Towsif Ekram M, Shekhar Shashank
Departments of Physics, Cell Biology and Biochemistry, Emory University, Atlanta, GA 30322.
Proc Natl Acad Sci U S A. 2025 Feb 4;122(5):e2411318122. doi: 10.1073/pnas.2411318122. Epub 2025 Jan 28.
Cellular actin networks exhibit distinct assembly and disassembly dynamics, primarily driven by multicomponent reactions occurring at the two ends of actin filaments. While barbed ends are recognized as the hotspot for polymerization, depolymerization is predominantly associated with pointed ends. Consequently, mechanisms promoting barbed-end depolymerization have received relatively little attention. Here, using microfluidics-assisted three-color single-molecule imaging, we reveal that cyclase-associated protein (CAP), long known for its roles in nucleotide exchange and pointed-end depolymerization, also acts as a processive depolymerase at filament barbed ends. CAP molecules track barbed ends for several minutes, inducing depolymerization rates of up to 60 subunits per second. Importantly, CAP modulates barbed-end dynamics even under cytosol-mimicking assembly promoting conditions. We further show that CAP can colocalize with both formin and capping protein (CP) at barbed ends. CAP enhances formin processivity by 10-fold, allowing CAP-formin complexes to track fast-elongating barbed ends. In contrast, CAP destabilizes CP-bound barbed ends and accelerates dissociation of CP by fourfold. Our findings, combined with CAP's previously reported activities, firmly establish CAP as a key regulator of cellular actin dynamics.
细胞肌动蛋白网络呈现出独特的组装和解聚动力学，主要由肌动蛋白丝两端发生的多组分反应驱动。虽然带刺端被认为是聚合的热点，但解聚主要与尖端相关。因此，促进带刺端解聚的机制受到的关注相对较少。在这里，我们使用微流控辅助三色单分子成像技术，揭示了环化酶相关蛋白（CAP），长期以来因其在核苷酸交换和尖端解聚中的作用而闻名，它在细丝带刺端也作为一种持续性解聚酶发挥作用。CAP分子在带刺端追踪数分钟，诱导每秒高达60个亚基的解聚速率。重要的是，即使在模拟胞质溶胶的组装促进条件下，CAP也能调节带刺端的动力学。我们进一步表明，CAP可以在带刺端与formin和封端蛋白（CP）共定位。CAP将formin的持续性提高了10倍，使CAP-formin复合物能够追踪快速伸长的带刺端。相比之下，CAP使CP结合的带刺端不稳定，并使CP的解离加速四倍。我们的研究结果，结合CAP先前报道的活性，牢固地确立了CAP作为细胞肌动蛋白动力学的关键调节因子。
