beta-catenin Ser45 in development and disease
Lead Research Organisation:
University of Edinburgh
Department Name: The Roslin Institute
Abstract
Mutations in the beta-catenin signalling pathway are found in many types of cancer. It is widely assumed that all these mutations activate this pathway. We are interested in the development of Wilms' tumours, kidney cancers found in young children that are the result of normal kidney development before birth going wrong. A subset of these tumours has mutations in beta-catenin. Interestingly, whereas there are four different mutations possible, almost all patients have exactly the same mutation, suggesting this mutation is specifically important for this type of cancer. In the work leading up to this proposal we have made mouse models that carry different mutant copies of beta-catenin, including the mutation found in Wilms' tumours. We found that this mutation resulted in kidney abnormalities that would fit with a role in Wilms' tumours but, surprisingly, this mutation did not seem to activate the protein. Unfortunately, these models were made in a way that differs from the way the gene is mutated in patients, and it is therefore not possible to conclude this mutation will have the same effect in tumours.
In this project we will therefore first make the mutation in exactly the same way as found in human tumours. We will repeat the analyses we have done before on the other model to see if the surprising findings we did are confirmed. We will study the effect of this mutation in mice, with an emphasis on the kidney and neuron / brain.
In this project we will therefore first make the mutation in exactly the same way as found in human tumours. We will repeat the analyses we have done before on the other model to see if the surprising findings we did are confirmed. We will study the effect of this mutation in mice, with an emphasis on the kidney and neuron / brain.
Technical Summary
It is assumed that oncogenic mutations found in beta-catenin result in activation of the pathway. We have a strong interest in the early steps in the development of Wilms' tumours, in particular the subset that is caused by loss of WT1 and activation of beta-catenin. Interestingly, almost all cases of Wilms' tumour with mutant beta-catenin carry a mutation affecting the Ser45 residue. To study the reason for this selection we have previously generated several mouse models that express different beta-catenin mutations from the Rosa26 locus. We found that the Ser45 mutant does not have an increased signalling capacity, despite being stabilized at the protein level as expected. Still, the Ser45 mutation has a biological effect on Wt1-deficient kidney mesenchyme cells which were driven a step closer towards Wilms' tumours. This finding suggests a complete new dimension to the role of beta-catenin mutations in cancer. However, the fact that our models express their mutant beta-catenin from Rosa26 means we cannot directly translate these finding to patients. In this project we will therefore generate a Cre-inducible Ser45 mutation in the endogenous mouse beta-catenin gene. We will test its capability to increase target gene transcription in ES cells as we did for the Rosa26 models. In vivo we will activate the expression of the mutant allele through a Cre deleter strain and analyse the phenotype in development and/or postnatal. To study the role of this mutation in Wilms' tumour development we will cross the mice with our kidney mesenchyme-specific Wt1 knockout. We will use genome-wide expression analysis to identify molecular changes that could explain the selection for this mutation in WT1-mutant Wilms' tumours. The mice will be followed for Wilms' tumour development and other phenotypes. Finally, we will specifically analyse any phenotypes in the neuronal system caused by the Ser45 mutation.
Planned Impact
The proposed project will have a potential impact on the following stake holders.
The academic community working on cancer genetics in general, and Wilms' tumour genetics in particular. The information gained in this project will increase our understanding of cancer genetics, Darwinian selection in tumourigenesis and the roles of specific cancer-related mutations in the origin of tumours.
The health sector. Better understanding of the earlier steps in cancer development will ultimately lead to improvement in cancer treatment options.
Cancer patients. Better understanding of the origins of cancer and identification of the processes that, when disturbed, lead to cancer development will allow the design of more efficient and more specific therapies. This will result in better treatments with fewer side-effects, leading to a significant increase in quality of life.
General public. As the public pays for MRC-funded research through their taxes there is an obvious right of information on what is done with the money.
