A De Novo Noncoding RARB Variant Associated with Complex Microphthalmia Alters a Putative Regulatory Element

Area of Science:

• Genetics and Developmental Biology
• Molecular Biology

Background:

• Retinoic acid receptor beta (RARB) is vital for eye development.
• RARB variants are linked to eye malformations like microphthalmia.
• A de novo variant was found in an individual with complex microphthalmia and developmental delay.

Purpose of the Study:

• To investigate the functional impact of a de novo RARB intronic variant.
• To determine if the variant affects mRNA splicing or gene regulation.
• To explore the variant's role in RARB overexpression and downstream gene expression.

Main Methods:

• In silico analysis for splicing and regulatory potential.
• In vitro minigene assays to assess mRNA splicing.
• Luciferase reporter assays to evaluate promoter activity.
• Overexpression studies in human lens epithelial cells.

Main Results:

• In silico tools suggested potential regulatory function and alternative splicing.
• Minigene assays confirmed no effect on RARB mRNA splicing.
• The variant significantly increased RARB promoter activity in vitro.
• RARB overexpression led to increased cell proliferation and FOXC1 expression.

Conclusions:

• The intronic conserved region (CR1) of RARB has a regulatory role.
• The de novo variant enhances RARB promoter activity, potentially causing overexpression.
• Altered RARB regulation may contribute to abnormal eye development and developmental delay.