Abstract
The membrane currents of chicken embryo fibroblasts (CEFs) transformed by Rous sarcoma virus (RSV) were compared with the currents of their nontransformed counterparts by using the whole-cell patch-clamp technique. In nontransformed CEFs, the main membrane current is a delayed outward K+ current that is sensitive to tetraethylammonium ion but insensitive to 4-aminopyridine. This K+ current is almost independent of the intracellular Ca2+ concentration and becomes completely inactivated at positive membrane potentials with a time constant of about 10 s at +30 mV. In contrast, transformed CEFs exhibit a noninactivating K+ current that strongly depends on the intracellular Ca2+ concentration. This Ca(2+)-dependent K+ current is blocked by the scorpion toxin charybdotoxin with an IC50 value of 19 nM, whereas the K+ current of normal CEFs is insensitive to charybdotoxin (up to 300 nM). The K+ current properties of transformed CEFs were also found after microinjection of purified, enzymatically active pp60v-src into normal CEFs but not after infection of CEFs with the Rous-associated virus RAV5, which lacks the v-src oncogene. Our results suggest that the oncogene product pp60v-src modulates existing K+ channel proteins, leading to profound electrophysiological and pharmacological alterations of the K+ current properties in RSV-transformed CEFs. Furthermore, our experiments identify for the first time K+ channels as possible substrates of pp60v-src.
Full text
PDF




Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Becker D., Kurth R., Critchley D., Friis R., Bauer H. Distinguishable transformation-defective phenotypes among temperature-sensitive mutants of Rous sarcoma virus. J Virol. 1977 Mar;21(3):1042–1055. doi: 10.1128/jvi.21.3.1042-1055.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- DeCoursey T. E., Chandy K. G., Gupta S., Cahalan M. D. Voltage-gated K+ channels in human T lymphocytes: a role in mitogenesis? Nature. 1984 Feb 2;307(5950):465–468. doi: 10.1038/307465a0. [DOI] [PubMed] [Google Scholar]
- Deutsch C., Krause D., Lee S. C. Voltage-gated potassium conductance in human T lymphocytes stimulated with phorbol ester. J Physiol. 1986 Mar;372:405–423. doi: 10.1113/jphysiol.1986.sp016016. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dreyer F. Peptide toxins and potassium channels. Rev Physiol Biochem Pharmacol. 1990;115:93–136. [PubMed] [Google Scholar]
- Fukami Y., Lipmann F. Purification of the Rous sarcoma virus src kinase by casein-agarose and tyrosine-agarose affinity chromatography. Proc Natl Acad Sci U S A. 1985 Jan;82(2):321–324. doi: 10.1073/pnas.82.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Glossmann H., Presek P., Eigenbrodt E. Association of the src-gene product of Rous sarcoma virus with a pyruvate-kinase inactivation factor. Mol Cell Endocrinol. 1981 Jul;23(1):49–63. doi: 10.1016/0303-7207(81)90116-7. [DOI] [PubMed] [Google Scholar]
- Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F. J. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 1981 Aug;391(2):85–100. doi: 10.1007/BF00656997. [DOI] [PubMed] [Google Scholar]
- Hunter T., Cooper J. A. Protein-tyrosine kinases. Annu Rev Biochem. 1985;54:897–930. doi: 10.1146/annurev.bi.54.070185.004341. [DOI] [PubMed] [Google Scholar]
- Kaczmarek L. K. The regulation of neuronal calcium and potassium channels by protein phosphorylation. Adv Second Messenger Phosphoprotein Res. 1988;22:113–138. [PubMed] [Google Scholar]
- Kamps M. P., Buss J. E., Sefton B. M. Rous sarcoma virus transforming protein lacking myristic acid phosphorylates known polypeptide substrates without inducing transformation. Cell. 1986 Apr 11;45(1):105–112. doi: 10.1016/0092-8674(86)90542-8. [DOI] [PubMed] [Google Scholar]
- Kamps M. P., Sefton B. M. Identification of multiple novel polypeptide substrates of the v-src, v-yes, v-fps, v-ros, and v-erb-B oncogenic tyrosine protein kinases utilizing antisera against phosphotyrosine. Oncogene. 1988 Apr;2(4):305–315. [PubMed] [Google Scholar]
- Krueger J. G., Garber E. A., Goldberg A. R. Subcellular localization of pp60src in RSV-transformed cells. Curr Top Microbiol Immunol. 1983;107:51–124. [PubMed] [Google Scholar]
- Obrig T. G., Culp W. J., McKeehan W. L., Hardesty B. The mechanism by which cycloheximide and related glutarimide antibiotics inhibit peptide synthesis on reticulocyte ribosomes. J Biol Chem. 1971 Jan 10;246(1):174–181. [PubMed] [Google Scholar]
- Parsons J. T., Weber M. J. Genetics of src: structure and functional organization of a protein tyrosine kinase. Curr Top Microbiol Immunol. 1989;147:79–127. doi: 10.1007/978-3-642-74697-0_3. [DOI] [PubMed] [Google Scholar]
- Rane S. G. A Ca2(+)-activated K+ current in ras-transformed fibroblasts is absent from nontransformed cells. Am J Physiol. 1991 Jan;260(1 Pt 1):C104–C112. doi: 10.1152/ajpcell.1991.260.1.C104. [DOI] [PubMed] [Google Scholar]
- Resh M. D. Membrane interactions of pp60v-src: a model for myristylated tyrosine protein kinases. Oncogene. 1990 Oct;5(10):1437–1444. [PubMed] [Google Scholar]
- Trotter J., Boulter C. A., Sontheimer H., Schachner M., Wagner E. F. Expression of v-src arrests murine glial cell differentiation. Oncogene. 1989 Apr;4(4):457–464. [PubMed] [Google Scholar]
- Wyke J. A., Stoker A. W. Genetic analysis of the form and function of the viral src oncogene product. Biochim Biophys Acta. 1987 Apr 20;907(1):47–69. doi: 10.1016/0304-419x(87)90018-7. [DOI] [PubMed] [Google Scholar]
- van der Valk J., Verlaan I., de Laat S. W., Moolenaar W. H. Expression of pp60v-src alters the ionic permeability of the plasma membrane in rat cells. J Biol Chem. 1987 Feb 25;262(6):2431–2434. [PubMed] [Google Scholar]