School of Clinical Medicine

Most of the staff of the Department of Medical Genetics have their research groups in the Cambridge Institute of Medical Research (CIMR). This is a cross-departmental institute which provides a unique interface between basic and clinical science with the major goal of determining and understanding the molecular mechanisms of disease.

The department is also closely associated with the NHS East Anglian Medical Genetics Service at Addenbrooke’s Hospital.

Research ranges from the functional biology of genetic disorders to molecular diagnostics and novel approaches to therapy and can be grouped into three broad topic areas:

• Diabetes and inflammation (Professor Todd, Professor Wicker, Professor Clayton and collaborators);
• Neurological disorders (Professor Rubinsztein, Dr Raymond, Dr Woods, Dr Reid);
• Renal disorders (Professor Karet and Dr Sandford)

More detailed information on the various research groups within the department is set out below

Our aim is to discover the molecular basis for the autoimmune inflammatory disease type 1 (insulin-dependent) diabetes.  We use an integrated combination of genetics, in large collections of type 1 diabetic families and case/control, statistics, genome informatics and data mining, and gene expression and functional studies. 

Our major effort now is to correlate susceptibility genotypes with biomarkers and phenotypes e.g. we have correlated plasma levels of the soluble form of the interleukin-2 receptor with the genotypes of the IL-2RA gene that are associated with type 1 diabetes susceptibility.  This is a first step towards identifying disease precursors that could be used in the evaluation of future therapeutic studies.  To achieve this we have helped build a local biobank of healthy volunteers in whom we can study the effects of disease-associated genotypes (The Cambridge BioResource: www.cambridgebioresource.org.uk/). 

Our research efforts are part of the Juvenile Diabetes Research Foundation/Wellcome Trust Diabetes and Inflammation Laboratory, which includes the laboratories of Linda Wicker and David Clayton, as well as collaborations with the Department of Haematology, the Department of Paediatrics (David Dunger), the Wellcome Trust Sanger Institute and the MRC Epidemiology Unit.

Our aim is to develop and apply statistical methods in genetic epidemiology and, to a lesser extent, in other aspects of biostatistics. The main focus of this work is provided by the Diabetes and Inflammation Laboratory (DIL), although I have long-standing interests in studies into the genetic epidemiology of other complex diseases, including hypertension, and senile macular degeneration. Implementation and dissemination of analytical methods as an important part of our work. A major commitment over the last two years has been the Wellcome Trust Case-Control Consortium (WTCCC). This is a genome-wide association (GWA) study in which 2,000 cases of each of 7 different diseases will be compared with two shared control groups, together comprising 3,000 subjects. Each subject was typed for ~500,000 SNPs. I co-chaired the analysis group and we have developed analysis software, which we have made freely available on the Internet. My group has also made a major contribution to the management of the huge quantities of data generated by this study. The main results of the study were published in early 2007. The WTCCC has been remarkably successful in identifying new disease susceptibility loci, particularly for type 1 diabetes, and has generated much follow-on work. Fine mapping of causal variants remains a considerable challenge and refinement of methods for the design and analysis of fine mapping studies remains a high priority for my group. In addition the Type 1 Diabetes Consortium has funded a further genome-wide association study of 4,000 new cases and 2,500 new controls (all drawn from the existing DIL collections) and my group will be responsible for the analysis of the new data and for a combined analysis of the new data in conjunction with the existing WTCCC T1D data (2,000 cases and 3,000 controls). I will also provide advice and software for several proposed GWA association studies.

Our group is focused on understanding the molecular and cellular mechanisms of autoimmune syndromes such as type 1 diabetes (T1D) by identifying and characterizing the function of genes that contribute to disease susceptibility in both humans and mice. 

Although the MHC region remains the major genetic determinant causing T1D in humans and nonobese diabetic (NOD) mice, the identification of non-MHC genes contributing T1D susceptibility has made significant progress in recent years.  It is now apparent that a conservative estimate of the number of T1D regions is at least 50 in both species, with most regions having plausible candidate genes that could alter immune function.  Importantly, overlap with T2D causal regions has not been observed whereas overlapping associations are present in the case of other autoimmune diseases such as RA and Crohn’s.  Humans and NOD mice have variations in some of the same T1D genes or gene pathways and there are data in both species that an early autoimmune response to insulin and other beta cell proteins that is caused by a partial failure in central and peripheral tolerance mechanisms precedes beta cell destruction.  In addition to sharing structural aspects of the MHC class II molecules that confer T1D susceptibility or resistance, variation of two molecules that negatively regulate T cells, CTLA-4 and the tyrosine phosphatase encoded by PTPN22/Ptpn22, are also associated with T1D in both species.  T1D-associated gene variants encoding the interacting molecules IL-2 and the alpha chain of the IL-2 receptor (CD25), which are members of a pathway essential for immune homeostasis via peripheral tolerance, are present in mice and humans, respectively. Signalling through the IL-2 receptor is critical for the function of FOXP3+CD25+CD4+ regulatory T cells, a T-cell subset that dampens the immune response.  Importantly, several groups have observed that the FOXP3+CD25+CD4+ regulatory T cell subset has reduced function in T1D patients and NOD mice.  We are examining FOXP3+CD25+CD4+ regulatory T cells in the peripheral blood of T1D patients and healthy volunteers accessed through the Cambridge BioResource.  Using multi-colour flow cytometry, expression levels of molecules associated with the function of regulatory T cells, most notably CD25 and FOXP3, are being correlated with gene variants that cause T1D susceptibility. 

