Teamwork Boosts Insight
Marina Feric was stuck.
It’s not an uncommon feeling for a research scientists working on a particularly difficult problem, but Feric, who was studying the internal structure of frog cells, worried that she was not moving forward. As part of her project, she had injected magnetic, micron-sized beads into the frog cells’ nuclei and made observations about how the beads diffused; the idea was to learn about any structure by describing dispersion patterns – but the larger beads were not diffusing.
“I was making all these observations but I was stuck on how to prove whether actin was the material forming a structure inside the cell,” said Feric, a second-year graduate student in bioengineering. She brought up the difficulty in meetings with her advisor, Clifford Brangwynne, and Stephanie Weber, a post-doctoral researcher in Brangwynne’s lab.
“Cliff told me to take a step back and try the passive experiments instead of using magnetic forces on the beads,” Feric said. “They were like, ok, just try the simplest experiments first.”
Feric could see that the fluctuations of the beads indicated a structure inside the frog cells – large beads seemed to stick in place, while smaller ones passed through – indicating a structural scaffold. She built on this by removing actin, a type of protein, from the nucleus by treating the cells with pharmacological drugs.
“The large nuclear bodies sunk like pebbles to the bottom of the nucleus,” said Brangwynne, an assistant professor of chemical and biological engineering. “That was genuinely shocking.”
Before Feric’s experiment, scientists generally believed that cells were too small for gravity to have much effect on their structure, and, for most cells, that is true. But for very large cells, such as immature egg cells of the African clawed frog that Feric was studying, the experiment showed that a cellular structure was needed to serve as a brace against gravity.
In an article in Nature Cell Biology, Feric and Brangwynne concluded that when cells reach a certain size they become subject to gravity and require an internal scaffold to function. Gravity, which was previously discounted as a factor in cell development, now becomes an important factor in limiting cell sizes to, generally, below 10 microns.
“Gravity becomes really important at a smaller scale than you might have guessed,” Brangwynne said.
Feric, who is from Maryland, completed her undergraduate work in chemical engineering at the University of Maryland at College Park and started at Princeton just before Brangwynne set up his new lab space in Hoyt Laboratory. Hoyt, which once housed the university’s chemistry labs, was completely renovated in 2012 and 2013. The work created state of the art equipment and research space for the School of Engineering and Applied Science.
“It was a very cool to see the entire process and what it takes to start a lab,” Feric said. “Plus, I got to see how everything works.”
Bioengineering combines biology’s focus on living systems with engineering’s concentration on the mechanics of how the system functions, and Feric was particularly interested in how structures scale in biology.
“When civil engineers build bridges, the size of the structure matters for a number of reasons,” she said. “We are interested in a similar fashion in cellular structures.”
Feric felt that the frog cells’ unusual size – with a volume five orders of magnitude greater than an average cell – made them an excellent candidate for study. Brangwynne agreed and suggested that she spend some time working at Cold Spring Harbor Labs in New York to get more expertise working with frogs.
“Cliff sponsored it; he thought it was a good idea to meet with other people in the field,” Feric said. “I started in the spring and, by the summer, we were ready to work with frogs.”
Feric’s goal when she started the experiment was to gain insight into the cell’s internal scaffolding. Researchers had previously noted the presence of actin in cells, and she thought it could be the key to describing the structure inside the large nuclei.
“It was always in the back of my mind that actin could be important,” she said. Actin tends to form filaments that could be the basis of a scaffold-like structure. “But no one had been able to see these structures in in-vivo conditions inside the nucleus.”
Feric’s approach was to use small beads of various sizes coated with an anti-sticking surface and try to disperse them within the cell. By monitoring the motion of the beads, she could map out obstacles – where the beads stuck – and open areas.
“Large beads stuck and small beads were able to move; medium sized beads had this jumping behavior which suggested they were moving around filaments and hopping between pores,” she said.
When the researchers removed the actin from the cells, they were able to confirm that it formed the bead-capturing structure. And they also learned the important role that gravity can play above a certain cell size.
“Uncovering the role of nuclear actin felt as if we were solving a mystery,” Feric said. “Finding out that gravity was the dominant force in the end helped us close the case.”