King's, DNA & the continuing story
The double helix
James Watson and Francis Crick’s paper proposing a structure for DNA in 1953 was based on undeniably inspired model-building but published without their undertaking a single experiment. Instead, the experiments underpinning those models were undertaken over the previous three years in the newly-formed Medical Research Council Biophysics Unit of King’s College London.
The discovery that DNA is a double helix and that this structure comprises the hereditary material of living cellular organisms is perhaps the most momentous of our era. The insight that the discovery provided into how human characteristics arise from our individual genes created a veritable super-highway of research, ushering in gene therapy for inherited diseases and culminating in the sequencing of the human genome. The discovery also paved the way for a whole new arena of human endeavour, the biotechnology industry.
The prime movers in obtaining the data at King’s were Maurice Wilkins (1916-2004), who had commenced pilot studies on the use of X-rays to analyse DNA structure, and Rosalind Franklin (1920-1958), who advanced the Xray resolution of DNA structure to a new level of clarity and sophistication. Wilkins studied physics at Cambridge then in John Randall’s (1905-1984) department at the University of Birmingham.
In his early years his work was involved with the Second World War, culminating in the Manhattan atomic bomb project. Partly as a reaction to the results in Japan and continuing threat to human life, he was attracted to biophysics, so that he could apply his knowledge to a more useful and intellectually attractive field. He teamed up with Randall again, first at St Andrew’s and then at King’s when Randall was invited to head the new MRC Biophysics Unit in 1946. This unit broke fresh ground, being the first to bring biologists, physicists and chemists together to study biological problems.
At that time it was not generally accepted that DNA was the cell’s genetic material, but Randall and Wilkins, persuaded that it was, pressed ahead to examine its structure by X-ray diffraction. Wilkins, working with PhD student Raymond Gosling (b 1926), later Professor of Physics in Medicine at Guy’s, obtained in 1950 the first clear X-ray diffraction pattern of DNA. Their colleague Alec Stokes (1919-2003), a physicist and mathematician, deduced that this pattern was due to a helical structure.
In this same year Randall invited Rosalind Franklin to join the unit and study the structure of DNA. Aged 30, Franklin already had a reputation as an excellent scientist; she had graduated from Cambridge in chemistry, then pursued significant work on the microstructure of graphite and coal before spending three years in Paris gaining extensive experience in X-ray diffraction. Franklin joining King’s certainly strengthened the DNA project, but not entirely through the forging of a single team as she and Wilkins, although discussing their results, regarded themselves as working on separate programmes.
Franklin took over the supervision of Gosling’s PhD thesis and they found DNA to have two forms – ‘A’ and ‘B’ – depending on water content, and with different diffraction patterns, ‘A’ being the form already characterized by Wilkins and Gosling. Meanwhile Crick and Watson were building models of DNA structure at Cambridge, there being a continual exchange of information between the two institutions. No model had yet satisfied the complex and expanding information on DNA’s 3D structure and its chemical composition.
In early 1953 Wilkins showed Watson a picture of ‘B’ DNA from Franklin’s work, seemingly without her knowing, but Wilkins believing that the information was communal. Shortly afterwards the Cambridge group received the MRC’s report on its site visit (December 1952) to Randall’s department. The new information led Crick and Watson to a further model which was viewed by Wilkins in March 1953 and recognized by him to accord with his and Franklin’s work, yet giving him new insight to DNA’s structure too. These events took place against the background of Franklin’s imminent departure to Birkbeck College, where she was planning to change her work to the structure of tobacco mosaic virus.
The latest Cambridge model was the basis of the famous publication by Watson and Crick in Nature, April 1953, proposing the structure of DNA, which appeared as a sequence of three papers on DNA (the other two being by Wilkins, Stokes and Wilson, and Franklin and Gosling, on the X-ray diffraction data and structural conclusions of DNA preparations).
