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MEMBRANE TRANSPORTERS, ION CHANNELS, AND PUMPS

Mercury and zinc differentially inhibit shark and human CFTR orthologues: involvement of shark cysteine 102

¹Nephrology Division, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut; ²Department of Physiology and Pharmacology, Oregon Health and Science University, Portland, Oregon; and ³The Mount Desert Island Biological Laboratory, Salisbury Cove, Maine

Submitted 28 April 2005 ; accepted in final form 14 October 2005

The apical membrane is an important site of mercury toxicity in shark rectal gland tubular cells. We compared the effects of mercury and other thiol-reacting agents on shark CFTR (sCFTR) and human CFTR (hCFTR) chloride channels using two-electrode voltage clamping of cRNA microinjected Xenopus laevis oocytes. Chloride conductance was stimulated by perfusing with 10 µM forskolin (FOR) and 1 mM IBMX, and then thio-reactive species were added. In oocytes expressing sCFTR, FOR + IBMX mean stimulated Cl^– conductance was inhibited 69% by 1 µM mercuric chloride and 78% by 5 µM mercuric chloride (IC₅₀ of 0.8 µM). Despite comparable stimulation of conductance, hCFTR was insensitive to 1 µM HgCl₂ and maximum inhibition was 15% at the highest concentration used (5 µM). Subsequent exposure to glutathione (GSH) did not reverse the inhibition of sCFTR by mercury, but dithiothreitol (DTT) completely reversed this inhibition. Zinc (50–200 µM) also reversibly inhibited sCFTR (40–75%) but did not significantly inhibit hCFTR. Similar inhibition of sCFTR but not hCFTR was observed with an organic mercurial, p-chloromercuriphenylsulfonic acid (pCMBS). The first membrane spanning domain (MSD1) of sCFTR contains two unique cysteines, C102 and C303. A chimeric construct replacing MSD1 of hCFTR with the corresponding sequence of sCFTR was highly sensitive to mercury. Site-specific mutations introducing the first but not the second shark unique cysteine in hCFTR MSD1 resulted in full sensitivity to mercury. These experiments demonstrate a profound difference in the sensitivity of shark vs. human CFTR to inhibition by three thiol-reactive substances, an effect that involves C102 in the shark orthologue.

chloride transport; Xenopus laevis oocytes; dithiothreitol; glutathione; p-chloromercuriphenylsulfonic acid; cystic fibrosis transmembrane regulator

This article has been cited by other articles:

				M. S. Bewley, J. T. G. Pena, F. N. Plesch, S. E. Decker, G. J. Weber, and J. N. Forrest Jr. Shark rectal gland vasoactive intestinal peptide receptor: cloning, functional expression, and regulation of CFTR chloride channels Am J Physiol Regulatory Integrative Comp Physiol, October 1, 2006; 291(4): R1157 - R1164. [Abstract] [Full Text] [PDF]