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New research from MRC Harwell has used the exciting CRISPR-Cas9 technology to repair a mutation known to be involved with progressive age-related hearing loss

The International Mouse Phenotyping Consortium (IMPC) is working to decipher the functions of every gene in the mouse genome, by removing one gene at a time and observing what happens to the mice born without that gene. The idea is that if we see an abnormality in those mice then we can deduce that the gene which is missing is linked to the abnormality, or “phenotype”.

Research on mice around the world is usually   carried out using a number of specific “strains”. These strains are mice that have been inbred for many generations creating a very defined set of genes. However, some of the commonly used strains carry mutations which developed at random, so-called spontaneous mutations. Because all mice belonging to each strain are only bred with other mice from the same strain, all mice in that strain will carry the same mutation.

A common mutation is the ahl (age-related hearing loss) mutation in a gene called Cadherin23, or Cdh23. The ahl mutation causes mice to progressively lose their hearing as they get older, like humans who suffer from age-related hearing loss. The ahl mutation is found in several inbred mouse strains, including the C57BL/6NTac strain used for the IMPC at MRC Harwell. When our mouse strain carries a mutation such as ahl, this can cause problems deciphering the function of genes. If we remove a gene and notice that our mouse has problems with its hearing, it is difficult to assess whether it is due to the gene we have removed or because of the ahl mutation that mouse strain already carried.

Now, a new study from MRC Harwell published in Genome Medicine on 15th February 2016 has used the CRISPR-Cas9 gene editing technique to ‘repair’ the Cadherin23 ahl mutation in our C57BL/6NTac mouse strain.

The ahl mutation is a single nucleotide polymorphism in the Cadherin23 gene. This means that, of the over ten thousand DNA bases that make up Cdh23, just one base has changed: an adenine (A) replaces a guanine (G). This single change has huge repercussions for the protein produced by the Cdh23 gene, resulting in a section of the protein being missing, and predisposing mice carrying the mutation to age-related hearing loss as the hair cells in their ears degenerate as they get older.

The team at MRC Harwell used the CRISPR-Cas9 gene editing technique to ‘repair’ this mutation. The CRISPR-Cas9 system was originally identified in bacteria as part of their immune system. This technique allows scientists to precisely snip out a section of DNA and replace it with another of their choosing: the team at MRC Harwell were thus able to replace the incorrect DNA base in the ahl mutation with the correct base, in single-cell mouse embryos.  The adult mice resulting from these embryos carried a ‘repaired’ Cdh23 gene and, importantly, they did not suffer from the same progressive hearing loss as the animals that carry the ahl mutation.

This research is important for several reasons. Primarily, the strains generated by IMPC can now be bred with the mice with a ‘repaired’ Cdh23 gene, meaning the IMPC gene mutations can be used to characterise deafness phenotypes in the future. Mouse studies such as this one also demonstrate a great potential for the use of CRISPR-Cas9 to repair genetic mutations in mice and other organisms.

On Wednesday 3rd February 2016, we welcomed a group of Sixth Form Applied Science students from St Birinus School and Didcot Girls’ School.

The visit started with a tour of the Mary Lyon Centre. After putting on a sterile suit and going through our air showers, the students were shown the mouse facility, including the high-tech specially designed cages complete with their own ventilation system where the mice are housed in small groups to keep each other company. They also had the chance to see our animal technicians in action, ensuring that each and every mouse in the facility is checked every day of the year, including Christmas and Boxing Day. After re-emerging from the Mary Lyon Centre, the students came over to the Mammalian Genetics Unit to see some of our labs putting the other side of our research into practice. In particular, the students spent some time in histology and microscopy, where tissues are examined at a cellular level to identify small changes which might not be noticed without magnification.

The students then moved into our special teaching lab to do some work of their own! As usual, our DNA extraction from strawberries activity went down very well. As Nanda Rodrigues said, “making DNA from strawberries always grabs their attention…it’s always satisfying to be able to explain what we do and to see their faces light up with enthusiasm and wonder”. Finally, after having witnessed many aspects of working at MRC Harwell, the students sat down with some of our researchers to discuss careers in the biomedical sciences.

