Optogenetic dephosphorylation of phosphatidylinositol 4,5 bisphosphate in Xenopus laevis oocytes

Here, we present a protocol for optogenetic dephosphorylation of the phosphoinositide PI(4,5)P₂ at the plasma membrane of Xenopus laevis oocytes. We first describe the co-injection of oocytes with cRNAs encoding (1) a light-activated PI(4,5)P₂ 5-phosphatase fusion protein, (2) its dimerization partner fused to the plasma membrane, and (3) the potassium channel reporter for PI(4,5)P₂ dephosphorylation. We then detail blue light illumination to induce PI(4,5)P₂ dephosphorylation, combined with simultaneous two-electrode voltage clamp electrophysiological recording to assess potassium channel current responses. For complete details on the use and execution of this protocol, please refer to Xu et al. (2022).¹.