Principal Supervisor: Prof. Teresa CarlomagnoLink opens in a new window, University of Birmingham

Co-supervisor: Dr. Andrew Bowmann, University of Warwick

PhD project title: Understanding nucleosome assembly: the interplay of order and disorder in histone chaperones

University of Registration: University of Birmingham

Project outline:

The genome of eukaryotic cells displays a hierarchy of structures that generate a tightly packed arrangement called chromatin. At the smallest scale, DNA molecules wind around protein complexes known as histone octamers to form nucleosomes. These nucleosomes are then packed in regular chromatin fibre arrangements that are in turn folded up to yield chromosomes.

Nucleosome are disassembled and reassembled during cell division. The process is highly controlled and assisted by a plethora of proteins (chromatin assembly factors, CAFs) with diverse functions, comprising histone modification factors, histone binding factors and ATP-dependent chromatin remodelling factors. This project will deal with the first two classes of CAFs.

Histone modification factors covalently modify histone proteins through an array of site-specific modifications, including acetylation, methylation, phosphorylation and ubiquitination. These modifications modulate DNA–histone and protein–histone interactions. They impact chromatin packing and represent a unique mechanism to regulate gene expression.

Histone binding factors act as histone chaperones, aiding histones in folding and binding to different partners, including histone modification enzymes, as well as delivering histones to the DNA.

Most histone modification factors, histone chaperones and the histone themselves contain large, disordered domains, in addition to well-folded globular domains. The interplay of order and disorder in the modulation of protein–protein interactions as well as protein function, is a burning question in biophysics and an emerging field in structural biology.

Molecules are known bind to each other through their complementary shapes (lock and key model). In the past 30 years this model has been expanded to include the notion that some molecules, whose three-dimensional shape is malleable, interact with more rigid structures by adapting to them, similar to pieces of play dough. This model provides opportunities for regulation of the intermolecular interactions by influencing the malleability of the play dough. Even more recently, the paradigm of “(folded) structure-function relationships” has been challenged by a revolutionary paper¹ describing a functional complex between two intrinsically unfolded proteins, which both remain disordered upon interaction (like in a tangle of ropes). In our recent work, we found a second example of this². We believe that this atypical interaction mode is much more common than anticipated and may represent a general mechanism for chaperoning disordered substrates.

In this project we will study selected histone chaperones and histone modification enzymes to establish the principles of functional regulation through the interplay of folded and unfolded intermolecular molecular interactions. These include the histone chaperones Asf1 and NASP³, as well as the acetylation enzyme Rtt109 and the ubiquitination enzyme UBR7.

This work goes beyond the conventional structure determination process, as the concept of “complex structure” is substituted by a large ensemble of transient conformations, a scenario that allows a favourable balance between the enthalpic and entropic terms of the free energy. Given the abundance of disordered motifs in the eukaryotic proteome, this research will improve our understanding of most eukaryotic cellular processes.

Furthermore, we will establish new interaction motifs (including disordered motifs) as drug targets in diseases whose molecular mechanism relates to chromatin assembly and modification. This work will expand the druggable targets space and will define general principles of how to target molecular flexibility or disorder.

References:

¹ Borgia et al. Extreme disorder in an ultrahigh-affinity protein complex Nature 555, 61–66 (2018)

² Danilenko et al. Histone chaperone exploits intrinsic disorder to switch acetylation specificity Nature Comm. vol 10, Article: 3435 (2019)

³ Bowman et al. Nucleic Acids Res 44, 3105–3117(2016).

BBSRC Strategic Research Priority: Understanding the rules of life – Structural Biology

Techniques that will be undertaken during the project:

NMR spectroscopy

Electron paramagnetic resonance spectroscopy

FRET and microscopy

Molecular dynamics simulations

Proteomics

Cell and molecular biology approaches (for example, mutational analysis, pulse chase experiments, pull-downs, etc.)

 

Contact: Prof. Teresa CarlomagnoLink opens in a new window