The students seemed to thoroughly enjoy their day at MRC Harwell, and their teacher Mrs Wall said “I have mentioned it in about 4 lessons now as different aspects of the visit were relevant to them!”. Several of the students are now very keen to return to MRC Harwell for work experience. A successful day all around!

Richmond village talk by Dominic Norris

Dominic Norris visited Richmond Letcombe Regis Retirement Village to talk about his research on cilia

On 14 January 2016, Dominic Norris visited Richmond Letcombe Regis Retirement Village to talk about his research on cilia. He began by taking them on a journey through the history of microscopy, getting them to think about the cell, and how we use microscopy to see its components. Most people think that the detail that can be seen through a microscope is determined by the magnification power, but it‘s actually more commonly the resolution – being able to distinguish two close together objects from one another. Dominic gave the example of a pair of headlights, which when far away can appear to be a single spot of light, but once they loom into view can be distinguished as two distinct lights. He spoke about how the invention of the electron microscope has given us the resolution required to see the components of a cell, including his own research interest – cilia.

The next part of his talk went into what these cilia are, what they do and what happens when they don’t function properly. He showed video clips of tiny hair-like structures at the edge of the cell waving in unison. He used the example of cilia that brush dust and bacteria out of our lungs, and explained how one of the many reasons why they stop working can be due to diseases, such as Primary Ciliary Dyskinesia (PCD).

At this point he began describing the symptoms of PCD, which are not only limited to trouble clearing the lungs, but also relate to the many other roles motile cilia play in the body. To explain one of the more drastic of these symptoms, he took out a Mannequin showing the internal arrangement of organs in a human body.

This, he said, was how the body is normally arranged, pointing out how organs such as the heart have a set orientation, which allows them to fit in with the rest of the organs and function effectively. In some PCD patients, this is completely switched around to have a mirror image of the usual organ arrangement. He explained how, once again, this was due to malfunctioning cilia, this time during the early stages of development. These cilia swirl to create a flow that the embryo uses to determine which side of the body is left and which is right. Without this flow, organ arrangement goes awry.

The talk was very well received by the residents, with a good level of engagement and plenty of questions, and we hope to arrange more events with them in the future.   

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Deafness due to mismatched connections

Loss of a functional Neuroplastin gene in mice has been found to result in problems pairing hair cells with neurons in the inner ear, preventing sound being detected by the brain

 Normally hair cells in the inner ear of a mouse each pair up with a neuron (left), but in mice lacking a functional Neuroplastin gene, mismatches occur (right)

For the very first time, researchers at MRC Harwell have shown that the Neuroplastin gene is necessary for hearing in mice. Their findings, published in The Journal of Neuroscience on 6 January 2015, show that loss-of-function mutations in the Neuroplastin gene in mice mean the inner hair cells of the ear don’t match up properly with the neurons that transmit the signal to the brain, causing these mice to be profoundly deaf. It provides a new possible cause of deafness in people.

While mice are tuned towards higher pitched sounds than we are, the basic mechanism for how their hearing works is much the same; the sound travels along the ear canal as vibrations until these reach the inner ear, where they cause specialised structures on the upper surface of the inner hair cells to flex, pulling open ion channels at their tips and triggering electrical impulses along connecting neurons to the brain.

Around the time of the onset of hearing in mice, the researchers found that one of the two possible protein products from the Neuroplastin gene, Np65, is produced in the inner ear. Np65 is found at the base of inner hair cells, the region where they form synapses with neurons. Normally, a sound triggers the release of the neurotransmitter glutamate at these sites, which crosses the gap to pass on the signal to the connecting neurons and on to the brain.

However, the researchers found that when the function of the Neuroplastin gene was lost, the inner hair cells and neurons didn’t pair up properly. This suggests that Neuroplastin may be important for this matching process, either by helping to target neurons to inner hair cells, or by tethering them together. So while the sound vibrations are detected by the ears of the mice, the signal never reaches their brain, meaning that mice without a functional Neuroplastin gene are deaf from a very young age.

