Differential Transcription Factor Activity Analysis

Warning

Temporary documentation

Once coverage-based or fragmentomics-based features have been generated for multiple Transcription Factors (TFs), it is possible to derive condition/cluster/group-specific differentially active TFs. LBFextract offers “post_extraction_analysis_commands” which enables this analysis to be performed. To achieve this, a sample sheet (a “.csv” file) containing the paths to the signals in a column, the group membership in the group column, and other relevant information needs to be generated and filled.

LBFextract enables automatic generation of the sample sheet, which can be obtained with the following command:

$ lbfextract post_extraction_analysis_commands --path_to_res_summary DIRECTORY --path_to_sample_sheet FILE generate-sample-sheet

This command generates a sample sheet in the correct format, which can then be filled with the necessary information. path_to_res_summary represents the root folder where the signals have been stored. LBFextract will parse it and load all the CSV files present in there. The structure of the generated file will be as follows:

index, sample_name, group, tumor_fraction, cov, signal_type, bed_file_metadata, path_to_res_summary,paths_to_sample_result

Possible extra columns representing different group definitions can be added. For the differentially active analysis, the only essential columns are: index, paths_to_sample_result, and group. index and path are automatically filled. The group column needs to be filled. The sample_name, bed_file_metadata, and signal_type columns are automatically filled too, but may need to be updated (i.e., The name used in the sample_name column is the name of the file containing the signal. A shorter one may be desired).

After the generation of the sample sheet, the analysis can be performed with the following command:

$ lbfextract post_extraction_analysis_commands --path_to_res_summary DIRECTORY --path_to_sample_sheet FILE --signal_length INTEGER --center_signal_indices TEXT --flanking_signal_indices TEXT --output_path DIRECTORY --outer_group_column TEXT --correction_method TEXT --remove_outliers --save_individual_plots get-differentially-active-genomic-intervals

With this analysis, LBFextract will load all the signals for the defined TFs, extract the accessibility of each one of them for each sample, and generate a matrix of samples by TFs which will be used to find the differentially active TFs between groups.

The results will include:

  • A table containing all TFs with adjusted p-values, log2 fold changes, and other summary statistics.

  • A table containing the enrichment analysis.

  • Summary plots per TF containing the signal per group.

  • A heatmap containing the differentially active TFs found, the log2 fold values, and the adjusted p-values.

Heatmap of differentially active transcription factors