In a lab at the University of North Carolina at Chapel Hill, a tiny blue light flashes, and a protein switches on in a cell.
Light directs the protein to the mitochondria of the targeted cell. Alternatively, by a bit of genetic tweaking, the protein can be directed to the cell membrane or the cytoskeleton of the cell. This selective targeting and activation allows scientists to see exactly what the protein is doing in a single place and time.
This technology is possible due to the efforts of a team at the UNC Eshelman School of Pharmacy led by Fred Eshelman Distinguished Professor David Lawrence, Ph.D. Lawrence’s team has created a designer version of a ubiquitous protein kinase that can be activated with light. Their accomplishment is published in Cell Chemical Biology.
“Try to imagine the complex behavior of a single cell,” Lawrence said. “There is incredibly intricate chemistry taking place, sophisticated interactions in constant motion at the molecular level. How do you isolate and study one facet of such a complex system?
Anywhere, not Everywhere
“What if you could go into the cell and say, ‘I activate you protein XYZ and only you’?” Lawrence said “And now you can see how that cell behaves by activing that single protein. We have accomplished this by creating an engineered protein that can be activated with light.”
Lawrence’s team has designed an analog of the cAMP—dependent protein kinase, a mouthful-of-a-name that is more commonly referred to as PKA. PKA is an enzyme that performs a myriad of functions in a cell, including regulating metabolism, cell migration (important in cancer metastasis), and whether a cell lives or dies. PKA is activated by cyclic adenosine monophosphate, a messenger molecule known as cAMP. PKA is the best known and most studied member of a family of more than 500 enzymes.
Before Lawrence’s breakthrough, the only way to activate or deactivate PKA in a cell was to do so chemically, which activated or deactivated the enzyme everywhere in the cell. It was impossible to target a specific site.
OptoPKA in the Spotlight
Lawrence calls this PKA analog optoPKA and is working in an area of science known as optogenetics, which is the field of creating of light-responsive proteins. It hasn’t been easy.
“With flat-screen TVs, the challenge has been to make black truly black,” Lawrence said. “The challenge with light-activated proteins has been to ensure that, in the absence of light, optogenetic proteins are truly off. That is where we have been successful.”
Lawrence selected PKA to work with because it plays a variety of roles in cells and that the cellular behaviors (metabolism, migration, cell life/death) triggered by its action are initiated at different sites within the cell. In this regard, the team has now been able to examine the biochemical consequences of activating this enzyme at different sites within the cell. “No one has been able to do this before,” Lawrence said.
This technology has important ramifications. “For example, a defective cell should self-destruct,” Lawrence said. “Uncontrolled growth of defective cells is the basis of cancer. Correlating a cellular behavior with a biochemical cause is important because of the potential therapeutic implications.”
Authors, Funding and Citation
- Colin P. O’Banion,Ph.D., a postdoctoral fellow in the UNC Eshelman School of Pharmacy
- Melanie A. Priestman, Ph.D., a postdoctoral fellow in the UNC Eshelman School of Pharmacy and now vice president of research and development at Iris Biomed
- Robert M. Hughes, Ph.D., an assistant professor at East Carolina University
- Laura E. Herring, Ph.D., a research assistant professor in the UNC School of Medicine and director of the UNC Michael Hooker Proteomics Center
- Stephen J. Capuzzi, a graduate student in the UNC Eshelman School of Pharmacy
- David S. Lawrence, Ph.D., chair of the Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of Pharmacy