ABSTRACT

Acid extracts of thapsigargin-activated Jurkat cells have been shown to have intracellular activity in inducing a dose-dependent rapid chloride current upon microinjection in Xenopus laevis oocytes. The extracts act by elevation of calcium through calcium entry. The factor(s) responsible for this activity have been termed calcium influx factor (CIF) and have been found to be small, relatively polar molecules (<1000 daltons) whose activity is abolished by alkaline phosphatase treatment and potentiated by co-injection of okadaic acid (a protein phosphatase inhibitor). CIF is produced in a time-dependent manner following thapsigargin treatment of Jurkat cells, being first elevated above basal levels by 2 min. Intracellular CIF activity is completely absent from NG115-401L neuronal cells, which lack capacitative entry. On this basis, it appears that Jurkat cells, activated by stimuli that deplete internal calcium stores, produce one or more CIF activities acting intracellularly, and Xenopus oocytes may be a powerful tool to purify and characterize CIFs.

FOOTNOTES

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This research was supported in part by grants from the National Institutes of Health and the Council for Tobacco Research. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked ``advertisement'' in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

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Supported by a National Institutes of Health training grant in molecular and cellular biology.

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To whom correspondence should be addressed. Tel.: 916-752-8332; Fax: 916-752-3516; mrhanley{at}ucdavis.edu.

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The abbreviations used are: InsP, inositol 1,4,5-trisphosphate; TG, thapsigargin; CPA, cyclopiazonic acid; PHA, phytohemagglutinin; CIF, calcium influx factor; HBSS, Hanks' balanced salt solution; LPA, oleoyl lysophosphatidic acid.

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H-Y. Kim, D. Thomas, and M. R. Hanley, manuscript in preparation.

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