STIM1 phosphorylation triggered by epidermal growth factor mediates cell migration

• EGF triggers the phosphorylation of STIM1 at ERK1/2 target sites.
• Activation of Ras proteins leads to phosphorylation of STIM1 at ERK1/2 target sites.
• EGF induces STIM1–EB1 dissociation.
• Phosphorylation of STIM1 is a downstream effector of the EGF signaling pathway.
• Impairment of STIM1 phosphorylation reduces cell migration and EMT.

STIM1 is a key regulator of store-operated calcium entry (SOCE), and therefore a mediator of Ca2 + entry-dependent cellular events. Phosphorylation of STIM1 at ERK1/2 target sites has been described as enhancing STIM1 activation during intracellular Ca2 + emptying triggered by the inhibition of the sarco(endo)plasmic Ca2 +-ATPase with thapsigargin. However, no physiological function is known for this specific phosphorylation. The present study examined the role of STIM1 phosphorylation in cell signaling triggered by EGF. Using a human endometrial adenocarcinoma cell line (Ishikawa cells) EGF or H-Ras(G12V), an active mutant of H-Ras, was found to trigger STIM1 phosphorylation at residues Ser575, Ser608, and Ser621, and this process was sensitive to PD0325901, an inhibitor of ERK1/2. Both, ERK1/2 activation and STIM1 phosphorylation took place in the absence of extracellular Ca2 +, indicating that both events are upstream steps for Ca2 + entry activation. Also, EGF triggered the dissociation of STIM1 from EB1 (a regulator of microtubule plus-ends) in a manner similar to that reported for the activation of STIM1 by thapsigargin. Migration of the Ishikawa cells was impaired when STIM1 phosphorylation was targeted by Ser-to-Ala substitution mutation of ERK1/2 target sites. This effect was also observed with the Ca2 + channel blocker SKF96365. Phosphomimetic mutation of STIM1 restored the migration to levels similar to that found for STIM1-wild type. Finally, the increased vimentin expression and relocalization of E-cadherin triggered by EGF were largely inhibited by targeting STIM1 phosphorylation, while STIM1-S575E/S608E/S621E normalized the profiles of these two EMT markers.