Dataset Information

Direction pathway analysis of large-scale proteomics data reveals novel features of the insulin action pathway.

ABSTRACT:

Motivation

With the advancement of high-throughput techniques, large-scale profiling of biological systems with multiple experimental perturbations is becoming more prevalent. Pathway analysis incorporates prior biological knowledge to analyze genes/proteins in groups in a biological context. However, the hypotheses under investigation are often confined to a 1D space (i.e. up, down, either or mixed regulation). Here, we develop direction pathway analysis (DPA), which can be applied to test hypothesis in a high-dimensional space for identifying pathways that display distinct responses across multiple perturbations.

Results

Our DPA approach allows for the identification of pathways that display distinct responses across multiple perturbations. To demonstrate the utility and effectiveness, we evaluated DPA under various simulated scenarios and applied it to study insulin action in adipocytes. A major action of insulin in adipocytes is to regulate the movement of proteins from the interior to the cell surface membrane. Quantitative mass spectrometry-based proteomics was used to study this process on a large-scale. The combined dataset comprises four separate treatments. By applying DPA, we identified that several insulin responsive pathways in the plasma membrane trafficking are only partially dependent on the insulin-regulated kinase Akt. We subsequently validated our findings through targeted analysis of key proteins from these pathways using immunoblotting and live cell microscopy. Our results demonstrate that DPA can be applied to dissect pathway networks testing diverse hypotheses and integrating multiple experimental perturbations.

Availability and implementation

The R package 'directPA' is distributed from CRAN under GNU General Public License (GPL)-3 and can be downloaded from: http://cran.r-project.org/web/packages/directPA/index.html

Contact

jean.yang@sydney.edu.au

Supplementary information

Supplementary data are available at Bioinformatics online.

SUBMITTER: Yang P

PROVIDER: S-EPMC3957074 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature

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Publications

Direction pathway analysis of large-scale proteomics data reveals novel features of the insulin action pathway.

Yang Pengyi P Yang Pengyi P Patrick Ellis E Tan Shi-Xiong SX Fazakerley Daniel J DJ Burchfield James J Gribben Christopher C Prior Matthew J MJ James David E DE Hwa Yang Yee Y

Bioinformatics (Oxford, England) 20131027 6

<h4>Motivation</h4>With the advancement of high-throughput techniques, large-scale profiling of biological systems with multiple experimental perturbations is becoming more prevalent. Pathway analysis incorporates prior biological knowledge to analyze genes/proteins in groups in a biological context. However, the hypotheses under investigation are often confined to a 1D space (i.e. up, down, either or mixed regulation). Here, we develop direction pathway analysis (DPA), which can be applied to t ...[more]

PMID: 24167158

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Project description:BackgroundPhosphorylation is the most prevalent post-translational modification on eukaryotic proteins. Multisite phosphorylation enables a specific combination of phosphosites to determine the speed, specificity and duration of biological response. Until recent years, the lack of high quality data limited the possibility for analyzing the properties of phosphorylation at the proteome scale and in the context of a wide range of conditions. Thanks to advances of mass spectrometry technologies, thousands of phosphosites from in-vivo experiments were identified and archived in the public domain. Such resource is appropriate to derive an unbiased view on the phosphosites properties in eukaryotes and on their functional relevance.ResultsWe present statistically rigorous tests on the spatial and functional properties of a collection of approximately 70,000 reported phosphosites. We show that the distribution of phosphosites positioning along the protein tends to occur as dense clusters of Serine/Threonines (pS/pT) and between Serine/Threonines and Tyrosines, but generally not as much between Tyrosines (pY) only. This phenomenon is more ubiquitous than anticipated and is pertinent for most eukaryotic proteins: for proteins with > or = 2 phosphosites, 54% of all pS/pT sites are within 4 amino acids of another site. We found a strong tendency for clustered pS/pT to be activated by the same kinase. Large-scale analyses of phosphopeptides are thus consistent with a cooperative function within the cluster.ConclusionsWe present evidence supporting the notion that clusters of pS/pT but generally not pY should be considered as the elementary building blocks in phosphorylation regulation. Indeed, closely positioned sites tend to be activated by the same kinase, a signal that overrides the tendency of a protein to be activated by a single or only few kinases. Within these clusters, coordination and positional dependency is evident. We postulate that cellular regulation takes advantage of such design. Specifically, phosphosite clusters may increase the robustness of the effectiveness of phosphorylation-dependent response.ReviewersReviewed by Joel Bader, Frank Eisenhaber, Emmanuel Levy (nominated by Sarah Teichmann). For the full reviews, please go to the Reviewers' comments section.

Dataset Information

Direction pathway analysis of large-scale proteomics data reveals novel features of the insulin action pathway.

Motivation

Results

Availability and implementation

Contact

Supplementary information

Publications

Direction pathway analysis of large-scale proteomics data reveals novel features of the insulin action pathway.

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