Piezo1 are mechanogated Ca2+-permeable channels that are important participants of calcium signaling in cells. The changes of Piezo1 properties and regulation can lead to modulation of fundamental cellular processes and reactions. Current concepts suggest that Piezo1 activity is regulated via the combined action of plasma membrane lipids, components of extracellular matrix, and cytoskeleton. At the same time, the question about regulation of Piezo1 conductive properties by F-actin structure remains unexplored. Here, we selected human melanoma SK-MEL-2 cell line to register single Yoda1-induced Piezo1 currents before and after F-actin destruction by depolymerizing agent cytochalasin D (CytD). Single-channel analysis evidenced the decrease of Piezo1 amplitudes and conductance values after acute CytD treatment. F-actin dynamics visualization showed that time-dependent changes in actin organization coincide with the dynamics of decrease in single-channel Piezo1 currents. The results demonstrate a novel mechanism for regulating fundamental Piezo1 properties by the state of actin structures in living cells.