In gaelfortin/phosphogo: Analyze Your Phosphoproteomic Data

knitr::opts_chunk$set(
  collapse = TRUE,
  comment = "#>"
)

This vignette highlights essential functions of phosphogo and their use. For details on phosphogo installation, refer to the GitHub repository of phosphogo.

You can also use the web interface of pĥosphogo with:

library(phosphogo)
phosphogoApp()

First, load phosphogo and create a new folder to output all files:

library(phosphogo)
dir.create('myexperiment', showWarnings = FALSE) 

1. Generate NetworKIN and IV-KEA input from phosphoproteomic data

# Human phosphoproteomic data
phospho_input(
  phospho_file = "phospho_human.xlsx",
  phosphosites_column = "PhosphoSite",
  log2_column = "Log2",
  fdr_column = "Adj. Pvalue",
  species = 'hsa',
  output_folder = 'myexperiment/'
)

# Mouse phosphoproteomic data
phospho_input(
  phospho_file = "phospho_mouse.xlsx",
  phosphosites_column = "ProteinID-Phospho:Site",
  log2_column = "Log2",
  fdr_column = "Adj. Pvalue",
  species = 'mmu',
  output_folder = 'myexperiment/'
)

1. Run NetworKIN NetworKIN predictions were pre-computed on all human phosphosites. Thus, phosphogo fetches prediction for the given list of phosphosites.

run_networkin(
  input_file = "networKIN_input.res",
  output_folder = "myexperiment/"
)

1. Verify NetworKIN prediction scores distribution

networkin_qc(predictions_file = 'networkin_output.csv',
             output_folder = 'myexperiment/')

1. NetworKIN Fisher exact test The following Fisher exact test can be performed with NetworKIN and IV-KEA:
2. Up reg vs down reg
3. Up reg vs everything (signif and not signif)
4. Down reg vs everything (signif and not signif)

perform_Fisher_exact_test(predictions_file = 'networkin_output.csv',
                              predictions = "networkin",
                              output_folder = "myexperiment/",
                              FC_threshold = 1.2) 

1. Generate volcano plots Each Fisher exact test can be used to generate a volcano plot.

make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_networkin.csv', 
                  odds_ratio = up_vs_down_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = up_vs_down_FDR,
                  graph_title = "Kinases prediction enrichment (NetworKIN) up vs down",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = "myexperiment/",
                  file_name = "up_down_volcano_plot_networkin.pdf")
make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_networkin.csv', 
                  odds_ratio = up_vs_tot_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = up_vs_tot_FDR,
                  graph_title = "Kinases prediction enrichment (NetworKIN) up vs tot",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = 'myexperiment/',
                  file_name = "up_tot_volcano_plot_networkin.pdf")
make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_networkin.csv', 
                  odds_ratio = down_vs_tot_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = down_vs_tot_FDR,
                  graph_title = "Kinases prediction enrichment (NetworKIN) down vs tot",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = 'myexperiment/',
                  file_name = "down_tot_volcano_plot_networkin.pdf")

1. Perform IV-KEA

perform_ivkea(clean_phospho_file = 'phospho_clean.csv',
                          output_folder = 'myexperiment/')

Run Fisher Exact test

perform_Fisher_exact_test(top_predictions_file = 'ivkea_predictions.csv',
                              predictions = "ivkea",
                              output_folder = 'myexperiment/',
                              FC_threshold = 1.2)

Plot results on volcano plots

make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_ivkea.csv', 
                  odds_ratio = up_vs_down_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = up_vs_down_FDR,
                  graph_title = "Kinases prediction enrichment (IV-KEA) up vs down",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = 'myexperiment/',
                  file_name = "up_down_volcano_plot_ivkea.pdf")
make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_ivkea.csv', 
                  odds_ratio = up_vs_tot_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = up_vs_tot_FDR,
                  graph_title = "Kinases prediction enrichment (IV-KEA) up vs tot",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = 'myexperiment/',
                  file_name = "up_tot_volcano_plot_ivkea.pdf")
make_volcano_plot(kinase_enrichment_file = 'kinase_enrichment_ivkea.csv', 
                  odds_ratio = down_vs_tot_odds_ratio,
                  FDR_cutoff = 0.05, 
                  FDR = down_vs_tot_FDR,
                  graph_title = "Kinases prediction enrichment (IV-KEA) down vs tot",
                  x_axis_title = "log2(odds ratio)",
                  output_folder = 'myexperiment/',
                  file_name = "down_tot_volcano_plot_ivkea.pdf")

1. Compare enrichment tests from NetworKIN and IV-KEA

predictions_comparison(
  ivkea_enrichment_file = 'myexperiment/kinase_enrichment_ivkea.csv',
  networkin_enrichment_file = 'myexperiment/kinase_enrichment_networkin.csv',
  FDR_cutoff = 0.05, 
  graph_title = "Kinases prediction enrichment up vs down networkin vs iv-kea",
  output_folder = 'myexperiment/',
  file_name = "comp_plot.pdf"
)

gaelfortin/phosphogo documentation built on Nov. 26, 2020, 9:12 a.m.