Dr. Michael Reth’s career in some ways mirrors the history of science of the last decades. Reth, professor at the Max Planck Institute of Immunobiology and the University of Freiburg, and some of his colleagues have recently uncovered the mechanism that foreign substances use to activate B cells of the immune system. The researchers were using synthetic biology methods long before this particular branch of science existed in its present form. Their results require a paradigm change and a revision of the reference books. The spokesman of BIOSS (Centre for Biological Signalling Studies) excellence cluster at the University of Freiburg, which is specifically focused on synthetic biology, has stuck to his original ideas. The following paragraphs give an insight into the story of a major discovery and the decisions made by a highly committed researcher.
B cells play a crucial role in the immune response to pathogens and other foreign substances. When they come into contact with molecular structures with a potentially damaging effect (so-called antigens), the B cells make antibodies against antigens, which destroy the intruders. B cells only produce antibodies once they are activated. They are activated by way of receptor proteins known as B-cell receptors, proteins located on the surface of B cells that bind foreign structures. But how are B-cell receptors activated? And how are the signals transmitted into cells? Up until now, immunologists have worked on the basis of the "cross-linking model" postulated around fifteen years ago. This model assumes the presence of resting B-cell receptors on the cell surface. Only when an antigen cross-links two dispersed B-cell receptor monomers will the receptors form dimeric structures that trigger a signalling cascade inside the cells that then releases antibodies. "Our findings suggest that the cross-linking model is wrong," said Prof. Dr. Michael Reth from the Max Planck Institute of Immunobiology and Epigenetics and spokesman of the BIOSS (Centre for Biological Signalling Studies) excellence cluster at the University of Freiburg. "In fact, quite the opposite is true."
Reth travelled a long way before he came to this conclusion. "Like many researchers of my generation, I was mainly interested in biology in terms of ecology," said the immunobiologist who was born in Düsseldorf in 1950. "However, as a biology student in Cologne I discovered that the ecology of that time did not really focus on big global problems such as nutrition, the finiteness of resources or environmental catastrophes. Ecology dealt mainly with the behaviour of beetles. And this is why I found research areas such as genetics far more exciting than ecology." Reth did his diploma thesis in 1977 at the Institute of Genetics in Cologne in the department of Prof. Klaus Rajewsky who was among the first to use molecular biology methods to investigate immunobiological issues. Reth's interest in cellular and molecular immunology was further strengthened by a related family illness and a publication by two scientists who were later to become Nobel Laureates, Georges J. F. Köhler and César Milstein, on the production of monoclonal antibodies. "Our laboratory in Cologne was one of the first in the world to use Köhler and Milstein's method to produce monoclonal antibodies," said Reth who was motivated by questions relating to the diversity of the immune response on the antibody level and to the way molecules recognise their target structures on antigens.
Shortly after Köhler and Milstein's paper, the geneticist Susumu Tonegawa published his discovery that genetic material can rearrange itself to form the vast array of antibodies needed to protect humans against any type of antigen. Tonegawa was awarded the Nobel Prize in Physiology or Medicine in 1987 for this discovery. "I knew immediately that I had to adapt the new molecular biology methods to my research on the development of antibody diversity," recalls Reth. He finished his doctoral thesis in 1982 and moved on to the Columbia University in New York, joining the group of the molecular biologist Frederic Alt that was working on the variability and regulation of antibody genes. Reth and his colleagues discovered that antibody genes were regulated by some of their products, namely membrane-bound antibodies. Upon his return to Cologne in 1985, Reth found out that the transport of membrane-bound antibodies to the cell surface depended on two other membrane proteins that are required for the transmission of signals into cells. These proteins form the B-cell receptor. "We were the first to describe the signalling unit of the B-cell receptor in 1988," said Reth highlighting that this finding was an impulse for further research into B-cell receptor-related signalling. Reth was once again driven on by another question, this time relating to the elements that play a crucial role in the complex molecular signalling networks. Reth and his team have subsequently been able to discover many of the elements that they had been looking for.
