Actin-Regulator Feedback Interactions during Endocytosis.

Endocytosis is a key mechanism for receptor internalization and signaling regulation. In some contexts, this process relies on transient actin polymerization. While prior research has shown that actin and its regulators are involved in endocytosis, the precise interactions governing actin pulse formation remain unclear. This uncertainty motivated a deeper investigation into how actin regulators behave during endocytosis. Specifically, the role of feedback mechanisms in regulating actin dynamics has not been fully resolved. Earlier studies have demonstrated that actin polymerization is necessary for clathrin-mediated endocytosis in yeast. However, the exact sequence of protein interactions that lead to actin pulses is still unknown. The gap in understanding actin-regulator feedback interactions prompted the current study. Prior work has also shown that the Arp2/3 complex is essential for actin nucleation in this process. Yet, the effects of mutations and drug treatments on actin and regulator assembly remain poorly understood.

The goal of this study is to identify the key protein-protein interactions that drive actin pulse formation during endocytosis. The researchers aim to determine how mutations and drug treatments affect actin and regulator assembly. They focus on the budding yeast Saccharomyces cerevisiae as a model organism. The study combines computational modeling with experimental validation. The researchers want to understand how actin and its regulators behave in pulses during endocytosis. They also seek to clarify the effects of reduced regulator branching activity and altered actin disassembly rates. The study addresses the question of whether actin regulators function independently or in a coordinated manner. By integrating modeling and experiments, the researchers hope to uncover the underlying feedback mechanisms.

The study uses a combination of computational modeling and experimental techniques. Two models are employed: a detailed three-dimensional stochastic model of actin network growth and a simplified two-variable model inspired by the Fitzhugh-Nagumo system. Both models incorporate a negative feedback interaction between F-actin and Arp2/3 regulators. The researchers simulate how actin and regulator dynamics change under different conditions. They also perform experiments using quantitative fluorescence microscopy on yeast cells. Mutations in Arp2/3 regulators are introduced to test their effects on actin pulse formation. The experiments measure the accumulation of regulators in response to genetic changes. The models are validated against experimental data to confirm their accuracy.

The models successfully explain the observed actin pulse dynamics during endocytosis. They predict that reduced regulator branching activity leads to increased F-actin peak counts. The researchers observed that slowing actin disassembly also increases F-actin levels. Latrunculin treatment was found to prolong the lifetime of Arp2/3 regulators. Experiments confirmed that decreased regulator branching activity causes increased regulator accumulation. The data suggest that regulators function quasi-independently during endocytosis. Each regulator's accumulation is most strongly influenced by its own mutation. The models and experiments align in showing how feedback interactions shape actin dynamics. These findings provide a clearer picture of how actin pulses are regulated.

The study demonstrates that actin pulse formation during endocytosis is influenced by specific protein-protein interactions. The models and experiments confirm that reduced regulator branching activity increases F-actin levels. The researchers propose that negative feedback from F-actin onto Arp2/3 regulators is a key mechanism. They also suggest that slowing actin disassembly leads to higher F-actin accumulation. The study shows that latrunculin treatment extends the lifetime of Arp2/3 regulators. The findings indicate that regulator accumulation is most affected by mutations in the same regulator. The researchers conclude that regulators act quasi-independently during endocytosis. These results provide insights into how actin dynamics are regulated in yeast endocytosis.