ChIP-Seq and ATAC-Seq identify enriched regions or peaks in genome. Depending on the assay, the biological meaning of enrichment changes; in ChIP-Seq, enrichment indicates protein binding, while in ATAC-Seq, enrichment indicates open chromatin. To understand the importance of enriched regions in regulating gene expression, we can add information about overlapping or nearby genomic features. 

What is Annotate peaks?

Annotate peaks takes an input set of regions and checks for overlap between those regions and a gene/feature annotation. This gives regulatory context for enriched regions.

Running Annotate peaks

The input for Annotate peaks is a Peaks type data node. 

• Click a Peaks data node

• Click the Peak analysis section in the toolbox

  
  

• Click Annotate peaks
• Set the Genomic overlaps parameter

The Genomics overlaps parameter lets you choose one of two options (Figure 1).

• Report one gene region per peak (precedence applies) chooses one gene section for each peak using the precedence order to settle cases where more than one gene section overlaps a peak. The order of precedence is TSS, TTS, CDS Exon, 5' UTR Exon, 3' UTR Exon, Intron, Intergenic.
• Report all gene regions per peak creates a row for each gene section that overlaps a peak in the task report. 
  
  

User should define the transcription start site (TSS) and transcription termination site (TTS) limit in the unit of bp

• Choose a gene/feature annotation from the drop-down menu
• Click Finish to run

Annotate peaks output

Annotate peaks produces an Annotated peaks data node. The Annotated peaks task report adds a Gene section breakdown pie chart and adds columns with information about the Gene IDs, Transcript IDs, Gene section, Distance to TSS, and Distance to TTS of each peak to the standard Peaks report  (Figure 12). If run with the option to report all gene sections selected, each peak will have a row for each gene section it overlaps. If run with the option to report one gene section selected, each peak will have one row with the gene section it overlaps chosen using the order of precedence. 

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The table can be sorted and filtered by any of its columns (Figure 23).  The Gene section breakdown pie chart will update as the table is filtered (the current (Peaks) and the total number of peaks (in parenthesis) are given in the upper left corner).  Click on the Optional columns on the upper-left corner of the table to add more information on each region

Clicking the Image Removed button at the bottom of the task report generates a Filtered peaks data node containing only the peaks currently included in the table. 

Gene sections

TSS

Transcription start site (TSS) is -1000bp and +100bp (default setting) from the TSS for a transcript

TTS

Transcription termination site (TTS) is -100bp and +1000bp (default setting) from the TTS for a transcript 

CDS Exon 

Coding sequence (CDS) Exon is overlapping a coding exon in a transcript

5' UTR Exon

5' Untranslated Region (UTR) Exon is overlapping an exon in the 5' UTR of a transcript

3' UTR Exon

3' Untranslated Region (UTR) Exon is overlapping an exon in the 3' UTR of a transcript

Intron

Intron is overlapping an intron in a transcript 

Intergenic

Intergenic is not located within 1000bp of a transcript

Annotate peaks vs. Annotate regions

Annotate regions is almost identical to Annotate peaks. Annotate regions takes Region counts data as the input and outputs Annotated region counts. One important difference is that you cannot filter the Annotated region counts from the task report. 

Run Annotate regions prior to performing differential analysis if you will want to be able to view gene section information and filter regions by gene section in the differential analysis task report.