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

Abstract

Summary: 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 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cell culture
• Cell Membrane
• Gene Expression
• Model Organisms
• Molecular Biology