Determination of the molecular assembly of actin and actin-binding proteins using photoluminescence.

Actin, the most abundant protein in cells, polymerizes into filaments that play key roles in many cellular dynamics. To understand cell dynamics and functions, it is essential to examine the cytoskeleton structure organized by actin and actin-binding proteins. Here, we pave the way for determining the molecular assembly of the actin cytoskeleton using direct photonic in situ analysis, providing the photoluminescence characteristics of actin as a function of filament length and bundling, without labeling. In experiments for monomeric and filamentous actin reconstituted in vitro, structural forms of actin are identified from the peak positions and intensities of photoluminescence. Actin monomers exhibited small intensity emission peaks at 334 nm, whereas filamentous and bundled actin showed a shifted peak at 323 nm with higher intensity. The peak shift was found to be proportional to the length of the actin filament. With probing structural change of actin in various cells in vivo, our study provides an efficient and precise analytical in situ tool to examine the cytoskeleton structure. It will be beneficial for elucidating the mechanism of various cellular functions such as cell migration, differentiation, cytokinesis and adhesion. Furthermore, our technique can be applied to detect the alterations in the cell cytoskeleton that can occur during many pathological processes.