Citation

  • Authors: Ramesh G. et al.
  • Year: 2021
  • Journal: Cell Rep 34(11)
  • Applications: in vitro / DNA / jetOPTIMUS, jetPRIME
  • Cell types:
    1. Name: HEK-293
      Description: Human embryonic kidney Fibroblast
      Known as: HEK293, 293
    2. Name: MEF
      Description: Murine embryonic fibroblast cells 
    3. Name: SH-SY5Y
      Description: Human neuroblastoma cells
      Known as:

Method

For Bimolecular fluorescence complementation, HEK cells were transfected using jetOPTIMUS: 1.2x 10^5 HEK STIM1−/−;STIM2−/− cells were seeded in 6 well plates and transfected with a total amount of 2 μg DNA. For Fura2-based Ca2+ Imaging, jetPRIME was used: - MEF cells were seeded on 25mm coverslips 24 hours before transfection with 2μg of plasmids - SH-SY5Y cells were seeded in 35mm cell culture dishes 24 hours before transfection with 2μg of plasmids. Transfected SH-SY5Y were reseeded 4 hours post transfection on 25mm coverslips.

Abstract

Store-operated Ca2+-entry (SOCE) regulates basal and receptor-triggered Ca2+ signaling with STIM proteins sensing the endoplasmic reticulum (ER) Ca2+ content and triggering Ca2+ entry by gating Orai channels. Although crucial for immune cells, STIM1's role in neuronal Ca2+ homeostasis is controversial. Here, we characterize a splice variant, STIM1B, which shows exclusive neuronal expression and protein content surpassing conventional STIM1 in cerebellum and of significant abundance in other brain regions. STIM1B expression results in a truncated protein with slower kinetics of ER-plasma membrane (PM) cluster formation and ICRAC, as well as reduced inactivation. In primary wild-type neurons, STIM1B is targeted by its spliced-in domain B to presynaptic sites where it converts classic synaptic depression into Ca2+- and Orai-dependent short-term synaptic enhancement (STE) at high-frequency stimulation (HFS). In conjunction with altered STIM1 splicing in human Alzheimer disease, our findings highlight STIM1 splicing as an important regulator of neuronal calcium homeostasis and of synaptic plasticity.

Pubmed