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Nanotech News
Nanoscale Engineering Leads to New Way of Studying Cell-Cell Signaling An experiment that began as a “fantasy pipe dream” just three years ago is now a reality. Researchers with the Lawrence Berkeley National Laboratory and New York University School of Medicine, combining nanotechnology with biochemistry, have created unique synthetic membranes that, for the first time, enable them to directly control signaling activity in living T cells from the immune system. Already their experiments, the results of which were published in the journal Science, have yielded surprising results.
“This marriage of inorganic nanotechnology with organic molecules and cells enables us to go inside a living cell and physically move around its signaling molecules with molecular precision,” said Jay Groves, Ph.D. “Our experimental beaker has now become the inside of living cells and we can watch chemical reactions take place there. T cells are a type of white blood cell that attacks tumor cells and virus-infected cells. These immune system cells also produce a wide variety of chemical signals that regulate the body’s immune response. “Scientists, including ourselves, have been posing elaborate theories about how the strength and duration of signals that activate T cells are controlled, without having been able to do direct experimentation of key factors,” said Dr. Groves. “Three years ago, we had this fantasy pipe dream about an experiment to measure how alterations in the geometric shapes of the [contacts between T cells and tumor cells, for example] – what we call spatial mutations – would affect T cell signaling. Then we realized we have the tools to create nanoscale patterns, that can do this.” The human immune system is a remarkable collaboration of different types of cells, working together to protect our bodies from bacterial, parasitic, fungal or viral infections, and against the growth of tumors. The process starts when “antigens,” special markers on the surface of a cell, identify another cell as “non-self,” and signal the cellular warriors of the immune system to kill the invader. Leading this attack will be the T cells, cells produced by the thymus. A substantial body of research has established that the control center for T cell signaling is at the junction or point of contact between T cells and antigens, dubbed the “immunological synapse” because it resembles the synapse between two communicating nerve cells. At the immunological synapse, a central cluster of T cell receptors surrounded by a ring of adhesion molecules form what co-investigator Michael Dustin, Ph.D., of NYU, has described as a sort of “bull’s-eye.” The center of this bull’s eye has been dubbed the “central supramolecular activation cluster,” or c-SMAC, because it was believed to be the source of T cell activation. “The original idea behind the c-SMAC was that the larger the T cell receptor cluster, the stronger the T cell activation signal,” said Dr. Groves. “This simple vision of strength in numbers had begun to show cracks, and now we have demonstrated that just the opposite is true. The coalescence of the c-SMAC cluster extinguishes the T cell activation signal. The duration of the activation signal is related to the spatial organization of the T cell receptors rather than cluster size.” Dr. Groves and his colleagues constructed their synthetic membranes out of lipids, which they assembled onto a substrate of solid silica so that the membranes were able to float freely a few nanometers above the substrate. This enabled the researchers to preserve the membranes in their naturally fluid state, allowing lipids and T cell receptor proteins to diffuse and interact freely over macroscopic distances. “The fluidity of our membranes created artificial antigen-presenting cell surfaces that enabled the formation of functional immunological synapses with living T cells,” said Dr. Groves. The investigators were able to spatially mutate the geometric shapes of the immunological synapses by embedding the silica substrate with chrome lines that were only 100 nanometers wide. These ultra-narrow chrome lines served as barriers that restricted the motion of membrane lipids and T cell receptor proteins. Using electron-beam lithography, a technique developed for computer chip manufacturing, the researchers were able to configure the chrome lines into several distinct patterns, including simple parallel lines, grids, and a series of concentric hexagons. “By changing the shape of the immunological synapse, we showed that the synapse signal starts out in an amplified mode, and that the transport of the T cell receptors towards the center weakens and eventually extinguishes the signal, irrespective of the degree of clustering,” Dr. Groves said. Dr. Groves said this new technique for spatial mutation studies should be applicable to many intercellular signaling systems. Already, he and his colleagues have begun applying it to study neuronal synapse formation, and cell signaling mechanisms in the development of cancer. They are also using it to look at the dynamic range of signaling over which T cell receptors can respond. “Essentially, these experiments amount to using inorganic nanotechnology to physically grab a protein in a living cell and move it to another position in that cell – then watch how the cell responds,” said Dr. Groves. “We used it to study the T cell as a paradigm system, but the theme here is much more general. Whereas the spatial position of molecules is rarely thought to play an important role in the outcome of a chemical reaction, with our experimental technique we are seeing that, in living cells, this is not the case. The spatial position encodes information which can be directly translated into altered chemical outcomes.” This work is detailed in a paper titled, “Altered TCR signaling from geometrically repatterned immunological synapses.” An abstract is available through PubMed.
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