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J.K. Klarlund, A.R. Mazie, J.K. Spix, E.R. Block; Corneal Epithelial Cell Motility Is Triggered by Activation of the Epidermal Growth Factor Receptor Through Phosphatidic Acid Signaling . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5035.
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There is great interest in understanding the signals that induce motility in corneal epithelial cells, since signaling pathways are attractive pharmacological targets. We have previously shown that induction of motility upon wounding is strictly dependent on activation of the epidermal growth factor receptor (EGFR) through proteolytic activation of the precursor for one of its ligands (proHB–EGF). The purpose of these studies was to identify signals that lead to EGFR activation.
The studies relied on a previously published wounding assay that allows biochemical analysis of processes induced by wounding. Phospholipase D activities were assayed by labeling cells with [3H]myristic acid, adding 1–BuOH, and measuring formation of phosphatidylinositol butanol. Addition of short chain, water–soluble analogues of phosphatidic acid was used as a means of stimulating phosphatidic acid signaling.
Wounding activates phospholipase D, an enzyme that catalyzes the hydrolysis of phosphatidylcholine to generate phosphatidic acid. Short–chain analogues of phosphatidic acid induce motility robustly in corneal and other epithelial cell types, and we provide evidence that phosphatidic acid acts directly on one or more cellular targets. Phosphatidic acid was found to trigger transactivation of the epidermal growth factor receptor via proHB–EGF cleavage. This is surprising because phospholipase D has so far been considered a downstream target for the EGFR. Inhibition of phosphatidic acid signaling in the presence of EGF indicated that EGFR activation is not sufficient for induction of motility, and that phosphatidic acid has additional signaling activities.
These observations, together with other data, lead us to suggest that stimulation of phospholipase D is an immediate consequence of wounding, and that the EGFR is activated through phospholipase D activation. We propose the following signaling circuitry for induction of motility in corneal epithelial cells (PLD: phospholipase D; PA: phosphatidic acid; ERK: extracelular regulated kinase):
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