Other proposals for the structure of DNA were being made, and over the next seven years Wilkins and his colleague, Herbert Wilson (b 1929), a physicist specialising in X-ray diffraction, undertook further structural studies to confirm Watson and Crick’s proposed structure of the double helix. For this work and his earlier studies on DNA, Wilkins shared with Crick and Watson the Nobel Prize for Medicine or Physiology in 1962. Franklin had died from cancer in 1957; the Nobel prize is not awarded posthumously and is not shared between more than three laureates. Readers of the autobiographies or biographies of the main contributors to the unravelling of DNA and of the many accounts of the sequence of events will recognize not only the brevity of the above account, but also Adrian Hayday’s (King’s Professor of Immunobiology) comment that ‘the story is mired in utterly compelling complexity’.
It is perhaps better to focus on the facts: the combination of work eventually resulting in the discovery of DNA’s structure and function was recognized at the time by Nature in publishing the three seminal papers back to back; the award of the Nobel Prize, which by its constitution can recognize groups as much as individuals, to the three laureates; and above all to acknowledge the benefits already achieved and yet to come from the leap in understanding of heredity.
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Building on new knowledge
This new knowledge was soon put to use with the foundation at Guy’s of the Paediatric Research Unit by the Spastics Society in 1960. Its first director was Paul Polani (1914-2006) who shortly after became the first Prince Philip Professor of Paediatric Research. He graduated from Pisa in 1938 and came to the UK to start his career as a house surgeon at Lewisham Hospital. On the outbreak of the Second World War he joined the merchant navy, but was interned when Italy declared war on the UK in 1940. He was released, and found his way, as a locum, to the Evelina Children’s Hospital, where he stayed for six years, before moving to Guy’s. From the beginning of his appointment Polani recognized that an understanding of genetics and its application to clinical observation and practice were fundamental to medicine, and especially paediatrics.
At first interested in congenital heart disease, he noted that the incidence of aortic coarctation (a narrowing of the aorta) in a congenital disorder of girls, Turner’s syndrome, was similar to that in normal boys (higher than in normal girls). Turner’s syndrome affects 1 in 3,000 births with webbing of the neck, restricted growth and failed development at puberty, and congenital cardiovascular abnormalities, particularly coarctation of the aorta. Polani’s observations on coarctation led him to think that the girls with Turner’s syndrome might be genetically males.
This was before the days of chromosomal analysis, but buccal smears on patients with Turner’s syndrome showed that they did indeed have the chromatin pattern of males. He suggested that genetically they would not be XX (female) nor XY (male) but XO, and was later proven right. Polani was the first to indicate that a chromosomal abnormality could produce a defect in sexual development, and suggested that others could occur, which is now known to be the case.
Other original observations Polani made were that an individual could have two populations of cells with different chromosomes, and the explanation of the familial form of Down’s syndrome.
On his retirement in 1982 he was succeeded by Martin Bobrow (b 1938), who held the chair until moving to Cambridge in 1995. His tenure was marked by a change in emphasis from chromosomal analysis to molecular genetics. Genetic linkage was based on DNA markers and new techniques established for detection of mutations, with particular relevance for Duchenne muscular dystrophy, X-linked agammaglobulinaemia, Alport’s syndrome and Tay-Sachs disease.
With the rising public concern over many of the implications of the expanding knowledge of genetics and DNA biology and its applications, Bobrow became an influential member of national committees and an authoritative voice on the current problems.
Research in genetic diseases has moved from mapping Mendelian disease-related genes in two directions. The first is studying the functions of genes and their products, and the related cell and molecular biology. The second direction is complex genetic disease – common diseases such as heart disease and cancer are not due to a single gene defect, but are related to genes which increase the individual’s susceptibility. Identification of these genes has important implications for diagnosis, prognosis, prevention and therapy.
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Breast cancer is one of the commonest malignancies affecting women in the UK; one in nine will develop it. Some patients with breast cancer have a gene – BRCA1 –which predisposes to early-onset familial cancer of the breast and ovary. This gene gives a lifetime risk of 70 per cent for breast and 40 per cent for ovarian cancer.
