Research Article: Hypoxia induced hERG trafficking defect linked to cell cycle arrest in SH-SY5Y cells

Date Published: April 24, 2019

Publisher: Public Library of Science

Author(s): Damodara Reddy Vaddi, Lin Piao, Shakil A. Khan, Ning Wang, Nanduri R. Prabhakar, Jayasri Nanduri, Salvatore V. Pizzo.


The alpha subunit of the voltage gated human ether-a-go-go-related (hERG) potassium channel regulates cell excitability in a broad range of cell lines. HERG channels are also expressed in a variety of cancer cells and control cell proliferation and apoptosis. Hypoxia, a common feature of tumors, alters gating properties of hERG currents in SH-SY5Y neuroblastoma cells. In the present study, we examined the molecular mechanisms and physiological significance underlying hypoxia-altered hERG currents in SH-SY5Y neuroblastoma cells. Hypoxia reduced the surface expression of 150kDa form and increased 125kDa form of hERG protein expression in the endoplasmic reticulum (ER). The changes in protein expression were associated with ~50% decrease in hERG potassium conductance. ER retention of hERG 125kDa form by CH was due to defective trafficking and was rescued by exposing cells to hypoxia at low temperatures or treatment with E-4031, a hERG channel blocker. Prolonged association of hERG with molecular chaperone Hsp90 resulting in complex oligomeric insoluble aggregates contributed to ER accumulation and trafficking defect. Hypoxia increased reactive oxygen species (ROS) levels and manganese (111) tetrakis (1methyl-4-pyridyl) porphyrin pentachloride, a membrane-permeable antioxidant prevented hypoxia-induced degradation of 150kDa and accumulation of 125kDa forms. Impaired trafficking of hERG by hypoxia was associated with reduced cell proliferation and this effect was prevented by antioxidant treatment. These results demonstrate that hypoxia through increased oxidative stress impairs hERG trafficking, leading to decreased K+ currents resulting in cell cycle arrest in SH-SY5Y cells.

Partial Text

The human ether-a-go-go-related gene (hERG), the α subunit of a voltage gated potassium channel encodes a rapidly activating delayed rectifier current (Ikr) [1]. Congenital or drug induced disruptions of the hERG channel cause long QT syndrome type 2 (LQT2), a cardiac disorder that predisposes affected individuals to ventricular arrhythmias and cardiac arrest [2, 3]. A majority (~80%) of the hERG missense mutations thus far studied are due to defective trafficking of hERG protein to the cell surface [4–7]. hERG protein synthesized in the endoplasmic reticulum (ER), as an immature core glycosylated protein (cg) of about 125kDa, is exported to the Golgi apparatus for complex glycosylation and eventually inserted into the plasma membrane as fully glycosylated mature protein (fg) of ~150kDa [8, 9]. HERG maturation and trafficking of the protein to the cell surface is regulated by the molecular chaperone Hsp90, which protects proteins from misfolding and degradation [10].

Major findings of the present study are: a) continuous hypoxia (CH) reduces membrane expression with concomitant increase in ER accumulation of hERG protein in SH-SY5Y cells, b) reduced membrane expression was due to defective trafficking of hERG protein from ER , c) the effects of CH on hERG protein were associated with reduced hERG K+ conductance and inhibition cell proliferation, and d) CH increased ROS abundance and anti-oxidant treatment prevented the trafficking defect and restored the membrane expression of the protein, K+ conductance and cell proliferation.