Faculty Collaboration Database

Imaging the Nanoscale Distribution of Phosphoinositides in the Cell Plasma Membrane with Single-Molecule Localization Super-Resolution Microscopy. Methods Mol Biol 2021;2251:91-104

Date

01/23/2021

Pubmed ID

33481233

DOI

10.1007/978-1-0716-1142-5_6

Scopus ID

2-s2.0-85100326544 (requires institutional sign-in at Scopus site)

Abstract

Phosphoinositides make up only a small fraction of cellular phospholipids yet control cell function in a fundamental manner. Through protein interactions, phosphoinositides define cellular organelle identity and regulate protein function and organization and recruitment at the cytosol-membrane interface. As a result, perturbations on phosphoinositide metabolism alter cell physiology and lead to a wide range of human diseases, including cancer and diabetes. Among seven phosphoinositide members, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P₂, also known as PI(4,5)P₂ or PIP₂) is abundant in the plasma membrane. Besides its role in the second messenger pathway of phospholipase C that cleaves PtdIns(4,5)P₂ to form diacylglycerol and inositol-1,4,5-trisphosphate (IP₃), PtdIns(4,5)P₂ regulates membrane trafficking and the function of the cytoskeleton, ion channels, and transporters. The nanoscale organization of PtdIns(4,5)P₂ in the plasma membrane becomes essential to understand cellular signaling specificity in time and space. Here, we describe a single-molecule method to visualize the nanoscale distribution of PtdIns(4,5)P₂ in the plasma membrane by using super-resolution microscopy and the dual-color fluorescent probes based on the PLC_δ1 pleckstrin homology (PH) domain. This approach can be extended to image other phosphoinositides by changing the specific probes.

Author List

Fan F, Ji C, Lou X

Author

Xuelin Lou PhD Professor in the Cell Biology, Neurobiology and Anatomy department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Cell Culture Techniques
Cell Membrane
Humans
Membranes
Microscopy, Fluorescence
Phosphatidylinositol 4,5-Diphosphate
Phosphatidylinositol Phosphates
Phosphatidylinositols
Protein Transport
Single Molecule Imaging
Type C Phospholipases