In 1989, the Nobel Laureate Georges Köhler brought Reth to the MPI in Freiburg. In 1995, Reth began establishing a study priority on molecular immunology at the Faculty of Biology at the University of Freiburg. Reth adapted the biochemical method of native gel electrophoresis used by Prof. Dr. Nikolaus Pfanner’s research group that was studying the import of proteins into mitochondria. This method made it possible to investigate cell extract proteins in their native three-dimensional form. The surprising finding of the experiments Reth carried out with his former doctoral student Wolfgang Schamel was that the exposure of a cell to antigens resulted in the dissociation of the oligomers and to the transduction of signals. The researchers’ finding was therefore in complete opposition to the currently accepted model of the time that suggested that the antigen receptors on B cells are dispersed monomers that are activated when two receptors are cross-linked by an antigen. The new model of B-cell receptor activation suggests that it is the dissociation rather than the formation of a B-cell receptor complex that activates B cells. This led to uproar in the immunobiological community. The paper came close to not being published because of concerns about the methodological procedure. “Our results were totally and utterly rejected,” said Reth. However, Reth and his team were convinced that their results were correct and started looking for ways to confirm their discovery. In 1998, the researchers made a revolutionary decision. “Back then, research focused mainly on destruction,” recalls Reth. “Researchers used to investigate biological structures by disintegrating them or by switching off genes.” Reth and his colleagues chose to do exactly the opposite. They rebuilt the B-cell receptor of murine B cells in a Drosophila cell in order to study its structure and function in greater detail. Nowadays, this method is known as synthetic biology, but at the beginning of 2000, the immunobiological community was far from convinced that this would work. In 2010, Reth and his postdoc Dr. Jianying Yang from China eventually achieved a breakthrough. In the meantime, Reth had become spokesman of the BIOSS excellence cluster consisting of seven University of Freiburg faculties, the MPI and the Fraunhofer Institute for Physical Measurement Techniques. BIOSS’ objective is to focus on research “from analysis to synthesis”. Synthetic biology eventually led to the confirmation of the groundbreaking theory postulated by Reth and his team of researchers.
The researchers integrated individual B-cell receptors that were coupled to one of the two halves of the green fluorescent protein (GFP) into a Drosophila cell. The receptors fluoresced green when the two halves combined, i.e. when the B-cell receptors formed dimeric structures. The researchers hypothesised that the B-cell receptors are not present as dispersed monomers on resting cells such as postulated by the cross-linking model, but rather as oligomers that dissociate upon exposure to an antigen. The researchers were able to prove their hypothesis with experiments in which the surface of the Drosophila cells fluoresced green even though the B-cell receptors had not been activated. Since this is only possible when the two halves of the GFP combine with each other, the finding could only mean that the receptors also had to be closely connected in cells that were not activated by antigens. "The B-cell receptor occurs as an organised oligomer in a resting B cell," said Reth, summarising their discovery. "Other proteins are also involved in the regulation of this complex. These proteins ensure that the oligomers remain stable and do not release any activation signals. When an antigen destroys this order, a signalling cascade is induced that eventually causes a cell to release antibodies."
Reth and his colleagues have recently presented their model, which they refer to as dissociation activation mode, in the renowned journal Nature. The researchers' discovery has enormous implications for the field of medicine. "Many human diseases are associated with the dysregulation of B-cell receptors," said Reth. This includes diseases related to autoimmunity or the formation of B-cell tumours in leukaemia or lymphomas, for example. Once the researchers are able to discover the proteins that prevent the activation of the B-cell receptor oligomers on resting B cells, this might lead to completely new therapeutic strategies. "The world now knows about our discovery," said Reth going on to add "it remains to be seen what other immunological researchers think about the findings and whether immunological reference books will be revised."
Further information:Prof. Dr. Michael Reth Department of Molecular Immunology Max Planck Institute of ImmunobiologyFaculty of Biology - University of FreiburgTel.: 0049 (0)761/5108-421E-mail: michael.reth(at)bioss.uni-freiburg.de