Vascular smooth muscle cells (VSMC) can exhibit a contractile or a synthetic phenotype depending on the extracellular stimuli present and the composition of the extracellular matrix. Uncontrolled activation of the synthetic VSMC phenotype is however associated with the development of cardiovascular diseases. Here, we aimed to elucidate the role of the ARF GTPases in the regulation of VSMC dedifferentiation. First, we observed that the inhibition of the activation of ARF proteins with SecinH3, a blocker of the cytohesin ARF GEF family, reduced the ability of the cells to migrate and proliferate. In addition, this inhibitor also blocked expression of sm22α and αSMA, two contractile markers, at the transcription level impairing cell contractility. Specific knockdown of ARF1 and ARF6 showed that both isoforms were required for migration and proliferation, but ARF1 only regulated contractility through sm22α and αSMA expression. Expression of these VSMC markers was correlated with the degree of actin polymerization. VSMC treatment with SecinH3 as well as ARF1 depletion was both able to block the formation of stress fibres and focal adhesions, demonstrating the role of this GTPase in actin filament formation. Consequently, we observed that both treatments increased the ratio of G-actin to F-actin in these cells. The elevated amounts of cytoplasmic G-actin, acting as a signaling intermediate, blocked the recruitment of the Mkl1 (MRTF-A) transcription factor in the nucleus, demonstrating its involvement in the regulation of contractile protein expression. Altogether, these findings show for the first time that ARF GTPases are actively involved in VSMC phenotypic switching through the regulation of actin function in migration and proliferation, and the control of actin dependent gene regulation.

血管平滑肌细胞（VSMC）可能显示收缩或合成表型，具体取决于存在的细胞外刺激和细胞外基质的组成。然而，合成的VSMC表型的不受控制的激活与心血管疾病的发展有关。在这里，我们旨在阐明ARF GTPases在调节VSMC去分化中的作用。首先，我们观察到SecinH3（细胞粘附素ARF GEF家族的阻滞剂）对ARF蛋白激活的抑制作用降低了细胞迁移和增殖的能力。另外，该抑制剂还在转录水平上阻断了sm22α和αSMA这两个收缩标记的表达，从而损害了细胞的收缩能力。ARF1和ARF6的特异性敲低表明迁移和增殖均需要两种同工型，但ARF1仅通过sm22α和αSMA表达调节收缩力。这些VSMC标志物的表达与肌动蛋白聚合度相关。用SecinH3以及ARF1耗竭进行VSMC处理均能够阻止应激纤维的形成和粘着斑，这证明了该GTPase在肌动蛋白丝形成中的作用。因此，我们观察到两种处理均增加了这些细胞中G-肌动蛋白与F-肌动蛋白的比率。充当信号传递中间体的细胞质G-肌动蛋白数量增加，阻止了Mkl1（MRTF-A）转录因子在细胞核中的募集，表明其参与了对收缩蛋白表达的调节。共，