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Mitochondrial dysfunction caused by protein aggregation has been shown to have an important role in neurological diseases, such as Parkinson's disease (PD). Mitochondria have evolved at least two levels of defence mechanisms that ensure their integrity and the viability of their host cell. First, molecular quality control, through the upregulation of mitochondrial chaperones and proteases, guarantees the clearance of damaged proteins. Second, organellar quality control ensures the clearance of defective mitochondria through their selective autophagy. Studies in Drosophila have highlighted mitochondrial dysfunction linked with the loss of the PTEN-induced putative kinase 1 (PINK1) as a mechanism of PD pathogenesis. The mitochondrial chaperone TNF receptor-associated protein 1 (TRAP1) was recently reported to be a cellular substrate for the PINK1 kinase. Here, we characterise Drosophila Trap1 null mutants and describe the genetic analysis of Trap1 function with Pink1 and parkin. We show that loss of Trap1 results in a decrease in mitochondrial function and increased sensitivity to stress, and that its upregulation in neurons of Pink1 mutant rescues mitochondrial impairment. Additionally, the expression of Trap1 was able to partially rescue mitochondrial impairment in parkin mutant flies; and conversely, expression of parkin rescued mitochondrial impairment in Trap1 mutants. We conclude that Trap1 works downstream of Pink1 and in parallel with parkin in Drosophila, and that enhancing its function may ameliorate mitochondrial dysfunction and rescue neurodegeneration in PD.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 642.43K | Year: 2016

Recently, a completely new way of controlling gene expression has been discovered. This has come to light after the identification of a new class of genes, which unlike most genes do not produce proteins, but instead are processed into short RNA molecules called microRNAs. There are around 1000 different microRNAs within the human genome, all of which have different effects. They work by binding to the messenger RNA of protein encoding genes and inhibiting production of the protein encoded within the messenger RNA. Each of these 1000 small RNA molecules is believed to interact with and regulate around 200 protein encoding genes, thus adding to the complexity of the regulation of the human genome. Already it has become clear that malfunction of miRNA regulation is associated with virtually all human disease, including: cancer, diabetes, and viral infections. In 2002 Science magazine called miRNA the breakthrough of the year, and these small RNA molecules have been termed the Dark Matter of the cell. MicroRNAs were only discovered in 2001 and, amazingly, already within this short period, microRNA-based drugs are in clinical trials for a number of human diseases demonstrating the usefulness of the research within this field. However, despite the rapid advances within this field, how these small RNA molecules exert their effects on protein production is currently unclear and controversial. As manipulation of these small RNA molecules is a realistic approach for the treatment of a number of human diseases, understanding how these therapeutic agents work will be critical for their development and safe use. This proposal aims to determine the mechanism by which these small RNA molecules control the production of proteins within the human body and to resolve the controversy within the field by supplying testable models which can be probed by many laboratories around the world.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Training Grant | Award Amount: 94.13K | Year: 2014

Doctoral Training Partnerships: a range of postgraduate training is funded by the Research Councils. For information on current funding routes, see the common terminology at www.rcuk.ac.uk/StudentshipTerminology. Training grants may be to one organisation or to a consortia of research organisations. This portal will show the lead organisation only.


Grant
Agency: GTR | Branch: MRC | Program: | Phase: Intramural | Award Amount: 1.86M | Year: 2012

High–energy radiation (either environmental or used in clinical practice) produces toxic responses in human body. As the main consequence of radiation exposure is cellular DNA damage, we are aiming to understand how mammalian cells deal with the damage to its DNA. We are studying a group of proteins acting together in one DNA repair pathway – the non-homologous end joining (NHEJ). This pathway is responsible for the majority of DNA repair in cells. We have managed to identify several new proteins functioning in this pathway (XLS/PAXX factor and NR4A nuclear receptors). Currently our work focuses on the question of how these new factors operate at the molecular level in the DNA repair process. We also continuously search for novel components of cellular DNA repair machinery. We believe that these findings will facilitate approaches to manage toxic outcomes after radiation exposure.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 344.26K | Year: 2015

