Hsp90 Prevents Interaction between CHIP and HERG Proteins to Facilitate Maturation of Wild-type and Mutant HERG Proteins.

We examined the role of Hsp90 in expression and maturation of wild-type and mutant HERG proteins by using Hsp90 inhibitors, geldanamycine (GA) and radicicol, and Hsp90 overexpression.Methods and ResultsThe proteins were expressed in HEK293 cells or collected from HL-1 mouse cardiomyocytes, and analyzed 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 harboring disease-causing missense mutations were mainly in the immature form and had a higher binding capacity to CHIP than the wild-type; 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.Enhancement of HERG protein expression through Hsp90 inhibition of CHIP binding might be a novel therapeutic strategy for LQT2 caused by trafficking abnormalities of HERG proteins.
Cardiovasc. Res.
Date: Aug. 20, 2013
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