Principal Supervisor: Prof. Steve SmerdonLink opens in a new window

Co-supervisor: Prof. Joanne Morris

PhD project title: Reading, writing and molecular switching in the DNA-damage response

University of Registration: University of Birmingham

Project outline:

How cells detect and respond to a bewildering variety of chemical, physical and environmental stimuli is a fundamental question in biology. Although much remains to be understood, it is clear that post-translational alteration of protein structure and behaviour through the timely addition of a variety of modifications such as phosphorylation, ubiquitylation, acetylation etc plays a central role. This is a simple but highly effective strategy since deposition - 'writing' - of the modification and its removal by a separate class of 'eraser' enzymes provides the desirable property of rapid reversibility whereby the modification itself acts as a molecular 'switch' that initiates or terminates downstream signalling events. How these modifications are generated and regulated, and how these switching mechanisms are manifested at the molecular level form our core interests and we investigate them using a highly multidisciplinary approach that enables us to relate structural and biochemical information to important physiological signalling mechanisms.

Our signalling network of choice is the DNA-damage response (DDR) for two main reasons. Firstly, disruption of the DDR is a characteristic of many human diseases including cancer, neurodegeneration, growth abnormalities and immunodeficiency and is the target of many therapeutic strategies. Secondly, its inherent complexity allows us to study how DNA-damage detection and repair are integrated with signalling to and from many other cellular processes such as metabolic state, cell-cycle, developmental stage, cell type-specific effects and the execution of transcriptional programmes.

Tens of thousands of sites of post-translational modifications (PTM) have been identified and mapped yet the functional significance of the vast majority remains unknown. PTMs can affect protein activity in many ways including promotion of conformational changes and triggering protein complex assembly through the activity of PTM-specific interaction modules or 'readers'. These effects are not mutually exclusive and our goal is to understand at the molecular level how multiple modifications can combine to generate the spectacular flexibility and precision in signal transduction that we observe in cells.

Projects in several relevant areas are available which will use a combination of emerging techniques in chemical/synthetic biology alongside classical structural biology methods and other biophysical, biochemical and cell-biological approaches to answer outstanding questions such as:

• What is the molecular basis of cross-talk between signalling pathways that converge on specific post-translationally modified sequence motifs?
• How are molecular barcodes generated from multiple and/or different types of modification interpreted to control cellular activities such as tethering and mobilisation of specific chromatin-associated signalling and repair complexes before, during and after DNA-damage
• How, and to what extent do post-translational modifications of specific sites contribute to large-scale conformational rearrangements in macromolecular repair complexes and how are these effects influenced by interactions with 'reader' proteins?
  
  

References:

Fournier, M. et al (2020) ‘KAT2-mediated acetylation switches the mode of PALB2 chromatin association to safeguard genome integrity’ BioRxiv 575910; 735811 doi: https://doi.org/10.1101/735811.

Kyrieleis, O.J.P. et al (2016) ‘Three-dimensional architecture of the human BRCA1-A histone deubiquitinase core complex’ Cell Reports, 17 3099-3106.

Yata, K. et al (2012) ‘Plk1 and casein kinase 2 act in concert to regulate Rad51 during recombinational repair’ Mol. Cell 45, 371-383.

Lloyd, J. et al (2009) 'A supra-modular FHA/BRCT-repeat architecture mediates Nbs1 adaptor function in response to DNA-damage' Cell 139, 100-111.

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

 

Techniques that will be undertaken during the project:

Molecular biology (cloning, site-directed mutagenesis etc)

Recombinant protein expression and purification

Chemical/synthetic biology (protein/peptide ligation, site-specific labelling, orthogonal ribosome technology)

Cell-biology– localisation, knock-down/complementation, signalling pathway analysis

Protein-protein/protein-ligand interactions (eg BIACore, ITC, co-immunoprecipitation, mass spectrometry)

Molecular modelling/dynamics

Structural/biophysical analysis (SEC-MALS, X-ray crystallography, EM)

 

Contact: Prof. Steve SmerdonLink opens in a new window