2. Academic Validation
  3. High-Content Imaging Platform to Discover Chemical Modulators of Plasma Membrane Rafts

High-Content Imaging Platform to Discover Chemical Modulators of Plasma Membrane Rafts

  • ACS Cent Sci. 2022 Mar 23;8(3):370-378. doi: 10.1021/acscentsci.1c01058.
Nico Fricke 1,  Krishnan Raghunathan 1,  Ajit Tiwari 1,  Katherine M Stefanski 2,  Muthuraj Balakrishnan 3,  Alex G Waterson 4,  Ricardo Capone 1,  Hui Huang 2,  Charles R Sanders 2,  Joshua A Bauer 2 5,  Anne K Kenworthy 1
Affiliations

Affiliations

  • 1 Department of Molecular Physiology and Biophysics, Vanderbilt School of Medicine, Nashville, Tennessee 37232, United States.
  • 2 Department of Biochemistry, Vanderbilt School of Medicine, Nashville, Tennessee 37240, United States.
  • 3 Center for Membrane and Cell Physiology and Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, Virginia 22903, United States.
  • 4 Department of Pharmacology, Vanderbilt School of Medicine, Nashville, Tennessee 37232, United States.
  • 5 Vanderbilt Institute of Chemical Biology, High-Throughput Screening Facility, Vanderbilt School of Medicine, Nashville, Tennessee 37232, United States.
PMID: 35355811 DOI: 10.1021/acscentsci.1c01058
Abstract

Plasma membrane organization profoundly impacts cellular functionality. A well-known mechanism underlying this organization is through nanoscopic clustering of distinct lipids and proteins in membrane rafts. Despite their physiological importance, rafts remain a difficult-to-study aspect of membrane organization, in part because of the paucity of chemical tools to experimentally modulate their properties. Methods to selectively target rafts for therapeutic purposes are also currently lacking. To tackle these problems, we developed a high-throughput screen and an accompanying image analysis pipeline to identify small molecules that enhance or inhibit raft formation. Cell-derived giant plasma membrane vesicles were used as the experimental platform. A proof-of-principle screen using a bioactive lipid library demonstrates that this method is robust and capable of validating established raft modulators including C6- and C8-ceramide, miltefosine, and epigallocatechin gallate as well as identifying new ones. The platform we describe here represents a powerful tool to discover new chemical approaches to manipulate rafts and their components.

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