# Mechanism of interaction between the transactivation domain of N-myc and the DNA-binding surface of TFIIIC5

## TL;DR

This study reveals how the N-myc protein interacts with TFIIIC5, a component of a key RNA transcription complex, through detailed structural and biochemical experiments.

## Contribution

The study identifies the DNA-binding domain of TFIIIC5 as a specific binding site for N-myc and provides a structural model for their interaction.

## Key findings

- The DNA-binding domain of TFIIIC5 interacts with two regions in the transactivation domain of N-myc.
- AlphaFold modeling predicts a binding mode for N-myc that overlaps with the C-terminal acidic plug of TFIIIC5.
- Removing the acidic plug of TFIIIC5 enhances N-myc binding, suggesting a competitive interaction with DNA.

## Abstract

N-myc is a transcription factor, a powerful driver of cellular growth and an important oncoprotein. N-myc interacts with many factors, including the RNA Polymerase III assembly factor, TFIIIC, a six-subunit complex that is essential for the transcription of small, structured RNA. TFIIIC and N-myc mutually restrict each other’s chromatin association, and their complex contributes to quality control in mRNA transcription. We previously demonstrated that the intrinsically disordered transactivation domain of N-myc interacts directly with a sub-complex of TFIIIC, τA. Structural studies by others show that DNA binding of τA is largely mediated by TFIIIC3, leaving open the role of the DNA-binding domain of TFIIIC5. Here, we demonstrate that this domain is a binding site for two regions in the transactivation domain of N-myc, through an integrated approach combining NMR spectroscopy, hydrogen–deuterium exchange mass spectrometry, and interaction assays (pull-downs, ITC, fluorescence polarization, and co-immunoprecipitation). AlphaFold modelling predicts with high-confidence a binding mode for the higher affinity N-myc motif that overlaps with the predicted intramolecular binding site of the C-terminal acidic plug of TFIIIC5, removal of which enhances the binding of N-myc. This model elucidates how the N-myc:TFIIIC5 interaction competes with DNA and other interactions, providing a basis for their mutual regulation.

Graphical Abstract