The actin-associated protein Kaptin modulates F-actin barbed-end dynamics

Living cells require a dynamic and precisely regulated actin cytoskeleton to carry out normal cellular functions. In addition to well-established actin cytoskeleton regulators, such as nucleators, capping proteins, and bundlers, cells likely have uncharacterized modulators that regulate cytoskeleton dynamics, the detailed functions of which are not yet fully understood. In this study, we conducted biochemical exploration to identify the actin-regulatory activity of Kaptin (KPTN), a protein known to co-localize with actin-rich structures at the cells periphery. Using single-molecule assays, we demonstrated that KPTN inhibits actin nucleation. Our results revealed that KPTN possesses a novel barbed-end capping activity, which stabilizes and bundles actin filaments. Structural modeling, based on AlphaFold, suggests that KPTN is a member of the WD-repeat-containing protein family. Furthermore, we identified a crucial cationic residue in the putative N-terminal beta-propeller region of KPTN that plays a critical role in modulating actin dynamics. In summary, our data unveil the mechanistic underpinning functions of KPTN and establish its novel role as a regulator of the actin cytoskeleton.