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The Flow of Memory
The memory of the immune system is truly a wonderful thing—one of evolution's great innovations. It allows us to conquer many of the common and rare infections with which we are "challenged" over our lifetimes. In days past, when people were exposed for the first time to a virus like variola (which causes smallpox), they would endure the infection until their bodies had defeated the virus or they died. If they recovered, they would be immune for the rest of their lives. Even if "challenged" with another exposure to smallpox, their immune systems would mount a much more vigorous defense against the pathogen, fight it off, and clear it from the system subclinically. Knowing this, doctors began centuries ago to practice "variolation" and infect patients with what they hoped was a tiny dose of smallpox to immunize them against later infection. This led eventually to the innovation of vaccination, a similar but much safer practice that was developed in the late 18th century. Vaccination also took advantage of immunological memory, though perhaps a slightly distorted memory. If a person is exposed to a variola, like cowpox or vaccinia virus, he/she will acquire immunity against the similar smallpox virus. Today, in the wake of a successful concerted global effort to eradicate smallpox, it is hard to imagine the excitement generated by the knowledge that a single shot and a sore arm could provide a lifetime of protection against such a devastating scourge. Harder, still, because unlike 200 years ago, we can now describe the basic biology of how vaccination works, based on our understanding of immunological memory. Immunological memory is still not perfectly understood, though, and there many diseases for which we have no good vaccines despite sometimes enormous efforts to develop them. One of the newest members of the Department of Immunology at The Scripps Research Institute (TSRI), Associate Professor Michael McHeyzer-Williams would like to change that. He and his team in their brand-new laboratories on the east side of campus would like to deepen our understanding of the immune system's memory mechanisms. McHeyzer-Williams has worked towards this goal for as long as he can remember in his professional life. He did his graduate work with Gus Nossal at The Walter and Eliza Hall Institute in Melbourne, Australia, isolating and identifying these memory B cells. Memory B cells are one of the key mediators of immunological memory and produce a vigorous antibody response to fight off viral infections to which a person has previously been exposed. Towards the end of his graduate career, he realized that there was much more to the picture. "I realized to understand B cell memory," he says, "one had to understand helper T cell regulation of B cell memory." So he spent several years as a postdoctoral fellow at Stanford University with Mark Davis, identifying and isolating specific T cells and trying to understand how T cells regulate B cell responses—work that he continued in North Carolina, as a faculty member at the Duke University School of Medicine. "Then I realized that this T cell response that regulates B cell memory is, itself, regulated by dendritic cells," he says. So he and his laboratory started looking at the action of these dendritic cells, isolating the ones that seem to be involved in the regulation of helper T cells and asking how they do it. McHeyzer-Williams and his laboratory are continuing this work at TSRI, and they have come to appreciate that since B cell memory depends on helper T cell regulation, which depends on regulation of dendritic cells, they must study all three. "Memory is not one thing," he says. "It's a cascade of events with a multitude of outcomes." Dendritic Cells—the Early RegulatorsThe easiest way to understand immunological memory is to take a look at the process through which it forms—starting at the moment when a virus or other pathogen first enters the body and begins to circulate, infect, and replicate. Over millions of years of evolution, the immune system has built up myriad ways of countering such potentially lethal infections, including a strong first line of defense mediated by innate immune cells. One of these innate immune cells, the spider-like dendritic cells, circulate through the bloodstream or sit in the skin or other organs when they are alerted to the presence of a pathogen. Once alerted, they become active. As active dendritic cells, they mediate the general innate immune response and the clearance of the pathogens from the bloodstream and infected tissues. This involves making and releasing inflammatory chemicals and proteins that attract neutrophils and other "effector" innate immune cells. The innate immune response is, however, only half the picture. The slower and more specific adaptive immune response plays a critical role in ensuring our survival as well. Activated dendritic cells play a key role in this response. "They are the key regulators of everything that follows," says McHeyzer-Williams. Activated dendritic cells are the intermediary between the innate and adaptive arms of the immune system, because they act as "professional" antigen-presenting cells. They take up bacteria or viruses and process them—chopping their proteins and other "antigen" components into pieces and presenting them on their surface in a large assembly known as the major histocompatability complex. They also move into the local lymph nodes and show the antigen to naïve helper T cells, which periodically enter the lymph nodes as they circulate through the bloodstream. Helper T cells are one of the crucial players in the adaptive immune response. They develop in the thymus and circulate as naïve cells until they "see" the right professional antigen-presenting cell with their unique T cell receptor, which is something like a lock with only one key. When a circulating naïve helper T cell with the right T cell receptor sees an activated dendritic cell with the right antigen in its major histocompatability complex, then that naïve helper T cell itself becomes activated. Once activated, the helper T cell will produce a swill of chemicals to clear the infection and attract other immune cells, like killer T cells and B cells. "That starts the whole adaptive response," says McHeyzer-Williams. The T helper cells then turn around and regulate the response. Synapse I and Synapse IIBorrowing a term from the neuroscientists, immunologists call the interface between the naïve helper T cells and the activated dendritic cells synapse I, possibly to evoke the complex cellular communication processes that are focussed at this interface through the T cell receptor and the antigen-major histocompatibility complex. At this interface, the cells communicate with one another and transmit information through surface and secreted molecules. The nature of these interactions regulate the outcome of the immune response. In the case of naïve helper T cells, the outcome is their activation. Once the T cells are activated, all hell breaks loose. The now activated helper T cells seek out B cells that have