The pharmaceutical industry. There is an important lack in sufficiently good animal models for cancer. The use of models that are available for design and testing of therapies have in many cases been found not to be predictive for the efficacy of a therapy in patients. As a result too many new drug candidates are failing to make it to the clinic. Models as proposed here that mimic the genetic events in cancer in patients as closely as possible will undoubtedly lead to better and more predictive models for these purposes.
The staff on the project. Biomedical sciences increasingly require a multitude of techniques and approaches to answer a scientific question. The experimental models used become more sophisticated as technical possibilities improve. For both named staff this project will allow the extension of their expertise. For Dr. Ozdemir this project will help to further establish her reputation in the Wilms' tumour and mouse model field. She will be able to increase her expertise in the generation of genetically modified mouse models using the latest technologies, and introduce her to next-generation sequencing approaches. Mrs Sheraz has so far been running the mouse colony of the Hohenstein lab and been involved in developing and optimizing in vitro culture methods for mesenchymal kidney progenitor cells. This project will allow her to become experienced in all steps in the generation and analysis of transgenic mouse models.
The academic community working on cancer genetics in general, and Wilms' tumour genetics in particular. The information gained in this project will increase our understanding of cancer genetics, Darwinian selection in tumourigenesis and the roles of specific cancer-related mutations in the origin of tumours.
The health sector. Better understanding of the earlier steps in cancer development will ultimately lead to improvement in cancer treatment options.
Cancer patients. Better understanding of the origins of cancer and identification of the processes that, when disturbed, lead to cancer development will allow the design of more efficient and more specific therapies. This will result in better treatments with fewer side-effects, leading to a significant increase in quality of life.
General public. As the public pays for MRC-funded research through their taxes there is an obvious right of information on what is done with the money.
The pharmaceutical industry. There is an important lack in sufficiently good animal models for cancer. The use of models that are available for design and testing of therapies have in many cases been found not to be predictive for the efficacy of a therapy in patients. As a result too many new drug candidates are failing to make it to the clinic. Models as proposed here that mimic the genetic events in cancer in patients as closely as possible will undoubtedly lead to better and more predictive models for these purposes.
The staff on the project. Biomedical sciences increasingly require a multitude of techniques and approaches to answer a scientific question. The experimental models used become more sophisticated as technical possibilities improve. For both named staff this project will allow the extension of their expertise. For Dr. Ozdemir this project will help to further establish her reputation in the Wilms' tumour and mouse model field. She will be able to increase her expertise in the generation of genetically modified mouse models using the latest technologies, and introduce her to next-generation sequencing approaches. Mrs Sheraz has so far been running the mouse colony of the Hohenstein lab and been involved in developing and optimizing in vitro culture methods for mesenchymal kidney progenitor cells. This project will allow her to become experienced in all steps in the generation and analysis of transgenic mouse models.
Organisations
Publications
Rojo R
(2019)
Deletion of a Csf1r enhancer selectively impacts CSF1R expression and development of tissue macrophage populations.
in Nature communications
Grabert K
(2020)
A Transgenic Line That Reports CSF1R Protein Expression Provides a Definitive Marker for the Mouse Mononuclear Phagocyte System.
in Journal of immunology (Baltimore, Md. : 1950)
Li H
(2021)
Embryonic Kidney Development, Stem Cells and the Origin of Wilms Tumor.
