Identifying interactions between TDP-43's N-terminal and RNA-binding domains

Area of Science:

• Biochemistry
• Molecular Biology
• Neuroscience

Background:

• TAR DNA-binding Protein 43 kilodaltons (TDP-43) is implicated in amyotrophic lateral sclerosis pathogenesis.
• TDP-43 is an intrinsically disordered protein with distinct domains: N-terminal domain (NTD), RNA recognition motifs (RRMs), and a C-terminal glycine-rich domain.
• Previous work suggested allosteric alterations in TDP-43 due to a chemical probe, hinting at interdomain interactions.

Purpose of the Study:

• To investigate the interaction between the NTD and RRMs of TDP-43.
• To determine the impact of the NTD on TDP-43's RNA-binding capabilities.
• To elucidate the structural basis for TDP-43's role in amyotrophic lateral sclerosis.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy (2D [1H,15N] HSQC and Carr-Purcell-Meiboom-Gill) to compare TDP-43 constructs.
• Protein-protein docking to model NTD-RRM interactions.
• Microscale thermophoresis to assess RNA binding affinity.

Main Results:

• NMR revealed clustered shifts in RRM RNA-binding sites when NTD was present, even without RNA.
• Computational modeling proposed NTD stacking onto the RRM domains.
• Experimental evidence confirmed an interaction between NTD(1-102) and RRMs(102-260).
• TDP-43 constructs with NTD showed significantly altered RNA binding compared to RRMs alone.

Conclusions:

• The NTD of TDP-43 directly interacts with its RRM domains.
• This NTD-RRM interaction influences TDP-43's ability to bind RNA.
• Understanding these interdomain interactions is key to TDP-43's function and dysfunction in ALS.