BRCA1 has several functions; one is to form a complex with two other proteins to form an enzyme, ubiquitin ligase, which repairs DNA. Work at King’s College London has shown that most mutations in BRCA1 cause the enzyme to be inactive and so fail to repair DNA. However screening of many women with breast cancer has revealed many missense mutations whose effect is entirely unknown, raising serious difficulties in counselling the families. It was therefore important to study in detail these missense mutations; reproducing them in the protein complex has shown several different interactions between the three proteins and also loss of ubiquitin ligase activity in many of them. A functional assay for these mutations is being developed which will make the clinical counselling of the affected individuals more effective.
King’s is also studying acute promyelocytic leukaemia (APL), one of the commonest forms of acute myeloid leukaemia. APL is unique in that treatment with all-trans retinoic acid usually leads to complete remission in the short term (and long term with consolidation chemotherapy). The genetic fault is a translocation between chromosomes 15 and 17 to form a new fusion gene which produces a fusion oncogene (PML-RARa).
The mechanisms causing translocation are poorly understood, but a clue has come from APL secondary to treatment with mitoxantrone in patients with breast cancer – an example of treatment-related leukaemia, increasingly common following successful cancer therapy. Mitoxantrone is a cytotoxic drug which disrupts DNA by inhibiting topoisomerase, which regulates the coiling, transcription and replication of DNA. In these patients with secondary APL the translocation breakpoints were clustered on chromosome 15 at a site where topoisomerase inhibitors act, so implicating mitoxantrone in causing the secondary APL. This information is leading to studies to define the translocation(s) in primary APL.
Monitoring patients with cancer for relapse is a vital part of their management and the same group at King’s has developed a method (real time PCR) for APL which can detect one cell in 10,000, giving early warning of relapse and allowing treatment to recommence well before clinical relapse shows.
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Single gene diseases
Examples of single gene diseases to which King’s is making a contribution in understanding are Huntington’s disease and sickle cell disease.
Huntington’s disease presents in people between 30 and 50 years old with progressive dementia and abnormal and uncontrollable limb movements. The gene, an autosomal dominant and its product, huntingtin, have been identified. Work at King’s has developed a transgenic mouse model to understand pathogenesis and to evaluate treatments. Aggregates of huntingtin in the brain are related to cell damage and death and the mouse model is being used to examine methods of reducing these aggregates.
Sickle cell disease (SCD) is one of the inherited disorders of red blood cells involving abnormal haemoglobin production – the commonest genetic disorders worldwide. SCD is due to an autosomal recessive gene and occurs mainly in African and Afro-Caribbean people; it has a particular relevance for King’s and its partner hospitals in South East London owing to the large number of these ethnic minorities in the local population. The disorder is caused by the production of a defective form of haemoglobin, HbS. Red cells with HbS have a shortened life span; patients become anaemic and the red cells become sickle-shaped. This abnormal shape impairs red cells’ passage through small blood vessels, with sludging, thrombosis and reduced blood supply as consequences.
Normally, haemoglobin production switches from the fetal form, HbF, to the adult form, HbA, with a residual continuing production of HbF. In patients with SCD the greater the level of residual HbF, the less the severity of SCD, so ways to augment HbF production, which is beneficial, are being sought. The approach at King’s has been to search for genes regulating HbF production and two candidates have been identified on different chromosomes; once accurately defined, upregulating these genes offers a possible new treatment.
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Complex gene disease
One group of diseases being examined at King’s is inflammatory bowel disease – Crohn’s disease and ulcerative colitis. Two susceptibility genes have been identified. One on chromosome 16 increases the risk of Crohn’s disease by a factor of three in the heterozygote and a factor of 27 in the homozygote and is specific for ileal Crohn’s disease. The second gene also confers susceptibility to Crohn’s and its early onset but, interestingly, only if mutations are present in the first gene, suggesting the two interact in a common pathway.
King’s is taking part in an £8.6 million Wellcome Trust Case Control Consortium to identify susceptibility genes for common disorders, coordinating with Cambridge the study of inflammatory bowel disease. The others are Types 1 and 2 diabetes, hypertension, coronary heart disease, rheumatoid arthritis, tuberculosis and bipolardisorder.
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