Directed differentiation: controlling stem cell fate with optogenetics
Researchers have created a method to regulate embryonic stem cell differentiation with light, transforming them into neurons in response to an external cue.
The technique, developed at UC San Francisco, also uncovered an internal timer mechanism within stem cells that helps them decide whether to pay attention to a developmental signal or ignore it as inadvertent biological noise.
Previous studies have identified a range of molecular cues that tell stem cells when to divide into mature cells at the appropriate moment. The timing and direction of stem cell differentiation are crucial for normal embryonic development.
Scientists hope that by directing stem cells to differentiate they could repair damaged or aging tissues using the body’s innate regeneration mechanism. However, getting stem cells to do what we want has proven harder than expected.
Recent experiments have shown that the developmental cues are regularly being flipped on and off in undifferentiated stem cells, but no-one knew how the stem cells decided whether to respond or not. Cameron Sokolik, the lead author of the paper published in Cell Systems, said: “These cells receive so many varied inputs. The question is how does the cell decide when to differentiate?”
Using optogenetic tools, the researchers cultured mouse embryonic stem cells with a light sensitive switch that turned on the Brn2 gene, a strong cue for neural differentiation. By altering the strength and duration of the light pulses, they could control the dosage of Brn2 and observe how the cells respond.
They discovered that the Brn2 signal had to be both strong enough and long enough for the stem cells to start transforming. Otherwise, they completely ignored it.
To learn how the stem cells could distinguish between fast or enduring signals the team looked at the transcription factor Nanog, which normally halts stem cell differentiation. They added a fluorescent label to Nanog so that levels of the protein could be used to examine the cells’ decision making.
The group found that activation of Brn2 disrupts the feedback loop that stops the cell from differentiating causing levels of Nanog itself to fall. However, the protein does not completely disappear for around four hours. Therefore, Nanog acts like cellular stop-watch. If the Nanog levels drop entirely while the Brn2 signal is still on, the stem cells will begin to develop into neurons. If the Brn2 signal is unintentional, then the Nanog levels will return to their normal levels quickly when the signal is turned off.
Dr Matthew Thomson, co-senior author of the study, said: “It’s hard for a cell to be both tolerant and fast, to reject minor fluctuations, but respond very precisely and sharply when it sees a signal. This mechanism is able to do that.”
Dr Thomson hopes that this research will eventually lead to light-inducible differentiation technology that can allow a bath of stem cells to create complex three-dimensional tissues. Turning on a pattern of different coloured lights could potentially form an organ ready for transplantion into a patient.
“There’s lots of promise that we can do these miraculous things like tissue repair or even growing new organs, but in practice, manipulating stem cells has been notoriously noisy, inefficient, and difficult to control,” Thomson said. “I think it’s because the cell is not a puppet. It’s an agent that is constantly interpreting information, like a brain. If we want to precisely manipulate cell fate, we have to understand the information-processing mechanisms in the cell that control how it responds to the things we’re trying to do to it.”
Sokolik S, Liu Y, Bauer D, McPherson J, Broeker M, Heimberg G, Qi LS, Sivak DA, Thomson M Transcription factor competition allows embryonic stem cells to distinguish authentic signals from noise Cell Systems (2015) doi: 10.1016/j.cels.2015.08.001
Image caption: Human embryonic stem cells (A) Stem cell colonies that are not yet differentiated (B) Nerve cells, an example of a cell type after differentiation
Image credit: Nissim Benvenisty/PLOS Biology