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Barrier to autointegration factor interacts with the cone-rod homeobox and represses its transactivation function
Xuejiao Wang1, Siqun Xu, Carlo Rivolta
1Department of Ophthalmology and Visual Sciences, Molecular Biology and Pharmacology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Abstract:
Crx (cone-rod homeobox) is a homeodomain transcription factor implicated in regulating the expression of photoreceptor and pineal genes. To identify proteins that interact with Crx in the retina, we carried out a yeast two-hybrid screen of a retinal cDNA library. One of the identified clones encodes Baf (barrier to autointegration factor), which was previously shown to have a role in mitosis and retroviral integration. Additional biochemical assays provided supporting evidence for a Baf-Crx interaction. The Baf protein is detectable in all nuclear layers of the mouse retina, including the photoreceptors and the bipolar cells where Crx is expressed. Transient transfection assays with a rhodopsin-luciferase reporter in HEK293 cells demonstrate that overexpression of Baf represses Crx-mediated transactivation, suggesting that Baf acts as a negative regulator of Crx. Consistent with this role for Baf, an E80A mutation of CRX associated with cone-rod dystrophy has a higher than normal transactivation potency but a reduced interaction with Baf. Although our studies did not identify a causative Baf mutation in retinopathies, we suggest that Baf may contribute to the phenotype of a photoreceptor degenerative disease by modifying the activity of Crx. In view of the ubiquitous expression of Baf, we hypothesize that it may play a role in regulating tissue- or cell type-specific gene expression by interacting with homeodomain transcription factors.
Insights
Barrier to autointegration factor (Baf) interacts with cone-rod homeobox (Crx) transcription factor. Baf negatively regulates Crx activity, potentially influencing photoreceptor gene expression and disease.
Area of Science:
- Molecular Biology
- Genetics
- Neuroscience
Background:
- Cone-rod homeobox (Crx) is a crucial transcription factor for photoreceptor and pineal gene regulation.
- Understanding Crx interactions is key to deciphering retinal development and function.
Purpose of the Study:
- To identify novel protein interactors of Crx within the retina.
- To elucidate the functional role of identified interactors in Crx-mediated gene regulation.
- To explore the potential involvement of these interactions in retinal degenerative diseases.
Main Methods:
- Yeast two-hybrid screening of a mouse retinal cDNA library to identify Crx-binding proteins.
- Biochemical assays to confirm the Baf-Crx interaction.
- Transient transfection assays using a rhodopsin-luciferase reporter in HEK293 cells to assess Baf's regulatory effect on Crx.
- Analysis of a cone-rod dystrophy-associated CRX mutation (E80A) for altered Baf interaction and transactivation.
Main Results:
- Barrier to autointegration factor (Baf) was identified as a Crx-interacting protein.
- Baf is expressed in nuclear layers of the mouse retina, including photoreceptor and bipolar cells.
- Overexpression of Baf represses Crx-mediated transactivation of the rhodopsin gene, indicating Baf acts as a negative regulator.
- A CRX mutation linked to cone-rod dystrophy exhibits increased transactivation but reduced Baf interaction.
Conclusions:
- Baf negatively regulates the transcriptional activity of Crx.
- Baf may contribute to photoreceptor degenerative diseases by modulating Crx function, particularly in cases with CRX mutations.
- Baf's interaction with homeodomain transcription factors like Crx suggests a broader role in regulating tissue-specific gene expression.