Dimer-Specific FokT-seq Reveals DNA-Binding Dimerization and Novel Genomic Targets of TDP-43

Area of Science:

• Neuroscience
• Molecular Biology
• Genetics

Background:

• Sporadic amyotrophic lateral sclerosis (ALS) is characterized by impaired TAR DNA-binding protein 43 (TDP-43) dimerization and insoluble TDP-43 accumulation in postmortem brain tissues.
• The precise mechanism linking TDP-43 dimerization loss to ALS pathogenesis remains unclear, necessitating further investigation.

Purpose of the Study:

• To investigate the role of TDP-43 dimerization in DNA binding and its implications in ALS.
• To develop a novel system for studying TDP-43 dimerization and its DNA targets.

Main Methods:

• Development of the FokT system, fusing TDP-43 with FokI nuclease to restore dimerization-dependent nuclease activity.
• Establishment of the FokT-seq method (FokT combined with Guide-seq) for genome-wide detection of DNA double-strand breaks induced by dimerized TDP-43.

Main Results:

• The FokT system successfully reactivated FokI nuclease activity by restoring TDP-43 dimerization, enabling targeted DNA cleavage.
• FokT-seq identified genome-wide DNA sites bound by dimerized TDP-43, revealing essential targets.
• The study confirmed the critical role of TDP-43 dimerization in DNA binding.

Conclusions:

• TDP-43 dimerization is essential for its DNA-binding function.
• The developed FokT system and FokT-seq method provide powerful tools for studying dimerized transcription factors and their roles in diseases like ALS.