It is well known that lifestyle factors, particularly diet and exercise, have a big effect on long-term health and lifespan. In particular, many scientific studies have established that obesity increases risk of metabolic and cardiovascular diseases and cancer. It is less well known that maternal obesity during pregnancy can permanently influence the health of her offspring including their risk of being obese and developing insulin resistance. This phenomenon, known as developmental programming, has been studied in humans largely by observational and association studies, from which it is difficult to deduce underlying mechanisms. However, laboratory animals show developmental programming as a consequence of maternal obesity that is strikingly similar to that seen in humans, and therefore provide valuable systems in which to investigate mechanisms that will also be applicable in humans. We have been studying the offspring of female mice that have been fed a high-fat/high-sugar diet during pregnancy. We have found evidence that these offspring have impaired metabolic responses to the hormone insulin that persist into adulthood and that are known to increase the risk of metabolic and cardiovascular diseases in later life. Associated with these effects on insulin sensitivity of the whole animal, in fat tissue (which has a central role in orchestrating metabolism of other tissues) we detected changes in the levels of specific proteins that mediate responses to insulin. One of these proteins, IRS-1, was already known to be a key control point for insulin action on fat and muscle cells. By comparing fat cells taken from the offspring of mothers fed high-fat/high-sugar or normal diets, we found evidence that levels of IRS-1 protein are lower in the offspring of obese mothers because the synthesis of the protein is decreased. We believe this reflects the action of a class of small RNA molecules, microRNAs or miRs for short, that inhibit the translation of messenger RNA molecules containing the information required for synthesis of IRS-1 protein. In fact in cells from offspring of mothers fed the high-fat/high-sugar diet we have shown changes in the level of one of the microRNAs (miR-126) that is known to target IRS-1. This is an exciting breakthrough that points the way for further studies that will shed new light on what is happening at a molecular level to cause adverse developmental programming, and what might be done to prevent it. There are four things we would now like to do. First, we want to identify other microRNAs whose levels are programmed in fat cells by maternal over-nutrition. Advances in genome wide RNA sequencing and array technology make this a relatively straightforward exercise. The results will inform us about the scope of such changes and, using information on potential targets that is available in databases, what the overall effects might be. Second we will carry out experiments to identify targets of programmed microRNAs including miR-126. We have done similar experiments before in other contexts so we are confident of getting clear-cut data. Third, we will investigate the role of miRNAs including miR-126 in programming IRS-1 levels, insulin sensitivity and metabolic function in fat cells. We can by manipulating (up or down) the levels of individual microRNAs. Finally we will investigate whether simple lifestyle interventions, such as increased amounts of exercise during pregnancy, can alleviate developmental programming at the level of microRNAs and proteins in fat cells, and whether this correlates with improved metabolic responses at the whole organism level. We believe these studies will significantly advance understanding of the causes and consequences of developmental programming arising from maternal over-nutrition during pregnancy, and how such programming might be ameliorated. Importantly our experiments in animal models will inform the debate on how to manage the corresponding human condition.


Grant
Agency: GTR | Branch: MRC | Program: | Phase: Intramural | Award Amount: 1.07M | Year: 2013

This project is designed to identify exactly how the chemicals in cigarette affect which genes are turned on or off in human lung cells. This will reveal in fine detail exactly how cigarette smoke affects the behaviour of these cells, and will give clues as to how cigarette smoking leads to disease in human cells. It will also enable us to learn a great deal about how toxic chemicals affect a process called translation, which is the step at which the genetic code is converted into protein; this process is highly regulated, but is only now starting to be properly studied in human diseases. Genes which are affected by cigarette smoke will be assessed as markers of disease processes in archived human lung tissue. Also, we will look at how translation is altered in lung cancer compared to more normal cultured human cells. This is potentially of great value to the treatment of lung cancer, as a number of drugs are capable of targeting the translation process directly, and there is the possibility of using these to treat lung cancer.


Grant
Agency: GTR | Branch: MRC | Program: | Phase: Intramural | Award Amount: 927.24K | Year: 2013

Cancers often arise from alterations in a large variety of proteins that make cancer cells grow uncontrollably. One of the proteins that is very frequently altered in several different cancers is p53. Normally p53 protects against the formation of tumours by turning on several processes that stop cells from growing. An altered p53 protein can no longer fulfil these protective functions and, more importantly, obtains other functions that make cancer cells spread through the body and make them resistant to chemotherapy. Only recently have we started to understand a little bit more on how mutant p53 obtains these other functions and how these lead to spreading. With the proposed work I want to understand these processes better and I will investigate whether the cellular processes that are activated by mutant p53 and lead to spreading can also be involved in the resistance to chemotherapy. An increased knowledge of these processes can in the longer term lead to better therapies to target cancers with mutations in p53.


Grant
Agency: GTR | Branch: MRC | Program: | Phase: Intramural | Award Amount: 3.77M | Year: 2012

Humans are made up of literally billions of individual cells. These cells group together to make familiar organs such as lungs, heart, liver, kidneys and the brain. In order for these cells to function they have to communicate with each other. To do this cells in our body release chemicals that act as messengers – chemical messengers. These chemical messengers can be released in to the blood, for example, where they are transported to the “target” cells. The target cells have special proteins on the surface that are able to bind to very specifically to the chemical messengers. These proteins are called receptors. When the chemical messenger binds to a receptor it causes the target cell to respond. For example, if this were a heart cell and the chemical messenger was adrenaline then the heart cell would respond by beating faster and harder. The study described here will investigate this process of chemical stimulation of receptors. We will use new animal models to work out how animal’s respond when receptors are stimulated and how we might make better drugs that target these receptors.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 494.63K | Year: 2013

