1. Investigating a new protein in replication stress
Supervisor: Dr Eva Petermann
Accurate replication of the genome is essential for cell division and genome stability. Transcription is essential for gene expression and cellular function. Both processes use the same template – DNA – and can therefore clash with each other, leading to replication or transcription stress. Replication stress from clashes with transcription may play important roles in the genomic instability that can lead to cancer development.
We still do not know much about how replication and transcription interact with each other to prevent clashed and replication stress. To understand this better, our lab is interested in a protein of unknown function called TCEAL4. TCEAL4 has similarities to transcription factors, but our lab has found evidence that it could also play a role in replication stress.
This project aims to investigate further whether TCEAL4 plays a role in replication stress. You will work closely with lab members to grow human cell lines, transfect siRNA, run a Western blot and use a fluorescence microscopy assay to look at DNA replication or DNA damage with or without TCEAL4.
The student will be supervised by Dr Eva Petermann (https://www.evapetermann.org/)
2. Characterisation of the overexpression of single subunits of RNA Polymerase II as a novel marker for highly aggressive tumours
Supervisors: Martina Sadurni and Dr Marco Saponaro
Cells use a process called transcription to copy the information contained in our genomes and use this information to produce proteins in cells. The whole procedure needs to be tightly regulated to avoid damaging the DNA during the process. Indeed, deregulated transcription is associated with many diseases including cancer. The transcription of the mRNA, the protein-coding part of the genome, is carried out by RNA Polymerase II (RNAPII), a complex of 12 subunits. We have identified that upregulations of single subunits of the complex correlate with genome unstable cancers with reduced patients’ survival. In particular, the upregulation of the 4 largest subunits of the RNAPII (POLR2A-D) is associated with reduced survival across multiple tumour types, including acute myeloid leukaemia, breast invasive carcinoma, head and neck squamous cell carcinoma and brain lower grade glioma. Altogether, we want to fully understand how these deregulations impact on cells and whether the increased genomic instability in cancers is associated with increased transcription-induced DNA damage. We propose that the up-regulation of the four largest subunits of RNAPII could constitute a new biomarker for high-risk patients, therefore it is crucial to understand how increased levels of POLR2A-D impact on transcription and genome stability, as such understanding could help design new therapies for refractory tumours in poor survival cancer patients.
We have generated a doxycycline inducible cell line model using HeLa T-REx cell line to study the over expression of RNA Pol II subunits. Our data so far suggests that overexpression of the single subunits induces DNA damage in cells and transcription defects. We have now generated a new cell line for the over expression of POLR2A, the largest subunit of the complex and we aim to investigate whether the over expression of this single subunit induces DNA damage. The student will firstly assess the overexpression level of POLR2A by quantitative PCR (qPCR) and western blot. Subsequently, the student will determine whether the over expression of the subunit causes increased DNA damage by immunohistochemistry, performing a microscopy analysis of DNA damage markers. Therefore, the student will have the opportunity to learn and combine different molecular and cellular biology techniques and will contribute to understand more about mechanisms underlying these highly aggressive tumours.
The student will be supervised by Martina Sadurni, a PhD student in the Saponaro group.
3. Production of recombinant proteins from bacteria for use in in vitro studies of DNA replication.
Supervisor: Dr Neville Gilhooly
Our lab uses Xenopus egg extracts to study various mechanistic aspects of DNA replication. We commonly utilise recombinant DNA and protein purification techniques to generate proteins that we can add into our extracts to modify their behaviour. The student will purify 3 proteins using a standard molecular biology pipeline from start to finish i.e from starting DNA constructs to purified protein. In doing so, the student will learn essential molecular biology and biochemistry skills such as: working with and genetically manipulating E. coli, protein purification and analysis. The project will be of interest to a student aiming to acquire practical biochemical skills that will be essential for any molecular biology themed research programme they may be interested to pursue in the future.
The student will be hosted in Dr Aga Gambus’ lab and supervised by Dr Neville Gilhooly, a senior postdoc in the group.
