AQuA-HiChIP

AQuA-HiChIP quantifies protein-anchored chromatin interactions by using orthogonal normalization of HiChIP paired-end tags (PETs) to provide absolute measurements of differential chromatin interactions.

Key Features:

• Absolute Quantification: Provides absolute quantification of chromatin interactions by counting PETs and using an orthogonal reference for normalization rather than normalizing by sequencing depth.
• Protein-Anchored Interaction Analysis: Captures protein-anchored chromatin interactions to measure changes in 3D genome architecture relevant to transcriptional regulation and epigenetic memory.
• Orthogonal Normalization: Normalizes human PET counts to total mouse PET counts to differentiate induced, protein-anchored interactions from background and reduce false positives and negatives.
• Chromatin Contact Capture: Employs biotin-dATP incorporation, proximity ligation, gentle shearing, and ChIP to enrich biotin-captured chromatin fragments prior to sequencing.
• Paired-end Sequencing of PETs: Uses paired-end sequencing of biotin-captured fragments to generate PETs for interaction calling and quantification.
• Rapid Protocol: Experimental workflow from cell culture to ranking chromatin interactions can be completed within six days.

Scientific Applications:

• 3D Genome Folding: Enables quantitative analysis of topological determinants of genome organization and chromatin folding principles.
• Transcriptional Control and Epigenetic Memory: Facilitates study of how 3D chromatin architecture influences transcriptional regulation and tissue-specific epigenetic memory.
• Super Enhancer Function: Allows investigation of effects such as histone deacetylase inhibition on super enhancers and the formation of aberrant contacts that disrupt enhancer function.

Methodology:

Human paired-end tag (PET) counts from HiChIP sequencing are normalized to total mouse PET counts to achieve absolute quantification of chromatin interactions.