Characterization of the human pH- and PKA-activated ClC-2G(2 alpha) Cl- channel
| Autor: | John Cuppoletti, Elena Kupert, K. Stroffekova, Ann M. Sherry, Danuta H. Malinowska, L. M. Knapp |
|---|---|
| Rok vydání: | 1997 |
| Předmět: |
Adult
DNA Complementary Physiology Molecular Sequence Data Alpha (ethology) Cell Line chemistry.chemical_compound Xenopus laevis Fetus Chloride Channels Complementary DNA Extracellular medicine Animals Humans Amino Acid Sequence Protein kinase A Reversal potential Lung Ion transporter Chimera Stomach Cell Biology Tetraethylammonium chloride Hydrogen-Ion Concentration Adenosine Molecular biology Cyclic AMP-Dependent Protein Kinases chemistry Biochemistry Gastric Mucosa Oocytes Female Rabbits medicine.drug Hydrogen |
| Zdroj: | The American journal of physiology. 273(2 Pt 1) |
| ISSN: | 0002-9513 |
| Popis: | A ClC-2G(2 alpha) Cl- channel was identified to be present in human lung and stomach, and a partial cDNA for this Cl- channel was cloned from a human fetal lung library. A full-length expressible human ClC-2G(2 alpha) cDNA was constructed by ligation of mutagenized expressible rabbit ClC-2G(2 alpha) cDNA with the human lung ClC-2G(2 alpha) cDNA, expressed in oocytes, and characterized at the single-channel level. Adenosine 3',5'-cyclic monophosphate-dependent protein kinase (PKA) treatment increased the probability of opening of the channel (Po). After PKA activation, the channel exhibited a linear (r = 0.99) current-voltage curve with a slope conductance of 22.1 +/- 0.8 pS in symmetric 800 mM tetraethylammonium chloride (TEACl; pH 7.4). Under fivefold gradient conditions of TEACl, a reversal potential of +21.5 +/- 2.8 mV was measured demonstrating anion-to-cation discrimination. As previously demonstrated for the rabbit ClC-2G(2 alpha) Cl- channel, the human analog, hClC-2G(2 alpha), was active at pH 7.4 as well as when the pH of the extracellular face of the channel (trans side of the bilayer; pHtrans) was asymmetrically reduced to pH 3.0. The extent of PKA activation was dependent on pHtrans. With PKA treatment, Po increased fourfold with a pHtrans of 7.4 and eightfold with a pHtrans of 3.0. Effects of sequential PKA addition followed by pHtrans reduction on the same channel suggested that the PKA- and pH-dependent increases in channel Po were separable and cumulative. Northern analysis showed ClC-2G(2 alpha) mRNA to be present in human adult and fetal lung and adult stomach, and quantitative reverse transcriptase-polymerase chain reaction showed this channel to be present in the adult human lung and stomach at about one-half the level found in fetal lung. The findings of the present study suggest that the ClC-2G(2 alpha) Cl- channel may play an important role in Cl- transport in the fetal and adult human lung. |
| Databáze: | OpenAIRE |
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