Hsp90 prevents interaction between CHIP and HERG proteins to facilitate maturation of wild-type and mutant HERG proteins.

Abstract

AIMS:

We examined the role of Hsp90 in expression and maturation of wild-type (WT) and mutant ether-a-go-go related gene (HERG) proteins by using Hsp90 inhibitors, geldanamycin (GA) and radicicol, and Hsp90 overexpression.

METHODS AND RESULTS:

The proteins were expressed in HEK293 cells or collected from HL-1 mouse cardiomyocytes, and analysed by western blotting, immunoprecipitation, immunofluorescence, and whole-cell patch-clamp techniques. GA and radicicol suppressed maturation of HERG-FLAG proteins and increased their immature forms. Co-expression of Hsp90 counteracted the effects of Hsp90 inhibitors and suppressed ubiquitination of HERG proteins. Overexpressed Hsp90 also inhibited the binding of endogenous C-terminus of Hsp70-interacting protein (CHIP) to HERG-FLAG proteins. Hsp90-induced increase of functional HERG proteins was verified by their increased expression on the cell surface and enhanced HERG channel currents. CHIP overexpression decreased both mature and immature forms of HERG-FLAG proteins in cells treated with GA. Hsp90 facilitated maturation of endogenous ERG proteins, whereas CHIP decreased both forms of ERG proteins in HL-1 cells. Mutant HERG proteins harbouring disease-causing missense mutations were mainly in the immature form and had a higher binding capacity to CHIP than the WT; Hsp90 overexpression suppressed this association. Overexpressed Hsp90 increased the mature form of HERG(1122fs/147) proteins, reduced its ubiquitinated form, increased its immunoreactivity in the endoplasmic reticulum and on the plasma membrane, and increased the mutant-mediated membrane current. CHIP overexpression decreased the immature form of HERG(1122fs/147) proteins.

CONCLUSION:

Enhancement of HERG protein expression through Hsp90 inhibition of CHIP binding might be a novel therapeutic strategy for long QT syndrome 2 caused by trafficking abnormalities of HERG proteins.

• Authors: Chisato Iwai, Peili Li, Yasutaka Kurata, Yoshiko Hoshikawa, Kumi Morikawa, Nani Maharani, Katsumi Higaki, Tetsuro Sasano, Tomomi Notsu, Yuko Ishido, Junichiro Miake, Yasutaka Yamamoto, Yasuaki Shirayoshi, Haruaki Ninomiya, Akira Nakai, Shigeo Murata, Akio Yoshida, Kazuhiro Yamamoto, Masayasu Hiraoka, Ichiro Hisatome
• Journal: Cardiovascular research (Cardiovasc Res) Vol. 100 Issue 3 Pg. 520-8 (Dec 01 2013) ISSN: 1755-3245 [Electronic] England
• PMID: 23963841 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Benzoquinones
• ERG1 Potassium Channel
• Ether-A-Go-Go Potassium Channels
• HSP90 Heat-Shock Proteins
• KCNH2 protein, human
• Kcnh2 protein, mouse
• Lactams, Macrocyclic
• Macrolides
• STUB1 protein, human
• Ubiquitin-Protein Ligases
• monorden
• geldanamycin

Topics

• Animals
• Benzoquinones (pharmacology)
• Cell Membrane (enzymology)
• ERG1 Potassium Channel
• Endoplasmic Reticulum (enzymology)
• Ether-A-Go-Go Potassium Channels (genetics, metabolism)
• HEK293 Cells
• HSP90 Heat-Shock Proteins (antagonists & inhibitors, genetics, metabolism)
• Humans
• Lactams, Macrocyclic (pharmacology)
• Long QT Syndrome (enzymology, genetics)
• Macrolides (pharmacology)
• Membrane Potentials
• Mice
• Mutation, Missense
• Myocytes, Cardiac (drug effects, enzymology)
• Protein Transport
• Transfection
• Ubiquitin-Protein Ligases (genetics, metabolism)
• Ubiquitination