Identification of a new genetic cause of deafness, such as this, tells us more about the normal process of hearing and the essential components required. In the UK, more than 11 million people are affected by hearing loss. This newfound knowledge might one day be used to determine the exact cause of a person’s deafness, so they can receive the most appropriate treatment. 

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Mary Lyon Centre archiving promoted by NC3Rs

The National Centre for the Replacement, Refinement & Reduction of Animals in Research (NC3Rs) have run a web article and newsletter link promoting our mouse line archiving services

The NC3Rs, a UK-based organisation dedicated to promoting the 3Rs for the use of animals in research, posted a news article on their website about the MLC’s mouse line archiving and distribution service on the 9th December 2016. This service helps to reduce the number of mice required by storing stocks of mouse lines for use in research.

The non-profit archive at the Mary Lyon Centre is the UK archiving node for the European Mouse Mutant Archive (EMMA), and supports scientists from all around the world. Archiving a mouse line in this repository is free, with any costs recuperated from clients who order the lines.

For more information about our archiving services, please see our Frozen embryo and sperm archive (FESA) page or contact FESA.

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PhD student wins £500 for public engagement

After weeks answering a barrage of questions from curious school pupils, Sara Falcone has won her category of the national competition I’m a Scientist, Get me out of here. Image credit: I'm a Scientist (cc)

I’m a Scientist, Get me out of here is an online event, mainly funded by the Wellcome Trust, where school students can talk directly to scientists. Scientists compete for the students’ votes, answering their questions in online chats. The students ask questions, and then vote to evict scientists based on their answers. The winner is the last scientist left in their ‘zone’, and is awarded £500 to design a public engagement activity of their choice.

This time the competition ran from the 9th and 20th of November, with a total of 40 scientists and 160 schools taking part, split into eight zones. Sara entered the Ageing Zone, so answered questions based around her research in mice into the genetics of age-related diseases.

Sara is doing a PhD in Paul Potter’s group, Disease Model Discovery, which is responsible for managing the Harwell Ageing Screen, a large-scale project using mice to study the genetics of ageing and age-related diseases.

She had an eclectic mix of questions, ranging from light-hearted questions about herself, such as whether she liked to eat trifle, to huge, philosophical problems such as the meaning of life or whether we’ll ever be able to cure cancer. The students touched on big issues such as what it’s like to be a woman in science, how your lifestyle choices affect the way you age, and even whether elderly people with terminal illnesses should receive healthcare. 

Many students asked about her career path and her reasons for choosing to become a scientist. In Sara’s case this was far from a straightforward decision, showing it can sometimes take time to decide exactly what you want to do. So when a student asked ‘When did you realise you wanted to be a scientist?’ she answered:

“I always liked science, biology and medical science in particular, so I decided to study veterinary medicine because I liked medicine and animals. During the last year of uni I had to spend 6 months working in an animal hospital and I really didn’t really like it. Eventually, during my final test, one of the professors asked me ‘so, do you want to work with pets or with farm animals?’ I stopped and I thought really hard, and the only reply I could give was ‘Neither, I want to be a geneticist’. And that was my decision.”

They also asked all sorts of questions about what she does today and what she thinks of it - What do you enjoy most about your job? What’s your favourite experiment? How do you study the mice? Why do you experiment on mice rather than other animals? How will your work affect other people?

Sara answered all of the students’ questions with honesty, good humour and enthusiasm. In reward for this, she avoided being eliminated and was voted by the students as winner of the Ageing Zone. She has been awarded £500 to spend on a public engagement activity of her choice, which she’s planning to put into a new forensic, Cluedo-style activity for visitors to MRC Harwell.

PhD student wins £500 for public engagement

After weeks answering a barrage of questions from curious school pupils, Sara Falcone has won her category of the national competition I’m a Scientist, Get me out of here. Image credit: I'm a Scientist (cc)

I’m a Scientist, Get me out of here is an online event, mainly funded by the Wellcome Trust, where school students can talk directly to scientists. Scientists compete for the students’ votes, answering their questions in online chats. The students ask questions, and then vote to evict scientists based on their answers. The winner is the last scientist left in their ‘zone’, and is awarded £500 to design a public engagement activity of their choice.

