Congenital Heart Disease-associated Coding and Non-Coding Mutations alter TBX5-DNA Binding Affinity
Laboratory Genetics and Genomics
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Primary Categories:
- Genomic Medicine
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Secondary Categories:
- Genomic Medicine
Introduction:
Congenital heart diseases (CHDs) are the most common birth defect, affecting 1 in every 100 infants. CHDs are characterized by structural abnormalities that occur during heart development. Among the possible causes of CHDs are mutations that alter cardiac transcription factor (TF) function, such as TBX5. TFs are proteins that bind to specific non-coding regulatory DNA to regulate gene expression and control biological processes. TBX5 is crucial to regulating heart development, and mutations in the TBX5 gene have been shown cause CHDs. Additionally, genome-wide association studies (GWAS) have established a strong associations between coding and non-coding genetics variants and some types of CHD. In this work, we evaluated the potential of CHD-associated coding and non-coding mutations to alter TBX5 function.
Methods:
To evaluate the impact of missense mutations, we tested five TBX5 missense mutants (I54T, M74V, I101F, R113K, and R237W) and assessed how they altered TBX5 thermal stability and the DNA-binding affinity of TBX5 to its cognate site. For the non-coding variants, we used a machine learning-based predictive model (LS-GKM SVM) to prioritize CHD-associated mutations that create or disrupt TBX5 binding sites. We prioritized four non-coding variants (rs34444956, rs2465147, rs262828, and rs11904437) for in vitro validation on TBX5-DNA binding affinity.
Results:
Missense mutations experiments showed a decrease in binding affinity for two known TBX5 binding sites and a significant change in thermal stability for all five of the missense variants. Non-coding mutation experiments showed that rs34444956, rs2465147, and rs262828 increase binding affinity, whereas rs11904437 decreases binding affinity.
Conclusion:
In short, our finding show that coding variants can alter the thermal stability of TBX5 and impact the binding affinity to known TBX5 binding sites. Additionally, CHD-associated non-coding variants also impacted TBX5-DNA binding by creating and disrupting TBX5 binding sites. Completing this project will provide crucial information on the biochemical mechanism of coding and non-coding variants associated with CHDs. Our findings will aid in the development of novel treatments and early diagnostic methods for CHD patients.
Congenital heart diseases (CHDs) are the most common birth defect, affecting 1 in every 100 infants. CHDs are characterized by structural abnormalities that occur during heart development. Among the possible causes of CHDs are mutations that alter cardiac transcription factor (TF) function, such as TBX5. TFs are proteins that bind to specific non-coding regulatory DNA to regulate gene expression and control biological processes. TBX5 is crucial to regulating heart development, and mutations in the TBX5 gene have been shown cause CHDs. Additionally, genome-wide association studies (GWAS) have established a strong associations between coding and non-coding genetics variants and some types of CHD. In this work, we evaluated the potential of CHD-associated coding and non-coding mutations to alter TBX5 function.
Methods:
To evaluate the impact of missense mutations, we tested five TBX5 missense mutants (I54T, M74V, I101F, R113K, and R237W) and assessed how they altered TBX5 thermal stability and the DNA-binding affinity of TBX5 to its cognate site. For the non-coding variants, we used a machine learning-based predictive model (LS-GKM SVM) to prioritize CHD-associated mutations that create or disrupt TBX5 binding sites. We prioritized four non-coding variants (rs34444956, rs2465147, rs262828, and rs11904437) for in vitro validation on TBX5-DNA binding affinity.
Results:
Missense mutations experiments showed a decrease in binding affinity for two known TBX5 binding sites and a significant change in thermal stability for all five of the missense variants. Non-coding mutation experiments showed that rs34444956, rs2465147, and rs262828 increase binding affinity, whereas rs11904437 decreases binding affinity.
Conclusion:
In short, our finding show that coding variants can alter the thermal stability of TBX5 and impact the binding affinity to known TBX5 binding sites. Additionally, CHD-associated non-coding variants also impacted TBX5-DNA binding by creating and disrupting TBX5 binding sites. Completing this project will provide crucial information on the biochemical mechanism of coding and non-coding variants associated with CHDs. Our findings will aid in the development of novel treatments and early diagnostic methods for CHD patients.