Laboratory members are also examining CD25 expression levels in immune cell subsets that upregulate CD25 after activation, rather than having constitutive expression, as is the case for FOXP3+CD25+CD4+ regulatory T cells.  In analysing the immune responses of IL-2 congenic mice developed in the laboratory, we observed that an approximately 2-fold increase in IL-2 production is associated with the presence of a T1D resistance allele at Il2.  Augmented FOXP3+CD25+CD4+ regulatory T cell function and a decreased ability of islet-specific effector CD8+ T cells to proliferate in the draining lymph nodes of the pancreas and to subsequently migrate to the islets also positively correlated with T1D resistance alleles.  The T1D-resistant IL-2 alleles also increased the ability of costimulation blockade to induce tolerance to allogeneic and xenogeneic islet transplants even when the studies are performed in a mouse strain that does not develop autoimmunity. 

Our findings likely explain the observations that the IL-2 gene region controls susceptibility to thymectomy-induced ovarian dysgenesis and that neutralizing anti-IL-2 antibodies can induce autoantibodies as well as cellular infiltrates in a variety of tissues.  We hypothesize that there has been selective pressure to maintain optimal levels of IL-2 production since excessive IL-2 could reduce immune responsiveness via the induction of overly potent regulatory T cells, whereas too little IL-2 increases the likelihood of autoimmune responses to reproductive or other vital organs because effector mechanisms are not held in check by regulatory T cells. The elucidation of the molecular consequences of the causative single nucleotide polymorphisms (SNPs) in the genes encoding CTLA-4, IL-2 and CD25 are also under investigation in the laboratory.  For example, we have discovered that the causative SNP in mouse CTLA-4 modulates the amount of a CTLA-4 protein isoform that is produced by altering the efficiency of an alternative splicing event that must occur to produce the isoform’s mRNA.  In contrast, variation of the mouse gene encoding IL-2 in extended regions 5´ and 3´ of the gene led us to test the hypothesis that one or more of the SNPs alters chromosome accessibility mediated by epigenetic modification, thereby affecting expression levels of the mRNA.

Our current efforts mainly aim to characterise molecular mechanisms governing human renal acid-base homeostasis, the fine regulation of which is the chief job of a-intercalated cells (a-IC) in the distal nephron. Intact a-IC functions (secretion of protons in to the urine via an apical multi-subunit H+-ATPase, coupled to bicarbonate reclamation via the basolateral Cl-/HCO3- exchanger AE1) are necessary for appropriate excretion of the net acid load of a normal diet, and for generation of adequate amounts of bicarbonate for buffering. However, neither the identity of all the transporters, pumps and channels responsible, nor the regulatory pathways involved, are yet well understood. To elucidate the relevant physiology, we adopted an initial genetic approach, studying rare single-gene disorders (the distal renal tubular acidoses, dRTAs) where a-IC function is inadequate, imparting large quantitative effects on the kidney’s ability to maintain normal body fluid pH. DRTA is phenotypically defined by hyperchloraemic metabolic acidosis, abnormalities of bone density and calcium deposition in the renal tract. The recessively inherited syndromes present with very severe changes at a young age and sensorineural hearing loss (SNHL) is often associated.

Identification of responsible genes in these naturally occurring “human knockouts” provided the springboard for our subsequent functional studies of both wild type and mutant gene products. We have described mutations in the AE1 gene in dominant dRTA, and loss-of-function mutations in genes encoding two of the renal proton pump's subunits (B1 and a4) in recessive disease. For the H+ATPase, we have shown that organization of the pump complex involves physical interaction between the a- and G subunits and the glycolytic enzyme PFK-1. Without the latter, coupling between ATP hydrolysis and proton translocation is disrupted. We have also discovered and characterized four novel genes, encoding kidney-specific isoforms of the proton pump's C, G, d- and e-subunits. Our functional investigations have centred on AE1.

