Department of Immunobiology, University of Arizona College of Medicine
Dreaming Big When It Comes to the Possibilities
Metals play a very important role in biology, as metal ions are associated with an estimated 40% of all proteins. In bacteria, dedicated acquisition and efflux systems tightly control metal homeostasis. Some metals, such as iron, are so critical to bacterial physiology that in most cases only the acquisition system exists; while others, such as copper, are so toxic that only efflux systems exist. It is this copper toxicity that is of particular interest to Dr. Michael Johnson and his new laboratory in the Department of Immunobiology at the University of Arizona’s College of Medicine. Dr. Johnson notes that although bacteria can develop resistance, as seen against modern antibiotic therapies, they remain vulnerable to the toxic effects of copper. His research aims to unravel this age-old mystery by understanding how copper is toxic to bacteria, and how bacteria overcome copper toxicity.
Using S. pneumoniae as the initial model organism in a microarray analysis, Dr. Johnson validates bacterial pathways targeted in response to copper toxicity and alleviating copper stress. Armed with this information, he can study bacterial behavior in relation to cellular and animal models. Several wars are waged when bacteria enter a host, one of which is called nutritional immunity and occurs in the phagolysosome. Here, the bacteria try to uptake or acquire metals from the host, such as iron, calcium and manganese, while the host cell throws out defense mechanisms to sequester those necessary metals. At the same time, the host cell attacks the bacteria with toxic metals such as copper, to actively stop or kill the invader. The question is, how does a mutation to one of the bacteria’s targeted pathways impact this war? Could a mutation increase or decrease the bacteria’s susceptibility to copper, and could it attenuate the disease model? What are the many consequences of mismetallation, where unexpected metal ions, such as copper, substitute for intended metal ions in a reaction?
These studies were previously performed manually using 10 mL culture volumes. Dr. Johnson and his researchers worked nights and weekends during time-consuming kinetic studies. Recently, he added BioTek’s Cytation™ 5 Cell Imaging Multi-Mode Reader to his new lab. Now, his lab is able to increase sample throughput and decrease reagent volumes by moving into a 96-well microplate format, and program the Cytation 5 for overnight studies without requiring the presence of any personnel. Dr. Johnson is taking advantage of the multi-mode functionality; using absorbance, fluorescence intensity, fluorescence polarization and imaging when running existing assays and developing new methods down the road. Having these capabilities together in one instrument helps his lab to examine multiple aspects simultaneously. He can now study bacterial replication inside of a cell, along with gene regulation, in real-time instead of performing separate assays, as one example of how powerfully the instrument will impact his studies. Dr. Johnson also took advantage of the optional dual reagent injectors, and optional gas controller for incubating bacteria and cells directly in Cytation 5. “I’m using all aspects of my background to understand everything about these pathways, and Cytation 5 allows me to test it all in high throughput,” he notes. “I really appreciate this versatility because it allows me to dream big in terms of what’s possible.” Cytation 5’s integrated Gen5™ Microplate Reader and Imager Software is user-friendly and quick to program, yet also offers advanced features for in-depth data/image capture and analysis.
Dr. Johnson considers Cytation 5 as a flagship instrument in his laboratory, but states that BioTek’s service and support are just as important to his lab. He values knowing that he can depend on this team to actively help him expand the boundaries of what’s possible with Cytation 5, or any of BioTek’s life science instrumentation. “They are so helpful and proactive, with a natural scientific curiosity,” he asserts. “They also have confidence in their products and solutions, and that gives me more confidence in my own abilities and my ‘dream big’ aspirations.”
Dr. Johnson (left) with Miranda Neubert (right) in the lab with BioTek’s Cytation 5.
To learn more about University of Arizona College of Medicine, visit their web site.
Thanks to Dr. Johnson at University of Arizona College of Medicine for sharing his BioTek experience.