Synergistic stabilization by nitrosoglutathione-induced thiol modifications in the stromal interaction molecule-2 luminal domain suppresses basal and store operated calcium entry

Sci Rep. 2020 Jun 23;10(1):10177. doi: 10.1038/s41598-020-66961-3.

Abstract

Stromal interaction molecule-1 and -2 (STIM1/2) are endoplasmic reticulum (ER) membrane-inserted calcium (Ca2+) sensing proteins that, together with Orai1-composed Ca2+ channels on the plasma membrane (PM), regulate intracellular Ca2+ levels. Recent evidence suggests that S-nitrosylation of the luminal STIM1 Cys residues inhibits store operated Ca2+ entry (SOCE). However, the effects of thiol modifications on STIM2 during nitrosative stress and their role in regulating basal Ca2+ levels remain unknown. Here, we demonstrate that the nitric oxide (NO) donor nitrosoglutathione (GSNO) thermodynamically stabilizes the STIM2 Ca2+ sensing region in a Cys-specific manner. We uncovered a remarkable synergism in this stabilization involving the three luminal Cys of STIM2, which is unique to this paralog. S-Nitrosylation causes structural perturbations that converge on the face of the EF-hand and sterile α motif (EF-SAM) domain, implicated in unfolding-coupled activation. In HEK293T cells, enhanced free basal cytosolic Ca2+ and SOCE mediated by STIM2 overexpression could be attenuated by GSNO or mutation of the modifiable Cys located in the luminal domain. Collectively, we identify the Cys residues within the N-terminal region of STIM2 as modifiable targets during nitrosative stress that can profoundly and cooperatively affect basal Ca2+ and SOCE regulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calcium / metabolism*
  • Calcium Signaling / physiology
  • Cell Line
  • Cell Membrane / metabolism*
  • Endoplasmic Reticulum / metabolism
  • Glutathione / metabolism*
  • HEK293 Cells
  • Humans
  • Membrane Proteins / metabolism
  • ORAI1 Protein / metabolism
  • Protein Binding / physiology
  • Stromal Interaction Molecule 1 / metabolism
  • Stromal Interaction Molecule 2 / metabolism*
  • Sulfhydryl Compounds / metabolism*

Substances

  • Membrane Proteins
  • ORAI1 Protein
  • STIM2 protein, human
  • Stromal Interaction Molecule 1
  • Stromal Interaction Molecule 2
  • Sulfhydryl Compounds
  • Glutathione
  • Calcium