PhD Defence: Frederik Friis Theisen

Title: The behaviour of short linear motifs in the DREB2A transcription factor and the effect of molecular context

Prof. Karen Skriver (main supervisor)
Prof. Birthe B. Kragelund (co-supervisor)

Assessment committee:
Research director Malene Ringkjøbing Jensen, Institut de Biologie Structurale, Grenoble
Research professor Xavier Salvatella, ICREA and IRB Barcelona
Prof. Kresten Lindorff-Larsen, University of Copenhagen (chair)

Abstract : Fast and accurate response to external stimuli is essential for organisms to survive. In plants, these responses must be facilitated by regulatory pathways of which transcription factors are a key component. Many transcription factors are controlled by sequence motifs found in dis- ordered regions. These motifs can be short and are often found in close proximity and thus the identification and characterization can be complicated. Representative model systems are therefore valuable in the effort to understand these interactions.

The interactions of the Radical-induced Cell Death1 (RCD1) RST do- main represent such a model system for studying regulation of tran- scription factors. One of the key members of the RCD1-RST model interactome is Dehydration Responsive Element Binding protein 2A (DREB2A), a stress response related transcription factor which has also been shown to interact with subunit 25 (Med25) of the Mediator complex.

This thesis both expands the experimental and analytical toolbox for analysis of disorder-based interactions and explores the interactions of DREB2A with RCD1-RST and Med25-ACID. The effect of motif flank- ing regions on the interaction with RCD1:DREB2A was characterized, through a combination of thermodynamics and structural methods, revealing the stabilizing effect of a dynamic flanking region. This study exploited a thermodynamic analysis method, presented by R. S. Spo- lar and M. T. Record Jr. in 1994, for quantification of conformational entropy changes and adapted for ID-based interactions as part of this work. The Med25-ACID interaction motif in DREB2A was identified using isothermal titration calorimetry (ITC) and emerging features of bivalency and heterogeneity were shown using chemical exchange satu- ration transfer (CEST) nuclear magnetic resonance (NMR) spectroscopy.

In conclusion, the work presented in this thesis provides insight into the regulatory mechanisms and pathways of the DREB2A transcription factor.