics data we developed the Graphical Proteomics Data Explorer (GProX)(1). The program requires no special bioinformatics training, as all functions of GProX are accessible within its graphical user-friendly interface which will be intuitive to most users. Basic features facilitate the uncomplicated management and organization of large data sets and complex experimental setups as well as the inspection and graphical plotting of quantitative data. These are complemented by readily available high-level analysis options such as database querying, clustering based on abundance ratios, feature enrichment tests for e.g. GO terms and pathway analysis tools. A number of plotting options for visualization of quantitative proteomics data is available and most analysis functions in GProX create customizable high quality graphical displays in both vector and bitmap formats. The generic import requirements allow data originating from essentially all mass spectrometry platforms, quantitation strategies and software to be analyzed in the program. GProX represents a powerful approach to proteomics data analysis providing proteomics experimenters with a toolbox for bioinformatics analysis of quantitative proteomics data. The program is released as open-source and can be freely downloaded from the project webpage at http://gprox.sourceforge.net.

Fig. 1.

Overview of the GProX Structure. GProX accepts as input a tab- or character-delimited file containing as a minimum only protein accession keys and quantitative information. Initial analysis session setup is done via a simple input wizard, which also supports defining the experimental design. The user interface provides access to all functions and uses the R-environment for advanced analyses and generation of graphics. All files associated with an analysis session are saved locally and from the session file a stored session can be opened for continued analysis.

Fig. 2.

GProX User Interface. The multiple document user interface of GProX display, in dedicated windows, the data tables on which the analysis is performed, analysis results and all figures produced in the analysis session. All functions inside the program are accessible via a ribbon control (see also supplemental Fig. S1).

Fig. 3.

Analysis steps, Protein Dynamics and Database Information within GProX. A, Overview of the analysis steps performed to exemplify the application of GProX for analyzing quantitative proteomics data. B, Plots of the ratios of one or more proteins and experiments can readily be requested to acquire a rapid overview of protein regulation. In this case, each panel includes regulation profiles from different experiments for individual proteins. C, IPI- and UniProt database sheets can be readily requested for selected proteins to immediately provide the user with available information stored in these databases. Furthermore, these database entries link out to other primary resources or higher-level annotations.

Fig. 4.

Visualization of Data Distributions in GProX. Graphical overviews of the distribution of protein abundance ratios in the form of histograms (A), box plots (B), scatter plots (C), density scatter plots (D), and heatmaps (E). Individually or in combination, the visualizations provide a summary of the cumulative properties of the data, which is indispensable e.g. for comparing experiments or assessing the reproducibility of replicate experiments.

Fig. 5.

Clustering and Enrichment Analysis in GProX. A, Unsupervised clustering by fuzzy-c means of the example data revealing distinct patterns of regulation. B, Enrichment analysis for GO biological process terms over-represented in each cluster (see fig. 5A). From this analysis it is clear, that the different modes of regulation summarized in (A) correspond also to distinct cellular processes.

Fig. 6.

GProX Pathway Analysis of the Cellular Response to EGF Stimulation. The regulation after 30 min stimulation with EGF was mapped to the ErbB KEGG signaling pathway. Nodes corresponding to proteins identified in the example data are color-coded according to regulation so up-regulated proteins are colored in shades of red, down-regulated proteins are colored in shades of blue and not regulated proteins are gray. This analysis illustrate that after 30 min of EGF stimulation the proteins upstream in the pathway have returned to the control state or are down-regulated while downstream effector proteins remain up-regulated.