Blending theory with experiments reveals multiplayer actin dynamics
August 26, 2024 | Bharti Dharapuram
Using cutting-edge experiments and theory, researchers have devised a model of multi-protein interactions (above) that regulate active depolymerization. Image adapted from Arya, Choubey & Shekhar (2024) and shared under a CC by 4.0 license.
Actin is a protein that makes the scaffolding of our cells and organelles, helping them move and change shape. To do this, an actin filament changes in length by losing or stringing together subunits mediated by many regulatory proteins. While we know the regulatory function of single proteins, we do not fully understand how they collectively orchestrate actin dynamics.
Combining single-molecule experiments with theoretical models, researchers have developed a neat interaction map of proteins regulating actin depolymerization. Working with three regulatory proteins, they discovered previously unknown interactions and found that pairwise interactions explain their collective influence on actin disassembly. The study was carried out by the collaborative efforts of Ankita Arya and Shashank Shekhar from Emory University and Sandeep Choubey from The Institute of Mathematical Sciences, Chennai.
A cell responds to changes in its environment by breaking down and reassembling actin filaments to remodel its structural framework. Profilin, twinfilin and cofilin are three important regulatory proteins that mediate actin disassembly by binding to its barbed end. While we have an understanding of their one-way interactions with actin, traditional methods are limiting when studying their simultaneous transient interactions. In a new study, researchers used microfluidics and fluorescence imaging to make measurements of actin depolymerization rates by adding regulatory proteins both singly and together. They tested hypotheses about competitive or cooperative protein binding via an effective dialogue between theory and experiments.
The researchers found that profilin and cofilin can simultaneously bind to actin accelerating its depolymerization rate. Twinfilin competes with profilin for actin binding, while assisting cofilin in binding to the sides of an actin filament. Finally, they showed that these pairwise relationships can explain actin dynamics in the presence of all three regulatory proteins in a bottom-up manner.
“Our worldview is that you can understand living matter by using simple mathematical laws. We use ideas from statistical mechanics and statistics to understand these systems,” says Choubey, explaining his motivation. The ideas for this study developed over conversations with Shekhar, whose research group uses lab experiments to understand the long-standing mystery of how cells control actin remodeling. “Actin is a nice model system because you can make high-throughput measurements of changes in the length of hundreds of filaments in a precise quantitative manner. Without these lab measurements, one really cannot test models,” Choubey says.
They proposed simple models of competitive or cooperative interactions between regulatory proteins and tested predictions of actin depolymerization rates against experiments. “There is a dialogue between theory and experiments – you build a simple model, make a prediction and test it in experiments. If it doesn’t match, you come back and modify your model. You can falsify models to get to the right mechanism,” Choubey explains. “You can think of the modeling approach as a different microscope, an alternative method of observing what is happening within cells.”
This approach proved useful as they found evidence that multi-component interactions with actin are often contrary to what is expected from single protein studies. They also found that pairwise interactions between regulatory proteins are enough to explain their collective role in actin depolymerization. “This bodes well for the future. If we can map out pairwise interactions, we may have a way to understand what happens at the cellular level,” Choubey says.
Looking ahead, the group is working on a generalized theoretical framework for interpreting such high-throughput datasets. “We want to develop a framework for precision measurements, which is like a theory of the experiment that you can test and falsify,” Choubey says.
Reference: Arya, A., Choubey, S., & Shekhar, S. (2024). Actin filament barbed-end depolymerization by combined action of profilin, cofilin, and twinfilin. PRX Life, 2(3), 033002. https://doi.org/10.1103/PRXLife.2.033002
