Using a Modified Proximity Ligation Protocol to Study the Interaction Between Chaperones and Associated Proteins

Simone Baldan; Anatoli B Meriin; Michael Y Sherman

doi:10.1007/978-1-0716-3342-7_13

Using a Modified Proximity Ligation Protocol to Study the Interaction Between Chaperones and Associated Proteins

Methods Mol Biol. 2023:2693:163-174. doi: 10.1007/978-1-0716-3342-7_13.

Authors

Simone Baldan¹, Anatoli B Meriin², Michael Y Sherman³

Affiliations

¹ MRC Toxicology Unit, University of Cambridge, Cambridge, UK.
² Department of Biochemistry, Boston University, Boston, MA, USA. meriin@bu.edu.
³ Department of Molecular Biology, Ariel University, Ariel, Israel.

PMID: 37540434
DOI: 10.1007/978-1-0716-3342-7_13

Abstract

Molecular chaperones can interact with multiple proteins to form large networks. Understanding these interactions may shed light on the complexity of the chaperone functions. Here we developed a protocol for a modified proximity ligation-based methodology (PLA) for the detection of protein-protein interactions in order to understand how the Hsp70-Bag3 complex interacts with components of the Hippo signaling pathway. These experiments helped to elucidate the mechanisms of transmission of the proteotoxic stress signal to the Hippo pathway. The modified PLA technology has many advantages compared to co-immunoprecipitation protocols. It has higher sensitivity, is quantitative, and can be done in a 96-well format.

Keywords: Bag3; Hippo pathway; Hsp70; LATS1/2; Protein complex; Proximity ligation assay; Yap.

MeSH terms

HSP70 Heat-Shock Proteins* / metabolism
Hippo Signaling Pathway
Molecular Chaperones* / metabolism
Polyesters

Substances

Molecular Chaperones
HSP70 Heat-Shock Proteins
Polyesters