We have demonstrated that the dominant negative mechanism in at least two variants of disease is unusual, involving mis-targeting of mutant protein away from its normal basolateral location in polarized epithelial cells. Recently we have identified glycolytic enzymes as binding partners for AE1 in the kidney. Other studies are focusing on additional non-renal phenotypes involving these molecules, identifying the molecular pathways responsible for various forms of renal tubular disease, and clinically-based research.

We aim to identify mutations in novel genes that result in significant intellectual disability in humans. We also aim to understand the mechanisms by which the intellectual disability occurs. Two strategies are been used to identify these disease-causing genes. Firstly, work is in progress to identify DNA sequence at the breakpoints of balanced reciprocal X;autosome translocations. The technique is powerful but limited by the number of patients who carry these rare chance rearrangements and thus cannot be used as a systematic strategy to identify all X linked disease-causing genes. Nevertheless, it remains an invaluable approach for identifying disease causing genes on the X chromosome and our group is working on a number of these translocations at the present time. Secondly, in collaboration with The Wellcome Trust Sanger Institute, we are using a new approach to disease gene identification using systematic searches for mutations through the whole of the X chromosome. We have established a substantial collection of samples from families with X linked mental retardation (XLMR) by collaborating with genetics centres throughout the UK, Ireland, Australia, USA and Europe. We are performing high throughput DNA sequencing analysis on affected individuals from each of the families. The strategy has been made possible by the availability of the working draft human genome sequence and the development of computer based mutation detection systems. The approach will allow gene identification in those families where linkage/cytogenetic data is insufficient or unavailable and will rapidly extend the assignment of genes on the X chromosome to a mental retardation phenotype.

We are studying the pathogenesis of diseases caused by codon reiteration mutations, like Huntington's disease (HD) and oculopharyngeal muscular dystrophy (OPMD), which result from abnormally elongated polyglutamine and polyalanine codon stretches in the HD and PABPN1 genes, respectively. These diseases are associated with intracellular aggregate formation. We are addressing the following questions:

1. What are the early pathological changes that occur in HD and other codon reiteration diseases? We have developed a variety of models of HD, OPMD and related diseases in cells, flies (in collaboration with Dr Cahir O'Kane), zebrafish (in collaboration with Daniolabs/Dr Paul Goldsmith/Prof Bill Harris) and mice. We have been studying these with different approaches, including cDNA microarray analyses, reporter gene studies and biochemical techniques identifying huntingtin-interacting proteins. Data from cell models are then confirmed in vivo using transgenic animals and samples from HD patients.
2. What are the genetic pathways that modify polyglutamine toxicity? About 70% of the variance in the age-at-onset of HD can be accounted for by CAG repeat number in the disease-causing allele. The residual variance in age at onset unaccounted for by the CAG repeat numbers is likely to be partly due to genetic factors, because for a given CAG repeat number, age-at-onset appears to be more similar between siblings compared to unrelated individuals. Thus, even in a Mendelian disease that is associated with a high penetrance “deterministic” mutation, there are likely to be other genetic modifiers. We are using genetic approaches in mice (with Steve Brown, Harwell) and flies (with Cahir O’Kane, Cambridge) to identify such modifiers. Identification of such pathways can give clues to potential therapeutic strategies.
3. Can one attenuate polyglutamine toxicity by inducing autophagy? The polyglutamine expansion confers a novel toxic novel function on huntingtin. Thus, it is important to understand how its levels are regulated. We have shown that mutant huntingtin fragments are autophagy substrates and that autophagy upregulation is protective against mutant huntingtin toxicity in fly and mouse models. Recently we have shown that this strategy may have wider relevance to a range of polyglutamine diseases (including certain spinocerebellar ataxias), to mutant forms of a-synuclein that cause autosomal dominant forms of Parkinson’s disease, and also to mutant forms of tau that cause fronto-temporal dementias (in cells and flies). Since rapamycin is designed for chronic use in humans, this may represent a possible therapeutic strategy for a wide range of neurodegenerative diseases associated with aggregate-prone proteins. Currently, the only autophagy-inducing drug that is known to reduce mutant huntingtin levels effectively in mammalian brains is rapamycin. While it is designed for long-term use, it has significant side-effects and is not that well-tolerated. Mammalian autophagy is not that well understood, either in terms of the machinery or the signalling that impacts on this pathway. Thus, we have been trying to characterise this process better, with the hope of identifying possibly safer ways of inducing autophagy. Our data confirm that microtubule function is crucial for autophagy and that dyneins are the microtubule-associated motor proteins that facilitate autophagosome-lysosome fusion. We have also described new signalling pathways that regulate autophagy. For instance, we discovered that IP3 levels regulate autophagy via an mTOR/rapamycin-independent pathway.
4. Are there common mechanisms causing pathology in the different diseases associated with intracellular protein aggregation? It is important to test if the different diseases associated with intracellular aggregate formation share common pathways, as this may inform the fundamental understanding of the relationship between the aggregation process and cell dysfunction/death. We use cell and fly models of HD, OPMD and forms of Parkinson's disease, to compare pathological processes and test if strategies that are protective in polyQ disease models are also beneficial in other diseases associated with abnormal protein aggregation. We have developed an OPMD transgenic mouse model that has allowed us to be the first lab to test possible therapeutics for OPMD in vivo - we have shown beneficial effects with doxycycline and trehalose.