4. Understanding DNA damage repair pathway in human papillomavirus induced disease progression
Supervisor: Dr Christy Varghese
Persistent infection with high-risk human papillomaviruses (HPV) can cause anogenital as well as oropharyngeal cancers. HPV life cycle is closely linked to the differentiation of the host epithelial cell and several cellular pathways are manipulated to promote their life cycle. DNA damage repair (DDR) pathways such as ATM and ATR are activated in HPV infections and are critical in the completion of HPV life cycle. Activation of these pathways result in the recruitment of downstream factors such as CHK2, γH2AX, BRCA1 and RAD51 to discrete nuclear foci. Studies with biopsy materials have showed increased association of DDR factors with the grade of cervical lesion.
Apart from the activation of DDR pathways, persistent HPV infections accumulate mutations that contribute to cancer progression. TCGA data has revealed the presence of APOBEC mutational signature in HPV-positive head and neck as well as cervical cancers. APOBECs are a family of cytidine deaminases that convert cytosine to uracil in ssDNA. Kono et al. have shown that DDR pathway activation in HPV-positive cells closely corelates with APOBEC3B upregulation.
Therefore, we aim to study the DDR pathway activation and APOBEC expression in HPV induced cancer progression by utilizing our state-of-the-art physiological tissue models. Expression of DDR factors downstream to both ATM and ATR pathways such as CHK2, γH2AX, BRCA1 and RAD51 will be assessed in low passage and high passage primary keratinocytes derived from three different patient donors transfected with high-risk HPVs such as HPV16 or HPV18 to understand the DDR pathway activation in HPV induced disease progression. In addition, APOBEC3A and APOBEC3B expression will also be assessed in the same set of samples to study the linkage between DDR pathways and APOBECs in HPV induced disease progression. The student involved in this internship project will learn the following laboratory techniques - Cell culture, Extraction of RNA from cellular lysates, cDNA synthesis, Design of qPCR primers, qPCR and data analysis using GraphPad and Scientific data presentation.
The student will be supervised by Dr Christy Varghese, a post-doc in the Parish group.
5. Investigating how cellular enzymes alter resistance to cancer therapies
Supervisors: Ellie Sweatman and Dr Martin Higgs.
Accumulation of unrepaired DNA damage is a key step during cancer development. To prevent this, cells have evolved multiple pathways to repair DNA damage, known collectively as the DNA damage response (DDR). Defects in the DDR also provide opportunities to target cancer cells for treatment. One of the most successful anti-cancer drugs exploiting such defects are Poly (ADP-ribose) polymerase inhibitors (PARPi).
PARPi are currently approved for the treatment of breast, ovarian and pancreatic tumours that have defects in one of the main DDR pathways called homologous recombination (HR). However, the clinical success of PARP inhibitors is hampered by development of resistance. Recently we have shown that loss of a particular enzyme, called SETD1A, leads to PARPi resistance in certain cancer cells.
The aim of this project will be to assess whether this resistance occurs with a spectrum of PARPi that are used clinically, since they all act in different ways. During this project you will use a variety of techniques (cell culture, immunofluorescence, immunobotting) to investigate this question. The data you will produce will help us understand how loss of SETD1A affects cancer therapy and how we can optimise PARPi in the clinic to better treat cancer. You will be supervised by Ellie Sweatman, a PhD student in the Higgs group.
6. Studying the function of ATM in the development of the nervous system
Supervisors: Matthew Taylor and Dr Richard Tuxworth
Our lab is interested in the link between the DNA damage response and development of the nervous system. Mutations in several genes that encode key regulators of the DNA damage response lead to severe neurological defects in children. Why this is the case is not fully understood. We use the fruit fly, Drosophila, to study what the proteins are doing in the nervous sytem and what happens if mutate them or reduce expression. Drosophila offers powerful genetic tools to manipulate gene function, coupled with a functional and anatomical tools to interrogate neural development.