G protein-coupled receptors (GPCRs) are a very large family of cell surface proteins integral to how cells and tissues control their function. Because of this certain GPCRs are the targets for many medicines used to treat disease. In recent times it has also become clear that a number of GPCRs respond to alterations in concentration of nutrients such as fatty acids. Although initially surprising this makes sense as cells need to be able to alter their function as food and nutrient availability changes. One of these GPCRs is designated GPR120. Because of the capacity of GPR120 to respond to a group of fatty acids called omega-3 polyunsaturated fatty acids, which are present in high levels in the types of oily fish that we are encouraged to eat because these fatty acids have many health benefits, there has been great interest in whether synthetic chemicals could be identified that would activate GPR120 and if so might, in the longer term, provide the basis of novel medicines. GPR120 is expressed by a number of tissues in the body, including macrophages, that are important mediators of inflammation, and both pancreatic cells, the source of the hormone insulin, and white fat cells. In recent years it has become clear that inflammation is an important contributor to the development of chronic diseases such as diabetes as well as other diseases of aging. This has further raised interest in the possibility that manipulating the activity of GPR120 might be a useful, novel approach to treat diabetes and related conditions. Although very exciting, to date many of the studies implicating GPR120 as a good target in this area of health and disease have been indirect, because the type of fatty acids that stimulate GPR120 also activate other receptors and have many other effects that are not related to this receptor. Furthermore, because the omega-3 fatty acids are also converted into other mediators by the body it is possible that some of the functions suggested for GPR120 are not actually produced this way. The work we propose in this application is designed to unravel and define fully the functions of GPR120. In the last few months we have developed and characterised the only known group of synthetic chemical ligands that act selectively at GPR120 and at sufficiently low concentrations that we can be sure their effects do require activation of GPR120. We will use these to assess how activation of GPR120 in cells including macrophages, adipocytes and pancreatic cells controls their function, their production of hormones and other mediators and their interactions with other cell types. GPCRs can respond to different ligands in multiple and sometimes in distinct ways (this is termed bias). A common feature is that the receptor is rapidly modified by the addition of phosphate groups to specific amino acids. Such phosphorylation can either limit receptor function or initiate a panoply of new signals. We wish to also explore this for GPR120. We have determined exactly which amino acids in GPR120 become modified and made a version of the receptor in which this cannot happen. We wish to assess the implications of this and to do so we will generate mice in which this altered version of GPR120 replaces the normal form. These animals will then provide cells and tissues to assess which physiological functions of GPR120 require phosphorylation and which do not. In concert with this we will also make antibodies that only identify GPR120 when it is phosphorylated and will use these to determine the extent to which the receptor is activated in different conditions, for example when mice are fed a high fat diet. Interestingly, there is a variant form of GPR120 that is only found in humans and we also define its role. The ultimate objective of our studies is to define if there is a strong case to be made in investing large amounts and time and money in developing synthetic medicines that target GPR120 as a therapeutic strategy.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 258.22K | Year: 2015

The central dogma of cell biology is the information in the genetic material in the cell, the DNA, is converted via an intermediary substrate, mRNA, into proteins. For proteins to be synthesised the mRNA must interact with a large complex called the ribosome which consists of ribosomal RNA (rRNA, which do not carry instructions to make specific proteins like mRNAs but instead provide structure and contribute to the activity of the ribosome) and proteins. Ribosomes can be thought of as molecular factories where the genetic information that is held in the mRNA is decoded and then synthesised into proteins. Ribosomes are essential for cell growth and proliferation and for a cell to divide and grow, it is necessary for the number of ribosomes to double (so that more proteins can be made). For this to occur it is essential that all the proteins that make up the ribosome (there are approximately 80 of these) are made at the same time, in the correct amount and the right amount of rRNA has to be synthesised. The rRNA is made in the nucleus within a compartment that is called the nucleolus. The protein that is used to read the sequences in the DNA and covert them into rRNA in called polymerase I (Pol I). When cells are exposed to external agents that can damage them (such a sunlight or following viral infection), they undergo a complex process where they sense the damage that is caused and the proteins in the cells act to bring about a range changes to ensure the survival of the cell. During this time the cell stops growing and dividing and one of the ways in which this is achieved is by stopping the production of rRNA by inhibition Pol I activity and stopping the production of ribosomes. When this occurs the nucleolus is disassembled and the proteins within the nucleolus enter the rest of the nucleus (the nucleoplasm). While most of the proteins in the nucleolus are simply required to make ribosomes, some of these proteins have important other functions. Our data, and that of others, suggest that non-ribosomal functions of some of the proteins in the nucleolus are associated with the cellular responses to the damage that has been caused to it by external agents. In this application we propose to study how these proteins with non-ribosomal functions allow the cell to respond to different types of injury. The results obtained from these studies could have important implications for devising new ways in which to target disease that are associated with aberrant cell proliferation such as cancers.

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