title: Interaction of a Novel Chaperone PhLP2A With the Heat Shock Protein Hsp90.
abstract: PhLP2 is a small cytosolic protein that belongs to the highly conserved phosducin-like family of proteins. In amniote genomes there are two PhLP2 homologs, PhLP2A and PhLP2B. It has been shown that mammalian PhLP2A modulates the CCT/TRiC chaperonin activity during folding of cytoskeletal proteins. In order to better understand the function of PhLP2A in cellular protein quality control system, in the present study we have searched for its protein targets. Applying immunoprecipitation followed by mass spectrometry analysis we have identified Hsp90 as a partner of PhLP2A. With pull down experiments, we have confirmed this interaction in protein lysate and using purified proteins we have shown that PhLP2A interacts directly with Hsp90. Furthermore, the proximity ligation assay (PLA) performed on mIMCD-3 cells has shown that PhLP2A forms complexes with Hsp90 which are mainly localized in the cytoplasm of these cells. Further analysis has indicated that the level of PhLP2A increases after heat shock or radicicol treatment, similarly as the level of Hsp90, and that expression of PhLP2A after heat shock is regulated at the transcriptional level. Moreover, using recombinant luciferase we have shown that PhLP2A stabilizes this enzyme in a folding competent state and prevents its denaturation and aggregation. In addition, overexpression of PhLP2A in HEK-293 cells leads to increased heat stress resistance. Altogether, our results have shown that PhLP2A interacts with Hsp90 and exhibits molecular chaperone activity toward denatured proteins. J. Cell. Biochem. 118: 420-429, 2017. © 2016 Wiley Periodicals, Inc.
authors: Laboratory of Cytoskeleton and Cilia Biology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, Warsaw, 02-093, Poland.
authors: Łucja
authors: Ł
authors: Krzemień-Ojak
authors: Laboratory of Calcium Binding Proteins, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, Warsaw, 02-093, Poland.
authors: Agnieszka
authors: A
authors: Góral
authors: Laboratory of Cytoskeleton and Cilia Biology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, Warsaw, 02-093, Poland.
authors: Ewa
authors: E
authors: Joachimiak
authors: Laboratory of Calcium Binding Proteins, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, Warsaw, 02-093, Poland.
authors: Anna
authors: A
authors: Filipek
authors: Laboratory of Cytoskeleton and Cilia Biology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, Warsaw, 02-093, Poland.
authors: Hanna
authors: H
authors: Fabczak
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D018869:Heat-Shock Response / Q000187:drug effects*; D006801:Humans; D018942:Macrolides / Q000494:pharmacology*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D011485:Protein Binding; D017510:Protein Folding / Q000187:drug effects*; D055550:Protein Stability; D051381:Rats
chemical_list: D002352:Carrier Proteins; D018841:HSP90 Heat-Shock Proteins; D018942:Macrolides; D009419:Nerve Tissue Proteins; C498322:PDCL3 protein, human; C035359:monorden
keywords: CHAPERONES; HEAT SHOCK; Hsp90; PhLP2