This time the competition ran from the 9th and 20th of November, with a total of 40 scientists and 160 schools taking part, split into eight zones. Sara entered the Ageing Zone, so answered questions based around her research in mice into the genetics of age-related diseases.

Sara is doing a PhD in Paul Potter’s group, Disease Model Discovery, which is responsible for managing the Harwell Ageing Screen, a large-scale project using mice to study the genetics of ageing and age-related diseases.

She had an eclectic mix of questions, ranging from light-hearted questions about herself, such as whether she liked to eat trifle, to huge, philosophical problems such as the meaning of life or whether we’ll ever be able to cure cancer. The students touched on big issues such as what it’s like to be a woman in science, how your lifestyle choices affect the way you age, and even whether elderly people with terminal illnesses should receive healthcare. 

Many students asked about her career path and her reasons for choosing to become a scientist. In Sara’s case this was far from a straightforward decision, showing it can sometimes take time to decide exactly what you want to do. So when a student asked ‘When did you realise you wanted to be a scientist?’ she answered:

“I always liked science, biology and medical science in particular, so I decided to study veterinary medicine because I liked medicine and animals. During the last year of uni I had to spend 6 months working in an animal hospital and I really didn’t really like it. Eventually, during my final test, one of the professors asked me ‘so, do you want to work with pets or with farm animals?’ I stopped and I thought really hard, and the only reply I could give was ‘Neither, I want to be a geneticist’. And that was my decision.”

They also asked all sorts of questions about what she does today and what she thinks of it - What do you enjoy most about your job? What’s your favourite experiment? How do you study the mice? Why do you experiment on mice rather than other animals? How will your work affect other people?

Sara answered all of the students’ questions with honesty, good humour and enthusiasm. In reward for this, she avoided being eliminated and was voted by the students as winner of the Ageing Zone. She has been awarded £500 to spend on a public engagement activity of her choice, which she’s planning to put into a new forensic, Cluedo-style activity for visitors to MRC Harwell.

New movement disorder gene identified 

Loss of a new gene has been found to cause progressive neurodegeneration in mice, specifically in the spinal cord and peripheral nervous system. Image: muscle fibres (credit: Macroscopic Solutions)

Abraham Acevedo-Arozena has led a study into mice that lack the gene Zfp106, which develop muscle wastage and the death of neurons in the spinal cord, leading to difficulties walking, curvature of the spine and a reduced lifespan in these mice. The findings from this research, published in the journal Human Molecular Genetics, suggest that these mice could be used for research into neurodegenerative diseases such as peripheral neuropathies (PN) and Amyotrophic Lateral Sclerosis (ALS).

ALS, also known as Lou Gehrig’s disease, is the most common type of motor neuron disease – a fatal, progressive condition without cure, where patients lose the ability to move, speak and breathe.  Awareness of motor neuron disease was recently raised in the UK by the ice-bucket challenge, a social media campaign that went viral.

PN are a group of diseases affecting the nerves outside of the central nervous system. There are different forms of PN depending on the nerves affected. Collectively, they can lead to problems with movement, sensation or organ function.

Similarities to ALS and PN

The researchers found many similarities between the traits of Zfp106 deficient mice and the characteristic features of ALS and PN.

Firstly, the mice only began to show signs of muscle loss when they are a few weeks old, with these gradually worsening as they got older. It was not that the neurons did not develop properly, but that they were fine when the mice were first born and later deteriorated over time.  

Secondly, only neurons extending from the spinal cord and peripheral nervous system were affected – those in the brain, for example, did not die – particularly affected were motor neurons, which convey the electrical messages that trigger the muscles to move.

Lastly, there were similarities at the cellular level. A common feature of many neurodegenerative diseases, such as ALS and PN, is malfunctioning mitochondria, which provide energy for the cell. Some cell types, such as neurons, are more energy intensive, so are hit harder when the mitochondria can’t function properly. The researchers found signs the mitochondria were not working properly in the Zfp106 deficient mice, raising the possibility that this could be the root cause, and that Zfp106 has a role in ensuring mitochondria stay healthy.

