APOSTL

Overview

Automated Processing of SAINT Templated Layouts (APOSTL) is a Galaxy-based analysis pipeline for reproducible analysis of affinity proteomics (AP) data. APOSTL utilizes Significance Analysis of INTeractome (SAINT), a popular command-line software for analyzing AP data. APOSTL can process AP results from MaxQuant, Scaffold, PeptideShaker, or any software that can export mzIdentML. APOSTL can create a number of publication-quality visualizations including interactive bubble plots, protein-protein interaction networks, and pathway enrichment/gene ontology plots. All visualizations are accomplished through Shiny, an interactive and open-source visualization package for the R programming language. APOSTL is open-source software released under GPLv3, and it is freely available on the Galaxy Tool Shed and GitHub.

Features

Multiple Formats

Pre-process your data from Scaffold, MaxQuant, PeptideShaker or mzIdentML

Interactive Plotting

Analyze your data in real time

Publication Quality Figures

Save any image as publication quality figure in multiple formats

GalaxyP

Interface with GalaxyP for an entire proteomics workflow in Galaxy!

How It Works

Pre-processing

APOSTL is able to recognize the Scaffold Samples Report file, the peptides.txt file in the Maxquant txt output folder, the PeptideShaker Protein Report file, or the output from other tools in mzIdentML format. Scaffold, MaxQuant, and PeptideShaker output files must be saved as tab delimited text, and no other modifications should be made to these files. Using the Bait Create option in the SAINT Pre-processing tool allows you to quickly create a bait.txt file. It is important that the individual bait names exactly match the bait names within your output from the above software. This tool also includes an option to automatically generate a prey.txt file.

Note: See the SaintExpress manual for more information about file formatting requirements.

When making a prey.txt file, APOSTL queries a user provided FASTA database in order to extract protein amino acid lengths and gene names. This process may take several minutes. Some users may want to run SAINTexpress using the same data set while changing which baits are considered test or control. It is useful to toggle Make Prey off in order to save time by circumventing this step as the same prey file can be used for both SAINTexpress runs.

Post-processing

Once SAINTexpress has been run, APOSTL is able to read the resulting list.txt file. From here APOSTL does a number of things:

1) APOSTL calculates the normalized spectral abundance factor (NSAF) values for each prey based on the average spectra observed for each bait

2) Optionally, APOSTL calculates the probability of a specific interaction based on prey prevalence in the CRAPome.

Other Analysis Options

1) Protein-protein interaction network query using ConsensusPathDB following filtering. Allows for easy Cytoscape import for visualization using the import network from file option.

2) Dot plot and clustering analysis from the the Prohits Visualization Tool.

3) Galaxy passes results to Shiny for interactive visualizations (Shiny is configured on this server).

Quick Start Guide

This is a quick start guide more appropriate for users familiar with Galaxy. Before beginning, it is recommended that first-time users familiarize themselves with how SAINTexpress works and the contents of required files (link to manual). A long format walkthrough with screen captures can be found on GitHub.

1) Register: Please create a free account by clicking on the User menu button and selecting Register. Accounts are created immediately and can be used immediately.

2) Once signed in as a user, click on the Shared Data menu button and select Published Histories.

This will allow you to view sample EGFR LFQ Scaffold and MaxQuant results files along with every step taken to produce the results.
These Histories can be imported into the History Panel on the Home Page by clicking on them and selecting Import History in the upper right corner.
The last step in each History (found at the very top of the History Panel) is an APOSTL-generated output that features interactive analysis with Shiny. To access the interactive data analysis, simply click View Data button (which looks like an eye) and clicking the APOSTL Interactive Analysis link.

3) To access workflows that can be used on user-provided datasets, click on the Shared Data menu button and select Published Workflows.

Choose the appropriate workflow for Scaffold or MaxQuant data and click Import Workflow.
Upload output files from supported software: a Scaffold-formatted Samples Report output file or MaxQuant peptides.txt output file, and a bait file (which can be created using the Bait Create within the preprocessing tool).
Click the Workflow button at the top of the page and select the imported workflow. Select the appropriate files in the drop down menu and press Run Workflow.

Welcome