Non-stimulated, agonist-stimulated and store-operated Ca2+ influx in MDA-MB-468 breast cancer cells and the effect of EGF-induced EMT on calcium entry

PLoS One. 2012;7(5):e36923. doi: 10.1371/journal.pone.0036923. Epub 2012 May 30.

Abstract

In addition to their well-defined roles in replenishing depleted endoplasmic reticulum (ER) Ca(2+) reserves, molecular components of the store-operated Ca(2+) entry pathway regulate breast cancer metastasis. A process implicated in cancer metastasis that describes the conversion to a more invasive phenotype is epithelial-mesenchymal transition (EMT). In this study we show that EGF-induced EMT in MDA-MB-468 breast cancer cells is associated with a reduction in agonist-stimulated and store-operated Ca(2+) influx, and that MDA-MB-468 cells prior to EMT induction have a high level of non-stimulated Ca(2+) influx. The potential roles for specific Ca(2+) channels in these pathways were assessed by siRNA-mediated silencing of ORAI1 and transient receptor potential canonical type 1 (TRPC1) channels in MDA-MB-468 breast cancer cells. Non-stimulated, agonist-stimulated and store-operated Ca(2+) influx were significantly inhibited with ORAI1 silencing. TRPC1 knockdown attenuated non-stimulated Ca(2+) influx in a manner dependent on Ca(2+) influx via ORAI1. TRPC1 silencing was also associated with reduced ERK1/2 phosphorylation and changes in the rate of Ca(2+) release from the ER associated with the inhibition of the sarco/endoplasmic reticulum Ca(2+)-ATPase (time to peak [Ca(2+)](CYT) = 188.7 ± 34.6 s (TRPC1 siRNA) versus 124.0 ± 9.5 s (non-targeting siRNA); P<0.05). These studies indicate that EMT in MDA-MB-468 breast cancer cells is associated with a pronounced remodeling of Ca(2+) influx, which may be due to altered ORAI1 and/or TRPC1 channel function. Our findings also suggest that TRPC1 channels in MDA-MB-468 cells contribute to ORAI1-mediated Ca(2+) influx in non-stimulated cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biological Transport / drug effects
  • Breast Neoplasms / pathology*
  • Calcium / metabolism*
  • Calcium Channels / deficiency
  • Calcium Channels / genetics
  • Calcium Signaling / drug effects*
  • Calcium-Transporting ATPases / antagonists & inhibitors
  • Calcium-Transporting ATPases / metabolism
  • Cell Line, Tumor
  • Epidermal Growth Factor / pharmacology*
  • Epithelial-Mesenchymal Transition / drug effects*
  • Gene Silencing
  • Humans
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Mitogen-Activated Protein Kinase 3 / metabolism
  • ORAI1 Protein
  • Purinergic Agonists / pharmacology*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • S Phase / drug effects
  • S Phase / genetics
  • TRPC Cation Channels / deficiency
  • TRPC Cation Channels / genetics

Substances

  • Calcium Channels
  • ORAI1 Protein
  • ORAI1 protein, human
  • Purinergic Agonists
  • RNA, Messenger
  • TRPC Cation Channels
  • transient receptor potential cation channel, subfamily C, member 1
  • Epidermal Growth Factor
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • Calcium-Transporting ATPases
  • Calcium