Multimodal bHLH-PAS DNA binding controls specificity and drives obesity

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The basic-helix-loop-helix Per-Arnt-Sim (bHLH-PAS) transcription factor (TF) family are crucial regulators of physiological and environmental responses.
• Specificity mechanisms for DNA binding and target gene regulation by bHLH-PAS TFs are not fully understood.

Purpose of the Study:

• To systematically analyze the DNA binding hierarchies and specificity mechanisms of key bHLH-PAS TF family members.
• To elucidate the role of DNA shape and flanking sequences in TF binding specificity.
• To investigate the functional significance of SIM1 PAS-loop/DNA interactions in TF activity and disease.

Main Methods:

• Systematic analysis of cognate DNA binding hierarchies for prototypical bHLH-PAS TFs (ARNT, ARNT2, HIF1α, HIF2α, AhR, NPAS4, SIM1).
• Investigation of DNA binding footprints and sequence preferences.
• Analysis of TF-DNA interactions, including DNA shape and flanking sequence contributions.
• Functional studies in a SIM1.R171H knock-in mouse model.

Main Results:

• Identified large DNA binding footprints (12-15 bp) for bHLH-PAS TFs.
• Revealed flank-encoded DNA binding specificity, distinguishing SIM1 and HIF TF binding through N-terminal interactions.
• Demonstrated a relationship between DNA shape, core, and flank TF binding, enabling motif sequence flexibility.
• Discovered novel SIM1 PAS-loop/DNA interactions with AT-rich sequences critical for DNA binding and transcriptional activity.
• Established a monogenic cause for hyperphagic obesity linked to SIM1 function in a mouse model.

Conclusions:

• bHLH-PAS TF binding specificity is achieved through multimodal mechanisms involving core motifs, flanking sequences, and DNA shape.
• SIM1 PAS-loop/DNA interactions are essential for TF function and linked to hyperphagic obesity.
• This study resolves key aspects of bHLH-PAS TF DNA binding specificity and its implications for homeostasis and disease.