in Genes
Description | PhD student funded from MSc biomedical sciences |
Amount | £18,000 (GBP) |
Organisation | University of Edinburgh |
Sector | Academic/University |
Country | United Kingdom |
Start | 11/2014 |
End | 10/2017 |
Title | V5-beta catenin mouse ES cells |
Description | These cells carry a V5 epitope tag on the N-terminus of the endogenous beta-catenin gene. They were an intermediate step in the generation of the delSer45 mouse model in this proposal. |
Type Of Material | Cell line |
Provided To Others? | No |
Impact | These cells can be used for a variety of in vitro proteomic approaches and could be turned into mice for in vivo proteomics (not part of this proposal) |
Title | mouse embryonic stem cells with Ser33Tyr mutation in beta-catenin |
Description | This cell line was made as a control for the delSer45 mutation which was an intermediate step in the generation of the delSer45 mice from this project. |
Type Of Material | Cell line |
Provided To Others? | No |
Impact | These ES cells can be used to generate Ser33Tyr mutatn beta-catenin m ice (not part of this proposal) and can be used as an in vitro system for further analysis in their own right. |
Title | mouse embryonic stem cells with constitutive delSer45 mutation in beta-catenin |
Description | We generated mouse ES cells heterozygous and homozygous for the delSer45 mutation in the endogenous beta-catenin gene that we currently are analysing |
Type Of Material | Cell line |
Provided To Others? | No |
Impact | This ES line is an intermediate step in the generation of delSer45 knock-in mice we proposed in the project, and which can be used as an experimental in vitro system for further analysis |
Title | Additional file 1: Table S1. of Proteomic profiling of neuronal mitochondria reveals modulators of synaptic architecture |
Description | All proteins identified from synaptic and non-synaptic mitochondrial proteomic analyses. (XLSX 273Â kb) |
Type Of Material | Database/Collection of data |
Year Produced | 2017 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_1_Table_S1_of_Proteomic_profili... |
Title | Additional file 1: Table S1. of Proteomic profiling of neuronal mitochondria reveals modulators of synaptic architecture |
Description | All proteins identified from synaptic and non-synaptic mitochondrial proteomic analyses. (XLSX 273Â kb) |
Type Of Material | Database/Collection of data |
Year Produced | 2017 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_1_Table_S1_of_Proteomic_profili... |
Title | Additional file 2: Table S2. of Proteomic profiling of neuronal mitochondria reveals modulators of synaptic architecture |
Description | All proteins identified from synaptic and non-synaptic mitochondrial proteomic analyses with >2 fold-change, pâ L0.05, â L2 unique peptides. (XLSX 106Â kb) |
Type Of Material | Database/Collection of data |
Year Produced | 2017 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_2_Table_S2_of_Proteomic_profili... |
Title | Additional file 2: Table S2. of Proteomic profiling of neuronal mitochondria reveals modulators of synaptic architecture |
Description | All proteins identified from synaptic and non-synaptic mitochondrial proteomic analyses with >2 fold-change, pâ L0.05, â L2 unique peptides. (XLSX 106Â kb) |
Type Of Material | Database/Collection of data |
Year Produced | 2017 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_2_Table_S2_of_Proteomic_profili... |
Title | Understanding the molecular consequences of inherited muscular dystrophies: advancements through proteomic experimentation |
Description | Introduction: Proteomic techniques offer insights into the molecular perturbations occurring in muscular-dystrophies (MD). Revisiting published datasets can highlight conserved downstream molecular alterations, which may be worth re-assessing to determine whether their experimental manipulation is capable of modulating disease severity. Areas covered: Here, we review the MD literature, highlighting conserved molecular insights warranting mechanistic investigation for therapeutic potential. We also describe a workflow currently proving effective for efficient identification of biomarkers & therapeutic targets in other neurodegenerative conditions, upon which future MD proteomic investigations could be modelled. Expert commentary: Studying disease models can be useful for identifying biomarkers and model specific degenerative cascades, but rarely offer translatable mechanistic insights into disease pathology. Conversely, direct analysis of human samples undergoing degeneration presents challenges derived from complex chronic degenerative molecular processes. This requires a carefully planed & reproducible experimental paradigm accounting for patient selection through to grouping by disease severity and ending with proteomic data filtering and processing. |
Type Of Material | Database/Collection of data |
Year Produced | 2016 |
Provided To Others? | Yes |
URL | https://tandf.figshare.com/articles/dataset/Understanding_the_molecular_consequences_of_inherited_mu... |