The main focus of my group is the investigation of the molecular pathogenesis of autosomal dominant polycystic kidney disease (ADPKD), a common, inherited kidney disease that frequently leads to kidney failure and is associated with severe cardiovascular complications. Genes causing this disease encode a family of proteins called the polycystins. Polycystin-1 is a large cell-surface protein of unknown function. Using biochemical, molecular and structural techniques we aim to identify extracellular and intracellular ligands and show how they interact with the wide variety of protein domains found in polycystin-1. In addition, mechanisms that target polycystin-1 to the cell surface are being identified. The polycystins have been shown to function as a mechanosensitive cell surface calcium channel localised to the renal primary cilium. We have recently shown that members of the classical receptor protein tyrosine phosphatase family interact with the polycystins in the primary cilium which identifies a novel mechanism for mechanical regulation of ion channel function. Further work is focusing on identifying substrates for these phosphatases and also other members of this large protein complex. It is hoped that the elucidation of the normal function of polycystin-1 will identify key steps in the pathogenesis of ADPKD and potential ways of modifying disease progression.

The research areas are:

1. Identifying autosomal recessive disease genes using autozygsoity mapping and e-cloning; predominantly causing mental and physical handicap in the Northern Pakistani population.
2. Defining the phenotype, genetic heterogeneity and genes that cause autosomal recessive primary microcephaly (MCPH).
3. Determining the expression pattern, cellular interactions and mitotic functions of MCPH genes.
4. Determining the evolutionary changes in MCPH genes through the primate lineages to lower mammals.

Our predominant focus is MCPH, which appears to be a primary disorder of neurogenic mitosis. The MCPH brain is small but architecturally normal and the only phenotype is mental retardation – which can be mild to severe. The MCPH genes seem to act in the neuro-epithelium lining the interior of the brain, and from which the majority of neurones arise in foetal life. Our initial focus was to find the MCPH genes, now we are trying to find what these genes do and how perturbation of this process leads to a small human brain. Two major unexplained problems in neurogenesis are: how does the neuroepithelium containing the neural precursors know how many neurones to produce; and how is the switch from symmetric (progenitor expansion) to asymmetric cell division (which generates neurones) controlled. It is likely that MCPH is a disease of one of these processes, but which one? This work benefits families, as we can offer DNA testing. But also it may help unravel the complexities of neurogenesis, with implications for neural stem cell therapy. Finally, it appears that the same MCPH genes that when mutated can make a small brain, are also involved in making out species brain so large. Studies of the MCPH1 and MCPH5 genes show that they have undergone positive selection in the primate lineage. Throughout the primate lineage there has been a step wise increase in brain size, culminating in our brains being three times the relative size of our nearest ape relatives, the chimp and gorilla. Only Dolphins have a brain almost as large as our!

Our research is focused on the hereditary spastic paraplegias, genetic conditions in which the corticospinal tract axons degenerate. HSPs selectively involve axons while sparing the neuronal cell bodies, so we study them to understand molecular mechanisms crucial for axonal maintenance and degeneration.

We want to understand both the normal functions of HSP proteins and how these functions are disrupted in the disease.  An emerging theme in the HSPs is the involvement of many of the disease proteins in membrane traffic processes.  Our work concentrates on understanding the functions of this particular subgroup of HSP proteins and in this we are greatly helped by strong interactions with the membrane traffic community within CIMR.  Our work is based on three main themes:

1. Understanding the functions of spastin and atlastin.  These proteins are involved in processes at the interface between membrane traffic and microtubule regulation and we have shown that they are binding partners, strongly suggesting that they are functionally related. We are examining functional assays for selected membrane traffic pathways in cell models of spastin- and atlastin- HSP.
2. Understanding the role of spartin.  Spartin is mutated in Troyer syndrome, a type of complicated HSP.  We are exploring the role of spartin at endosomes using a variety of functional assays in cellular models.
3. Understanding the function of NIPA1.  This project builds on data generated from fly models, and is carried out in collaboration with Dr Cahir O’Kane in the Department of Genetics.  Its aim is to examine whether, like its fly homologue, mammalian NIPA1 is involved in Bone Morphogenic Protein signaling and if so, how it regulates this signaling pathway and how this could cause axonopathy.