taken up the antigen with the right receptor. When these antigen-activated helper T cells find these B cells, they communicate with them through what is known as synapse II—the interface between the activated helper T cells and the antigen-activated B cells. At this interface, another set of signals is exchanged in what is one of the crucial regulatory events in determining immunological memory. The outcome of this encounter determines which of two drastically different destinies a B cell will follow—to produce antibodies or to produce memory. Some of the B cells stay local and make buckets of antibodies. "This response is a clearance response—you are trying to get rid of the antigen load," says McHeyzer-Williams. Other B cells commit to the memory pathway and begin to develop memory B cells. The way that the development of memory works is that the B cells committed to the memory pathway move into small localized areas in lymph nodes (known as follicular areas), which develop into germinal centers that witness the prolific expansion of these B cells. During this expansion, the B cells also introduce point mutations into their own antigen receptors. These point mutations sometimes change the affinity of the receptor and occasionally will result in a B cell with a mutated receptor that has a much higher affinity than before. "What you have done on this micro-scale," says McHeyzer-Williams, "is to accelerate evolution." This process creates memory cell "compartments" that contain cells that are much more effective at fighting off the pathogens for which they are specific. Inside these germinal centers, the activated helper T cells help sort through these B cells according to their affinity and the highest affinity ones become long-lived memory B cells. Then the helper T cells also commit to becoming long-lived memory cells. Since the B cells have a much higher affinity for the antigen, they are able to detect smaller amounts of it. And since there are more memory helper T cells around that have the right T cell receptor, the B cells have an easier time presenting the antigen to the correct helper T cells. This means that the memory T cells can be activated rapidly and the overall response can be mounted more quickly. These accelerated kinetics and increased magnitude of the response offers better protection than would be possible without memory cells. If another challenge does come from a pathogen with the same antigen, the memory B cells and the helper T cells can create a rapid and severe response. "Now you get a reaction within two to three days that would have taken five to seven previously," says McHeyzer-Williams. Not All Naïve T Cells are Created EqualMcHeyzer-Williams and his group are particularly interested in the receptors, co-receptors, signaling molecules, and other molecules involved in various stages in the development of immunological memory. If the right signals are absent from Synapse I and Synapse II, there will not be memory cells later. Using the technique of flow cytometry, it is possible to separate and study sub-populations of cells in the body, or to find that single cell with the defined characteristics of interest. In fact, McHeyzer-Williams has built his career on the ability to find those very rare cells with high fidelity. Recently, what McHeyzer-Williams and his group found is that not all naïve helper T cells are the same. There is a major division among helper T cells even before they are activated on the basis of a mysterious glycoprotein-anchored protein called Ly6C that either is or is not expressed on their surface. A few years ago, they discovered this distinction almost by accident in naïve helper T cells that have been exported from the thymus but are not yet activated. There are hundreds of genes across these cells that are different, which is a consequence of the selection process that takes place in the thymus. But none of them were predicted to essentially distinguish between two types of naïve T cells in circulation. This was something McHeyzer-Williams and his laboratory discovered one day in a sweeping experiment in which they subjected naïve helper T cells to every reagent in their freezer. "There it was," he says. "Fifty percent of the helper T cells in the periphery had Ly6C and fifty percent didn't." The ligand that binds to Ly6C and the overall function of this protein are not known, though McHeyzer-Williams says it probably modulates receptor responses. However, its presence or absence on the surface of these cells seems to have a major effect on function of the cells. It is a major indicator of how they develop and regulate other cells downstream. "It looks like the two different types of helper T cells help B cells in different ways," says McHeyzer-Williams. In experiments in which he and his colleagues transferred only the Ly6C positive helper T cells into a model system and then activated them by challenging with the correct antigen, they saw "buckets" of antibody being produced, McHeyzer-Williams says. But when they did the same experiment with Ly6C negative helper T cells, they saw antibody production that was only five to ten percent of the norm. "Ly6C-positive T helper cells appear to be specialized for helping B cells," concludes McHeyzer-Williams. A New Neighbor and CollaboratorIn the slightly more than a year since he has come here, McHeyzer-Williams and his laboratory have managed to get things up and running. He arrived near the end of 2001 with his core group of two postdocs and one technician, and they all had to wait several months for their new, dedicated flow cytometer to arrive. Once it did, it took a few more months for them to get it running at peak performance. Shortly after his new instrument was finally on-line, TSRI immunology Professor Hugh Rosen [featured in a recent issue of News&Views] moved into the same contiguous laboratory space and was writing grants and getting his own laboratory started. After several conversations, McHeyzer-Williams recalls, "I said, 'let's do an experiment together.'" So they did. They designed an experiment involving T cell selection in the thymus using a chemical that Rosen had and an experimental approach designed by McHeyzer-Williams. When they analyzed the results, they saw something completely new—a result that should be forthcoming soon, as their first paper is currently under review. "We're very excited about it," says McHeyzer-Williams. "Both of us have started a whole new directions of research that we wouldn't have had." This is especially exciting, he says, since his previously existing research projects are now back on track after the move. McHeyzer-Williams and his laboratory are actively collecting data, writing papers and grants, and he has four new people starting soon. Last week, in fact, he was preparing to interview one postdoctoral fellow candidate and awaiting the arrival of another. And he says that after six months spent doing experiments, his group is finally at a place where they are really starting to take off. "Now we have too much to do," he says. "This next 12 months are going to be nuts."
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