Title | Understanding the molecular consequences of inherited muscular dystrophies: advancements through proteomic experimentation |
Description | Introduction: Proteomic techniques offer insights into the molecular perturbations occurring in muscular-dystrophies (MD). Revisiting published datasets can highlight conserved downstream molecular alterations, which may be worth re-assessing to determine whether their experimental manipulation is capable of modulating disease severity. Areas covered: Here, we review the MD literature, highlighting conserved molecular insights warranting mechanistic investigation for therapeutic potential. We also describe a workflow currently proving effective for efficient identification of biomarkers & therapeutic targets in other neurodegenerative conditions, upon which future MD proteomic investigations could be modelled. Expert commentary: Studying disease models can be useful for identifying biomarkers and model specific degenerative cascades, but rarely offer translatable mechanistic insights into disease pathology. Conversely, direct analysis of human samples undergoing degeneration presents challenges derived from complex chronic degenerative molecular processes. This requires a carefully planed & reproducible experimental paradigm accounting for patient selection through to grouping by disease severity and ending with proteomic data filtering and processing. |
Type Of Material | Database/Collection of data |
Year Produced | 2016 |
Provided To Others? | Yes |
URL | https://tandf.figshare.com/articles/dataset/Understanding_the_molecular_consequences_of_inherited_mu... |
Title | WT1 expression in breast cancer disrupts the epithelial/mesenchymal balance of tumour cells and correlates with the metabolic response to docetaxel |
Description | RNA profiles of breast cancer cells (MDA-MB-157) were generated by deep sequencing on the Illumina HiSeq 2000 platform. Untreated MDA-MB-157 cells, MDA-MB-157 cells transduced with a lacZ control vector, and MDA-MB-157 cells transduced with a lentiviral vector carrying a Wt1 shRNA were sequenced (titled untreated, lacZ and Wt1 respectively). |
Type Of Material | Database/Collection of data |
Year Produced | 2017 |
Provided To Others? | Yes |
Description | 7th course Developmental Biology, Institute Curie, Paris |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Postgraduate students |
Results and Impact | I gave a talk at the 7th international course for developmental biology, organised by the Institue Curiein Paris |
Year(s) Of Engagement Activity | 2016 |
Description | BDSRA - Patient family conference |
Form Of Engagement Activity | A formal working group, expert panel or dialogue |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Patients, carers and/or patient groups |
Results and Impact | Patient family conference for Batten Disease Scientific Research Association (USA based charity). Out of hours presentation on model systems for pre clinical investigation to family members of patients, other researchers and industry.. Approx 15 other short talks took place. |
Year(s) Of Engagement Activity | 2020 |
Description | Blog post NC3Rs |
Form Of Engagement Activity | Engagement focused website, blog or social media channel |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | Wrote blogpost about how transgenic technologies can help the 3Rs. |
Year(s) Of Engagement Activity | 2017 |
URL | https://www.nc3rs.org.uk/news/when-3rs-and-transgenic-technologies-meet |
Description | Coming of Age: The Legacy of Dolly at 20 public lecture |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Public/other audiences |
Results and Impact | At a series of public lectures to celebrate the 20th birthday of Dolly the Sheep I discussed kidney research with the public |
Year(s) Of Engagement Activity | 2016 |
Description | Consensus Meeting on Renal Progenitors and Kidney Regeneration |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | I gave a talk at this meeting organised to discuss the many different views in the field about renal progenitor cells. |
Year(s) Of Engagement Activity | 2016 |
Description | ILAR Roundtable 'gene editing to modify naimal genomes for reserarch - scientific and ethical considerations' |
Form Of Engagement Activity | A formal working group, expert panel or dialogue |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Policymakers/politicians |
Results and Impact | Round Table organized by the National Academy of Sciences in the USA to discuss the impact of genome editing on animal use in science. |
Year(s) Of Engagement Activity | 2015 |
URL | http://nas-sites.org/ilar-roundtable/roundtable-activities/gene-editing-to-modify-animal-genomes-for... |
Description | Invited seminar Cincinnati Children's Hospital Medical Center |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Invited visit to CCHMC to give seminar and discuss future collaborations. |
Year(s) Of Engagement Activity | 2017 |
Description | Invited seminar LUMC Leiden, the Netherlands |
Form Of Engagement Activity | A formal working group, expert panel or dialogue |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Invited seminar at the Leiden University Medical Center, Leiden, the Netherlands |
