In bioinformatics, pathway analysis is an important approach for understanding the role and interaction of proteins and other molecular entities in various biological pathways or processes. Reactome is an open-source database of human biological pathways designed to perform pathway analysis.
Here, we will discuss the steps to perform pathway analysis using Reactome, covering essential features, how to query data, and some tips on interpreting the result.
What is Reactome?
Reactome is a free, peer-reviewed, and curated online pathway database, that hosts a vast range of pathways and reactions, primarily of humans and also includes pathway data of over 90 other organisms. Reactome’s data is frequently updated, curated by experimental data, and supported by references to relevant literature. The pathway data in the Reactome includes all reactions and interactions related to metabolism, gene regulation, and immune response. Each pathway has a series of reactions annotated with protein or gene information, location and inter-pathway interactions.
Navigating Reactome’s interface:
Access Reactome’s Database:
To start Reactome, visit the Reactome website. A simple, user-friendly interface with prominent sections for search analysis and visualization will appear before you. No account is required for exploring this database however creating an account can allow you to save projects and analysis.
Search for protein:
In the main search bar, you can enter your protein IDs or gene names and keywords related to a specific biological pathway. For example, searching “TP53” will bring up all pathways involving the tumor suppressor protein p53. After submitting the search term, you will see results having terms, such as “Pathways”, “Reactions” and “Molecules”. Click on the Pathways tab to access detailed pathway information.
Exploring Pathways:
Each pathway has a detailed description page in Reactome and when you open a pathway, you will see an interactive pathway diagram along with reactions and participants. The pathway diagram presented by Reactome will be in a color-coded layout displaying the molecules involved. In the “Reaction” section you will see information regarding the type of reaction, each reaction will be further linked to references and external databases for cross-referencing like PubMed and UniProt. Reactome also offers a pathway browser to explore connections between different pathways. The browser uses icons like “star” to indicate key proteins and “zoom” icons for sub-pathways.
Conducting Pathway Analysis with Reactome:
Pathway Enrichment Analysis Tool:
For pathway analysis go to Analysis Tools at Reactome’s interface and then paste or upload a list of gene or protein IDs. Reactome accepts various formats, including Uniprot or Ensembl IDs. After submitting Reactome matches your list to pathway databases and calculates the P-value of each pathway based on the enrichment of proteins from your input list. Pathways with high enrichment scores suggest that your protein list strongly correlates with those pathways. The result table will show pathways with columns for “Pathway ID”, “Names”, “Entities found”, P-vale and “Entities ratio”.
Pathway Browser:
Pathway browser provides an interactive way to visualize pathways. This tool provides a detailed view of specific molecular interactions. You can zoom in and out to view the entire pathway or focus on specific interactions. You can also see additional details by clicking on proteins, molecules, or reactions. Reactome also allows users to examine how a pathway is connected to other pathways in the cell, thus providing a wider understanding.
Functional Interaction (FI) Network:
FI network is a Reactome tool for building a protein-protein interaction network. You can create an FI network by uploading a list of genes or proteins. The resulting network provides protein-protein interactions and key relationships between them. The nodes represent proteins, while the edges represent interactions, with different colors indicating the type of interaction between them. Furthermore, each interaction has supporting references for additional information.
Advanced features for Pathway Analysis:
Some advanced features of Reactome include;
· Reactome enables users to view pathways across species by utilizing orthologous links. Selecting species from the dropdown menu allows you to compare a pathway in humans to its homologs in other organisms, which is important for evolutionary studies or model organism research.
· Reactome also provides several data export options like SBML, BioPAX, and CVS, making it easy to integrate pathways in other analysis tools.
· You can also overlay or superimpose your experimental data on pathway diagrams which will allow you to interpret changes such as mutation rates and differential expression.
· Reactome’s pathway data also provides information about disease variations and their effects on biological processes. This feature is useful for researchers who are looking for how specific mutations alter protein functions within pathways, thus offering insight into the molecular basis of a disease.
Reactome is an invaluable tool for advanced research in the fields of Proteomics, Genomics, and Systems biology. Reactome can boost the depth and precision of your protein pathway analysis whether you are studying causes of disease, examining therapeutic targets, or conducting comparative biology.
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