i-Motif, not G-quadruplex, stability regulates insulin expression

Area of Science:

• Genetics
• Molecular Biology
• Biophysics

Background:

• The insulin-linked polymorphic region (ILPR) is a variable tandem repeat in the human insulin gene promoter.
• ILPR sequences can form G-quadruplexes (G4) and i-motifs (IM), DNA structures implicated in gene regulation.
• The precise roles of G4 and IM in regulating insulin gene expression, especially when co-occurring, are not fully understood.

Purpose of the Study:

• To investigate the relationship between ILPR sequence diversity, DNA structure formation (G4 and IM), and insulin gene regulation.
• To determine the conditions under which G4 and IM structures influence transcriptional activity in response to glucose.
• To elucidate the specific roles of G4 and IM structures in modulating insulin gene promoter activity.

Main Methods:

• Characterization of nine ILPR-based DNA sequences using biophysical techniques.
• Assessment of DNA structure stability and formation.
• Luciferase reporter assays to measure promoter activity under varying glucose conditions.

Main Results:

• Transcriptional activation of the insulin gene promoter was observed only when both G-quadruplex and i-motif structures could form.
• Insulin gene promoter activity showed a positive correlation with i-motif stability, but not with G-quadruplex stability.
• Different combinations of G4 and IM structures did not induce transcription, highlighting the necessity of both structures.

Conclusions:

• A model is proposed where G-quadruplexes may initiate transcription, while i-motifs modulate insulin gene expression.
• The formation and stability of both G-quadruplexes and i-motifs are crucial for glucose-responsive insulin gene regulation.
• These DNA structures represent a dynamic, interdependent system with potential as therapeutic targets for gene expression modulation.