Year(s) Of Engagement Activity | 2017 |
Description | Invited talk at BSDB/Nordic meeting |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Invited talk at joined meeting of the British Society for Developmental Biology and Scandinavian developmental biologists |
Year(s) Of Engagement Activity | 2017 |
Description | Lobbying reception Scottish Parliament |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Policymakers/politicians |
Results and Impact | Kidney research UK organized a lobbyng reception in the Scottish parliament to discuss the importance of kidney research with MSPs, patients and other researchers. |
Year(s) Of Engagement Activity | 2016 |
Description | Organiser and speaker at 3rd international workshop on the biology of WT1 |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Organized the 3 international workshop in the biology of WT1 and spoke on the meeting |
Year(s) Of Engagement Activity | 2015 |
Description | Roslin Institute Open Day |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Public/other audiences |
Results and Impact | I was involved in discussion about the use of animals in research as well as presenting the imaging work from my own laboratory. |
Year(s) Of Engagement Activity | 2017 |
Description | Talk at MRC Toxicology Unit, Leicester |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Professional Practitioners |
Results and Impact | Invited seminar |
Year(s) Of Engagement Activity | 2016 |
Description | Talk at Nephrotools meeting |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Ta\lk at a meeting of the EU funded Nephrotools consortium |
Year(s) Of Engagement Activity | 2015 |
Description | Teaching at 10th Advanced Paediatric Oncology Course (APOC), Edinburgh, United Kingdom |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Teaching at this course for paediatric oncology nurses |
Year(s) Of Engagement Activity | 2017 |
Description | Visit Kidney research UK delegation to Edinburgh |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Other audiences |
Results and Impact | A delegation from Kidney research UK visited Edinburgh to discuss renal research with all kidney researchers |
Year(s) Of Engagement Activity | 2016 |
Description | Visiting undergrad students Wyoming |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Undergraduate students |
Results and Impact | I discussed the effects of Brexit on science funding and networking with visiting undergrad students from Wyoming |
Year(s) Of Engagement Activity | 2018 |
Description | Welcoming committee |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Public/other audiences |
Results and Impact | At the open doors day Roslin Institute I was welcoming visitors to the building, while my lab was demonstrating the culture of embryonic mouse kidneys on our time-lapse imaging system |
Year(s) Of Engagement Activity | 2016 |
Description | hosting seminar Andy McMahon |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Professional Practitioners |
Results and Impact | hosted an external seminar in the institute |
Year(s) Of Engagement Activity | 2015 |
Description | hosting seminar Robert Kelsh |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Professional Practitioners |
Results and Impact | hosted external seminar from Robert Kelsh |
Year(s) Of Engagement Activity | 2015 |
Description | in conversation with session on Roslin Open Doors day |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Public/other audiences |
Results and Impact | General public had 'in conversation with' sessions with scientists from The Roslin Institute to discuss their science. I have focused my sessions on the use of animals in research, and in kidney development and disease in particular. |
Year(s) Of Engagement Activity | 2014 |
Description | invited seminar at Crick Institute Mill Hill |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Professional Practitioners |
Results and Impact | invited seminar |
Year(s) Of Engagement Activity | 2016 |
Description | regenerative Medicine Therapy network development meeting |
Form Of Engagement Activity | A formal working group, expert panel or dialogue |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Professional Practitioners |
Results and Impact | A meeting to discuss the development of a national network on Renal Regenerative Medicine with the support of Kidney Research UK |
Year(s) Of Engagement Activity | 2016 |
Description | student talk at high school |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | talk at local high school discussing animal use in science in the context of kidney development, kidney stem cells, kidney disease and imaging |
Year(s) Of Engagement Activity | 2015 |
Description | visit Biomed students Wyoming |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Undergraduate students |
Results and Impact | A group of undergrad students from Wyoming visited the institute and I gave a talk about 'being a scientist' and our kidney research in particular |
Year(s) Of Engagement Activity | 2017 |