Temporally gated molecular tools for tracking protein-protein interactions in live cells

Kayla E Kroning; Wenjing Wang

doi:10.1016/bs.mie.2020.04.029

Temporally gated molecular tools for tracking protein-protein interactions in live cells

Methods Enzymol. 2020:640:205-223. doi: 10.1016/bs.mie.2020.04.029. Epub 2020 May 7.

Authors

Kayla E Kroning¹, Wenjing Wang²

Affiliations

¹ Department of Chemistry, University of Michigan, Ann Arbor, MI, United States; Life Sciences Institute, University of Michigan, Ann Arbor, MI, United States.
² Department of Chemistry, University of Michigan, Ann Arbor, MI, United States; Life Sciences Institute, University of Michigan, Ann Arbor, MI, United States. Electronic address: wenjwang@umich.edu.

PMID: 32560799
DOI: 10.1016/bs.mie.2020.04.029

Abstract

Protein-protein interactions (PPIs) are essential in most biological processes. Even though many methods were designed to detect PPIs, detecting PPIs in a large volume of cells with a temporal resolution remains challenging. Recent development of light gated transcriptional reporters, such as SPARK and iTANGO, enabled detection of PPI in a large population of cells with a temporal resolution on the order of minutes. In this chapter, we discussed in detail the application of SPARK to detect PPIs between the activated β-2 adrenergic receptor (B2AR) and both Gα mimic and β-arrestin2. Because SPARK is a multi-component system, the protein expression level is critical for its optimal performance. We also discussed the detailed protocols for using SPARK with either transfection or lentiviral infection in HEK296T/17 cells.

Keywords: Light-gated; Protein-protein interaction; SPARK; Transcriptional readout.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Protein Interaction Mapping*
Proteins*
Signal Transduction
Transfection

Substances

Proteins