By identifying a new gene that may be involved in ALS, PN and other neurodegenerative diseases, this research has provided a new way to investigate how and why they develop.

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New introductory training course a hit

We ran a new 2 day training course for the first time last week, ideally suited to new PhD students, ‘Introduction to Mouse Genetics & Phenotyping’, which was a great success.

Starting a new PhD can be a daunting process, with huge volumes of information to absorb and a new environment and way of working to adapt to. To kick-start the process, on 19-20 October 2015 we ran a course for those starting out in mouse genetics to introduce them to the field.

Consisting of a series of talks interspersed with workshops or group discussions, the course covered a great swathe of relevant topics. These included important considerations such as ethical approval, how to choose a suitable background strain and technique, in vivo and post-mortem phenotyping techniques, and methods for studying mutations during development and adulthood. It also pointed them towards a wealth of useful resources, including mouse line archives and databases, to identify relevant mouse models and analyse the data available on them.

Overall, the course was very popular, with 80% of the participants saying that they would recommend it. A particular highlight for many of the participants was a tour around the Mary Lyon Centre, an optional addition to the course, but all parts of the course received praise from at least one participant. We have taken on board all of the feedback and suggestions from the course, including a general feeling that it should be extended to three days to give ample time for every section. One participant requested that the course is run on an annual basis, a prospect we intend to implement.

We hope that this course has set up those who attended with a solid basis on which to build their research in mouse genetics, equipping them with the knowledge they need to go on to great things, and we look forward to welcoming a new group to MRC Harwell again next year.

To receive updates on this course, please sign up to receive training emails. We will not use your email address for any purpose other than sending you information about the courses you select.

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Taking our science into schools

As a Brilliant Club tutor, PhD student Abi Harris taught sixth form students at local state schools about her own research, giving them the competitive edge they need to get into university [Image credit: The Brilliant Club]

Your career prospects shouldn’t depend on which school you go to, but the sad truth is that in the UK, children who are privately educated are far more likely to get into a high-ranking university than those at state schools - 48% of private school kids go to a top university, compared to just 18% from state schools. The Brilliant Club is seeking to close this gap by sending PhD students and postdocs into state schools to teach university-style tutorials based on their own research. 

In February, we invited someone from the Brilliant Club to come and give a talk to our PhD students. The Brilliant Club provides training, travel expenses and pay for its tutors, so it’s an excellent way for PhD students to gain experience explaining their research and teaching, earn some extra money to supplement their stipend and inspire a new generation to follow in their footsteps.

There was a lot of interest and great attendance, and afterwards many of the PhD students decided to apply to become a Brilliant Club tutor. The application process was very competitive, but we had one, Abi Harris, who was successful. And she didn’t just do one, but two, school placements.

Abi did a whole day each week for a term, travelling to Abbotsfield High School and Swakeleys School in Hillingdon, near Uxbridge. It was a long journey, as the Brilliant Club do not yet have any placements at schools in Oxfordshire. She works in the Disorders of Sex Development group at MRC Harwell, but her tutorials covered a wide spread of other relevant topics in genetics.

Before she started, she had just three weeks to produce a complete her 8000 word course booklet. If there was anything she could have changed, she said, she would have liked more time to prepare so she could have designed more activities. Nevertheless, she still managed to include some activities, such as a card game to match up the relevant genes with a disease.

Her booklet include some great analogies to illustrate concepts, such as one to teach them about the ‘central dogma’ of molecular biology - that DNA produces the RNA required to make a protein - where someone photocopies a library book so they can use the instructions to build an IKEA chair.

Once into the main part of her placements, she ran 75 minute tutorials for small groups of three to four sixth form students at the two schools. During her tutorials, she got each student to give a presentation about one model organism, and ran a debate on the ethics of using animals in research. She explained about mutations, their role in cancer, the lab techniques used to study them and how they can lead to developmental disorders such as dwarfism, additional fingers, a ‘flipped’ organ arrangement (a condition known as situs inversus) or disorders of sex development.

Overall, Abi said she found it a really positive experience, even though it was tough starting out with two placements. She plans to start another placement next term, this time working with 11-13 year olds, and says she’s really looking forward to it.

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