An examination of the role of increased cytosolic free Ca2+ concentrations in the inhibition of mRNA translation

Algis L. Laitusis, Charles O. Brostrom, Alexey G. Ryazanov, Margaret A. Brostrom

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15 Scopus citations

Abstract

Mobilization of Ca2+ sequestered by the endoplasmic reticulum (ER) produces the phosphorylation of initiation factor (eIF) 2, whereas an increase in cytosolic free Ca2+ ([Ca2+](i)) due to plasmalemmal Ca2+ influx increases the phosphorylation of elongation factor (eEF) 2. In nucleated mammalian cells, depletion of ER Ca2+ stores has been demonstrated to inhibit translational initiation, but evidence that increased [Ca2+](i) per se causes slowing of peptide chain elongation is lacking. L- type Ca2+ channel activity of GH3 pituitary cells, which are enriched in calmodulin-dependent eEF-2 kinase, was manipulated such that the impact of [Ca2+](i) on eEF-2 phosphorylation and translational rate could be examined for up to 10 min without inhibiting initiation. At 1 mM extracellular Ca2+, resting [Ca2+](i) values were high (154-255 nM) and eEF-2 was phosphorylated. The Ca2+ channel antagonist, nisoldipine, lowered [Ca2+](i) and reduced eEF-2 phosphorylation by half but had no effect on amino acid incorporation. The Ca2+ channel agonist, Bay K 8644, produced sustained elevations of [Ca2+](i) that were associated with 25-50% increases in eEF-2 phosphorylation, but no changes in protein synthetic rates occurred. Larger Ca2+ influxes were achievable with either 25 mM KCl or KCl plus Bay K 8644. These treatments further increased eEF-2 phosphorylation (50100% above control) and inhibited leucine incorporation by 20-70% but ATP content was reduced by 2550% and total cell-associated Ca2+ contents rose by 3-to 13-fold. elF-2α was not phosphorylated during these treatments. Addition of low concentrations of ionomycin, which do not lower ATP content, was associated with complex changes in [Ca2+](i) that resembled alterations in eEF-2 phosphorylation. The inhibition of leucine incorporation in response to ionomycin, however, coincided only with the phosphorylation of eIF2-α, not eEF-2. It is concluded that changes in [Ca2+](i) occurring in the absence of ATP depletion alter the phosphorylation state of eEF-2 but are not regulatory for mRNA translation.

Original languageEnglish (US)
Pages (from-to)270-280
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume354
Issue number2
DOIs
StatePublished - Jun 15 1998

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

Keywords

  • ATP depletion
  • GH cells
  • L-type Ca channels
  • Translational regulation
  • [Ca](i)
  • eEF-2 phosphorylation

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