Optogenetic Characterization of the Stress-associated Focal Adhesion to Cluster Transition of Profilin-VASP Biomolecular Condensates

Abstract

OptoProfilin, a light-sensitive biosensor derived from Arabidopsis thaliana Cryptochrome 2, triggers profilin oligomerization when exposed to blue light. The primary objectives of this study are to investigate the biochemical composition of stress-associated clusters formed with OptoProfilin and to investigate individual amino acid residues within OptoProfilin and its VASP binding partner to determine their role in its response to light activation. Ultimately, this work seeks to understand the binding proteins and cell stress pathways involving profilin and VASP that contribute to stress granule formation and biomolecular structures. The observed OptoProfilin response to energetic and oxidative stress may offer valuable information about diseases in which profilin has been implicated as a key component of disease progression. Our findings identify mutations that have significant effects on the OptoProfilin response, including the identification of a new knockout mutant of OptoProfilin activity. We have also identified focal-adhesion associated proteins (zyxin, vinculin, paxillin) that are co-localized in OptoProfilin in focal adhesion (zyxin, vinculin, paxillin) and in stress-associated clusters (zyxin, vinculin). This result supports our general hypothesis of a multi-layered model of focal adhesion composition.