GAP43 Couples Hypoosmolarity to Calcium Signaling

Abstract

Cells can respond to changes in osmolarity and mechanical stimuli--changes in osmolarity cause cell swelling and shrinking, which can deform the membrane; this is a form of "mechanical" stimulus. In specialized mechanosensory cells, mechanical forces alter channel gating. In sensory neurons, which respond to changes in osmolarity and mechanical forces with calcium transients, the osmo- and mechanosensor(s) have not been identified. Caprini et al. used expression cloning with cDNA from dorsal root ganglion to identify GAP43 as an osmosensor that promoted calcium transients in response to hypoosmotic conditions, but not hyperosmotic conditions. GAP43 is a palmitoylated, membrane-associated protein, which has been implicated in axon growth and synaptic plasticity. When expressed in human embryonic kidney 293 (HEK293) cells, GAP43 promoted an increase in cytosolic calcium that was dependent on intracellular calcium stores and not extracellular calcium. Furthermore, both the magnitude of the calcium transient and the number of cells responding increased with increasing hypotonicity. Mutation of the cysteines required for palmitoylation, which allows GAP43 to accumulate in lipid rafts, blocked the ability of GAP43 to activate calcium signaling in response to hypoosmotic conditions. Pharmacological inhibition of protein kinase C, which is known to phosphorylate GAP43, reduced the number of cells that responded to hypotonicity. GAP43 and phospholipase C (PLC)-δ1 coimmunoprecipitated in cells exposed to hypoosmotic conditions and not in cells in isotonic medium. The response to hypoosmotic conditions was completely blocked if PLC activity was inhibited. Thus, hypoosmotic conditions stimulate the association of GAP43 with PLC, activating PLC activity and promoting the release of inositol trisphosphate (IP3), which stimulates release of calcium from internal stores. Indeed, increased IP3 was detectable in GAP43-expressing cells exposed to hypoosmotic stimuli.M. Caprini, A. Gomis, H. Cabedo, R. Planells-Cases, C. Belmonte, F. Viana, A. Ferrer-Montiel, GAP43 stimulates inositol trisphosphate-mediated calcium release in response to hypotonicity. EMBO J. 22, 3004-3014 (2003). [Abstract] [Full Text]