Dynamic network-guided CRISPRi screen reveals CTCF loop-constrained nonlinear enhancer-gene regulatory activity in...

Area of Science:

• Genomics
• Developmental Biology
• Systems Biology

Background:

• Enhancer discovery is challenging due to weak gene expression effects, particularly in complex diseases.
• Leveraging nonlinear enhancer-gene regulation during cell state transitions can enhance discovery sensitivity.

Approach:

• A CRISPRi-based enhancer screen was performed during human embryonic stem cell (hESC) definitive endoderm differentiation.
• Network modeling and mid-transition screening identified comprehensive enhancer sets for lineage-specifying transcription factors.
• A CTCF loop-constrained Interaction Activity (CIA) model was developed by integrating enhancer activity and 3D interaction data.

Key Points:

• Many identified enhancers showed strong effects mid-transition but weak effects post-transition.
• The CIA model outperformed traditional models relying solely on Hi-C contact frequency for predicting functional enhancers.
• The study identified 4-9 enhancers per locus for core endoderm transcription factors.

Conclusions:

• Generalizable strategies were developed for sensitive and comprehensive enhancer discovery during cell state transitions.
• This approach improves the understanding of gene regulation in both normal development and complex diseases.
• The findings offer a more effective method for identifying functional enhancers crucial for cellular processes.