One of the key regulators of the DNA damage response is a kinase called ATM. We have used CRISPR/Cas9 gene editing to insert GFP into the Drosophila ATM gene so we can see where ATM is localised in cells and neurons. The GFP-ATM protein is able to rescue lethality of dATM mutations, which indicates it is functional, is detectable via immunofluorescence and western blotting, and in excitingly, in neurons it appears to localise to synapses. However, GFP-ATM flies appear to be infertile and we don’t know why. We do not yet know whether this is affecting the males, the females, or both. One aspect of the student’s project will be to determine the cause of infertility, performing dissections and techniques such as western blotting, immunofluorescence microscopy and qPCR on Drosophila testes and ovaries to ask where GFP-ATM is localised and to try and work out what it is doing.
ATM is activated by DNA damage and also by rises in reactive oxygen species. The student will also conduct a simple RNAi screen to disrupt DNA damage response and oxidative stress signalling while simultaneously removing ATM from neurons. This may help us understand what ATM is doing and how it is activated specially in neurons.
The student will receive training in genetics, western blotting, dissections and immunofluorescence microscopy. The student will be supervised by Matthew Taylor, a PhD student in the Tuxworth group.
7. Modelling cancer development in zebrafish
Supervisor: Professor Ferenc Mueller
Our group uses zebrafish to understand fundamental aspects of vertebrate development. Zebrafish embryos are transparent which means intricate processes going on during development can be visualised live and in real time – something that would be impossible in any other vertebrate model. Recently, we have taken advantage of this to ask also whether zebrafish can be used to study how cancers development in the body.
In this project the student will have an opportunity to familiarise with the use of zebrafish embryos in modelling developmental biology and cancer development problems. The placement will include preparing and processing zebrafish embryos for imaging applications and provide training in embryo staging , and phenotype analysis by fluorescence microscopy detection of transgenic reporters and immunostaining. The student will be supervised by Prof Ferenc Mueller:
8. Exploring the role of a new protein modification in cancer
Supervisors: Dr Sally Fletcher and Dr Regina Andrijes
We would like to welcome prospective summer students to join the Coleman lab to investigate the role of an exciting protein modification in cancer. We study how ‘hydroxylation’, the enzymatic incorporation of a single oxygen atom into a target substrate, is deregulated in tumour cells. We study a variety of cancer-related processes from cell adhesion to DNA damage and protein synthesis so there is a lot of learning potential on offer. We also use a multidisciplinary approach to tackle the questions we ask from genome editing with CRISPR and mass spectrometry-based proteomics to western blotting and microscopy imaging. The internship will be a great opportunity to learn new biology and new skills and to help inform your decisions about scientific interests and future careers.
The student will be supervised by Dr Sally Fletcher and Dr Regina Andrijes who are post-docs in the Coleman group: https://www.birmingham.ac.uk/staff/profiles/cancer-genomic/coleman-matthew.aspx
9. Defining our next patient, carer and public engagement events in cancer
Supervisors: Dr Rowena Sharpe and Mrs Karen Turner
Background: The involvement of patients, their carers and members of the public throughout the lifetime of a clinical trial ensures that the research is carried out ‘with’ and ‘by them rather than being ‘about’ or ‘for’ them. This is an active partnership where patients, carers and members of the public influence and shape research. The University of Birmingham is ready to launch an ambitious new Cancer Patient, Carer, Public Involvement and Engagement (PCPIE) strategy in cancer. Following an 18-month consultation exercise via our PCPIE Taskforce it became clear that despite our high level of patient involvement in research, our patient, carer and public communities want more engagement with cancer research, feeling this would increase the levels of trust and meaningful involvement.
A key aim of our strategyis to build trust with communities by developing a strong two-way relationship and enable meaningful problem solving. The first step of this is to hear what our patients, carers and public say they would like from us. The second step is to establish a series of engagement events. This could be large events (such as a formal day event with speakers, information stalls and debates) and/or small events (such as informal coffee and chats) as well as the utilisation of social media to provide timely information to patients, carers and the public.
Project outline: We are creating a very simple survey to send out to our patients and the public and the aim of this internship would be, with strong support from the project supervisors, to analyse the results of this survey and generate proposals for what our first series of patient, carer and public engagement events should be.
The student will be supervised by Dr Rowena Sharpe, Director of Precision Medicine Trials and Mrs Karen Turner, Cancer Research UK Senior Research Nurse