Anion exchanger isoform 2 operates in parallel with Na+/H+ exchanger isoform 1 during regulatory volume decrease of human cervical cancer cells

FEBS Letters

Ellory, J.Clive¹  Shen, Meng-Ru¹  Wilkins, Robert J.¹  Chou, Cheng-Yang² 

[1] University Laboratory of Physiology, Parks Road, Oxford OX1 3PT, UK;Department of Obstetrics and Gynecology, College of Medicine, National Cheng Kung University, Tainan 704, Taiwan

关键词: Na+/H+ exchanger;    Anion exchanger;    Regulatory volume decrease;   

DOI  :  10.1016/S0014-5793(01)03317-8

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Intracellular pH (pH_i) homeostasis was investigated in human cervical cancer SiHa cells undergoing regulatory volume decrease (RVD) to determine which transport systems were involved. Using isoform-specific primers, mRNA transcripts of Na⁺/H⁺ exchanger isoform 1 (NHE1) and isoform 3 were identified by reverse transcriptase polymerase chain reaction (RT-PCR) and the results confirmed by Western immunoblotting. From anion exchanger isoforms 1–3 (AE1–3), only the mRNA transcript of AE2 was identified by RT-PCR and the identity was confirmed by digestion with a specific restriction endonuclease. SiHa cells loaded with the fluorescent dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein and resuspended in isotonic media showed a stable pH_i. In contrast, a gradual internal acidification took place following resuspension in hypotonic media. The NHE inhibitors, HOE694 (10 μM) and amiloride (1 mM), showed a similar potency in enhancing the rate and extent of the hypotonicity-induced internal acidification. The absence of extracellular Na⁺ also substantially enhanced the acidification during RVD. These results suggest that internal acidification during RVD is mainly compensated by the operation of NHE1. Extracellular Cl⁻ was critically necessary for the pH_i acidification during RVD. The hypotonicity-induced acidification was significantly attenuated by 100 μM 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid, a concentration inhibiting more than 90% AE activity. This indicates that AE2 mediates a net Cl⁻ influx with compensating HCO₃ ⁻ efflux during RVD. We conclude that AE2 operates in parallel with NHE1 to regulate pH_i during RVD of human cervical cancer cells.

【 授权许可】

Unknown