Assistant professor Mingxu You, chemistry, recently received a five-year, $1.9 million NIH Maximizing Investigators’ Research Award (MIRA) to fund his research in developing new tools – DNA-based probes – to quantify intercellular mechanical forces and understand a variety of mechano-sensitive cell signaling events at the molecular level.
As he explains, intercellular forces are critical regulators in many physiological and pathological processes, but scientists have until recently lacked the tools and approaches to characterize these mechanical events. “It is a whole new way to understand growth, division, intercellular motion and interaction,” You says.
Cells are usually touching each other or a substrate, pushing and pulling each other to work together as a tissue, an organ and at the whole body level, he adds. But these forces are so tiny and ever-changing, it is very hard to see how cells are physically communicating with each other, for example, during development, cell differentiation, normal physiological and various disease processes.
The You Lab, which includes postdoctoral researcher Bin Zhao and chemistry Ph.D. students Yousef Baheri and Puspam Keshri, will team with biologists Barbara Osborne and Tom Maresca, Lisa Minter of veterinary and animal sciences and Yubing Sun, mechanical and industrial engineering, to further develop these DNA-based tools to visualize, monitor and quantify such cellular forces.
You says, “In the near future, people will be able to apply these tools broadly to depict the basic principles of tissue morphogenesis, growth, and homeostasis. They will serve as a critical foundation for developing novel strategies in tissue engineering, regenerative medicine, immunotherapy and cancer treatment.”
Specifically, You says, “We are interested in the Notch signaling pathway. It’s widely conserved in most cells and organisms, very common to find, and it’s interesting because it’s really simple. There are only five Notch ligands and four Notch receptors but they regulate quite a diverse range of downstream functions,” he adds.
Characteristics of Notch receptor-ligand expression vary in different physical environments, You says. “Even though they are similar they can have very different effects, including opposite ones like tumor promoting or tumor reducing. Cells need force to activate the Notch pathway and we want to know how the different stress levels – how strong the mechanical forces need to be – to contribute to tumor growth or reduction. Using this new probe we can tell which protein ligand-receptor pair contributes to a particular intercellular force.”
You says he learned about the force measurement challenge when he came to campus in 2016 and asked his friend, mechanical engineer Sun, to name an area in the emerging field of mechano-biology that needed attention. “At the time, I had a system already developed that I used to modify DNA-lipid probe onto the cell membrane, and we realized we might be able to design a DNA structure to probe and detect intercellular forces,” he notes.
DNA was a good candidate for the probe, he adds, because investigators can control its folding and hybridization, the sequence of nucleotides, very precisely. “Also, we can control not only the structure but the dynamics, which in this case refers to zipping together and unzipping of the DNA. Once you have the probe, you can detect a distance-sensitive reaction between fluorophore, a dye, and a ‘quencher’ that can suppress the fluorescence signal.”
“You can see this dye-labeled DNA directly under a fluorescence microscope,” he explains. “For some experiments we want to know just whether there is a force or not, but for others, we want to actually quantify the strength. We are now developing probes for both purposes.”