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Regulation of K-Cl cotransport by Syk and Src protein tyrosine kinases in deoxygenated sickle cells

  • Ion Channels, Transporters
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Abstract

Protein tyrosine kinases (PTK) of the Src family are thought to suppress K-Cl cotransport (KCC) activity via negative regulation of protein phosphatases. However, some PTK inhibitors reduce KCC activity, suggesting opposite regulation by different PTK families. We have reported previously that deoxygenation of sickle cells stimulates KCC and activates Syk (a Syk family PTK), but not Lyn (an Src family PTK). In this study the same results were obtained when PTK activities were measured under the conditions used to measure KCC activity and which prevent any change in intracellular [Mg2+]. Methyl-2,5-dihydroxycinnamate (DHC), a PTK inhibitor, was more selective for Syk than Lyn, while staurosporine (ST), a broad-specificity protein kinase inhibitor, inhibited Lyn more than Syk. Deoxygenation or 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d] pyrimidine (pp2, a specific Src inhibitor) stimulated KCC independently. These effects were not additive and were inhibited by DHC. In contrast, ST-induced KCC activation was resistant to DHC, suggesting a different pathway of activation. Overall, these data indicate that Syk activity is required for KCC activation, either induced by deoxygenation of sickle cells, or mediated by Src inhibition in oxygenated cells, and that Syk and Src PTKs exert opposing and interconnected regulatory effects on the activity of the transporter.

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Fig. 1.
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Acknowledgements

The authors thank Drs. D. Bachir and F. Galactéros (Centre de la Drépanocytose et des Thalassémies, Hôpital Henri Mondor, Créteil, France) for providing blood samples from their patients. This work was supported by the Centre National de la Recherche Scientifique (CNRS, UMR 8619), the Université Paris XI-Orsay, the Société Française d'Hématologie to P. M. and the National Institutes of Health, Grant P60HL58421 (CHJ).

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Authors and Affiliations

  1. Biomembranes et Messagers Cellulaires, Centre National de la Recherche Scientifique UMR 8619, Bat 440, Université Paris XI, 91405, Orsay Cedex, France

    P. Merciris & F. Giraud

  2. Divisions of Pediatric and Medical Hematology/Oncology, Cincinnati Comprehensive Sickle Cell Center, Cincinnati, Ohio, 45229-3039, USA

    W. J. Claussen & C. H. Joiner

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  1. P. Merciris
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  2. W. J. Claussen
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  3. C. H. Joiner
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  4. F. Giraud
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Correspondence to F. Giraud.

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Merciris, P., Claussen, W.J., Joiner, C.H. et al. Regulation of K-Cl cotransport by Syk and Src protein tyrosine kinases in deoxygenated sickle cells. Pflugers Arch - Eur J Physiol 446, 232–238 (2003). https://doi.org/10.1007/s00424-003-1025-z

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