PMID- 26351694
OWN - NLM
STAT- MEDLINE
DCOM- 20160412
LR  - 20191210
IS  - 1091-6490 (Electronic)
IS  - 0027-8424 (Print)
IS  - 0027-8424 (Linking)
VI  - 112
IP  - 40
DP  - 2015 Oct 6
TI  - Nanoscale patterning of STIM1 and Orai1 during store-operated Ca2+ entry.
PG  - E5533-42
LID - 10.1073/pnas.1515606112 [doi]
AB  - Stromal interacting molecule (STIM) and Orai proteins constitute the core 
      machinery of store-operated calcium entry. We used transmission and 
      freeze-fracture electron microscopy to visualize STIM1 and Orai1 at endoplasmic 
      reticulum (ER)-plasma membrane (PM) junctions in HEK 293 cells. Compared with 
      control cells, thin sections of STIM1-transfected cells possessed far more ER 
      elements, which took the form of complex stackable cisternae and labyrinthine 
      structures adjoining the PM at junctional couplings (JCs). JC formation required 
      STIM1 expression but not store depletion, induced here by thapsigargin (TG). 
      Extended molecules, indicative of STIM1, decorated the cytoplasmic surface of ER, 
      bridged a 12-nm ER-PM gap, and showed clear rearrangement into small clusters 
      following TG treatment. Freeze-fracture replicas of the PM of Orai1-transfected 
      cells showed extensive domains packed with characteristic "particles"; TG 
      treatment led to aggregation of these particles into sharply delimited "puncta" 
      positioned upon raised membrane subdomains. The size and spacing of Orai1 
      channels were consistent with the Orai crystal structure, and stoichiometry was 
      unchanged by store depletion, coexpression with STIM1, or an Orai1 mutation 
      (L273D) affecting STIM1 association. Although the arrangement of Orai1 channels 
      in puncta was substantially unstructured, a portion of channels were spaced at 
       approximately 15 nm. Monte Carlo analysis supported a nonrandom distribution for a portion of 
      channels spaced at  approximately 15 nm. These images offer dramatic, direct views of STIM1 
      aggregation and Orai1 clustering in store-depleted cells and provide evidence for 
      the interaction of a single Orai1 channel with small clusters of STIM1 molecules.
FAU - Perni, Stefano
AU  - Perni S
AD  - Department of Cell and Developmental Biology, University of Pennsylvania, 
      Philadelphia, PA 19104;
FAU - Dynes, Joseph L
AU  - Dynes JL
AD  - Department of Physiology and Biophysics, University of California, Irvine, CA 
      92697;
FAU - Yeromin, Andriy V
AU  - Yeromin AV
AD  - Department of Physiology and Biophysics, University of California, Irvine, CA 
      92697;
FAU - Cahalan, Michael D
AU  - Cahalan MD
AD  - Department of Physiology and Biophysics, University of California, Irvine, CA 
      92697; Institute for Immunology, University of California, Irvine, CA 92697.
FAU - Franzini-Armstrong, Clara
AU  - Franzini-Armstrong C
AD  - Department of Cell and Developmental Biology, University of Pennsylvania, 
      Philadelphia, PA 19104; armstroc@mail.med.upenn.edu.
LA  - eng
GR  - R01 HL48093/HL/NHLBI NIH HHS/United States
GR  - R01 HL048093/HL/NHLBI NIH HHS/United States
GR  - R37 NS014609/NS/NINDS NIH HHS/United States
GR  - P01 AR052354/AR/NIAMS NIH HHS/United States
GR  - 2P01 AR052354/AR/NIAMS NIH HHS/United States
GR  - NS14609/NS/NINDS NIH HHS/United States
GR  - R01 NS014609/NS/NINDS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
DEP - 20150908
PL  - United States
TA  - Proc Natl Acad Sci U S A
JT  - Proceedings of the National Academy of Sciences of the United States of America
JID - 7505876
RN  - 0 (Calcium Channels)
RN  - 0 (Enzyme Inhibitors)
RN  - 0 (Luminescent Proteins)
RN  - 0 (Membrane Proteins)
RN  - 0 (Neoplasm Proteins)
RN  - 0 (ORAI1 Protein)
RN  - 0 (ORAI1 protein, human)
RN  - 0 (STIM1 protein, human)
RN  - 0 (Stromal Interaction Molecule 1)
RN  - 67526-95-8 (Thapsigargin)
RN  - EC 7.2.2.10 (Calcium-Transporting ATPases)
RN  - SY7Q814VUP (Calcium)
SB  - IM
MH  - Calcium/*metabolism
MH  - Calcium Channels/genetics/*metabolism
MH  - Calcium-Transporting ATPases/antagonists & inhibitors/metabolism
MH  - Cell Membrane/metabolism/ultrastructure
MH  - Endoplasmic Reticulum/metabolism/ultrastructure
MH  - Enzyme Inhibitors/pharmacology
MH  - Freeze Fracturing
MH  - HEK293 Cells
MH  - Humans
MH  - Luminescent Proteins/genetics/metabolism
MH  - Membrane Proteins/genetics/*metabolism
MH  - Microscopy, Electron
MH  - Microscopy, Fluorescence/methods
MH  - Monte Carlo Method
MH  - Mutation
MH  - Neoplasm Proteins/genetics/*metabolism
MH  - ORAI1 Protein
MH  - Protein Binding
MH  - Protein Transport/drug effects
MH  - Stromal Interaction Molecule 1
MH  - Thapsigargin/pharmacology
MH  - Videotape Recording
PMC - PMC4603497
OTO - NOTNLM
OT  - Orai1
OT  - SOCE
OT  - STIM1
OT  - electron microscopy
OT  - nanoscale patterning
COIS- The authors declare no conflict of interest.
EDAT- 2015/09/10 06:00
MHDA- 2016/04/14 06:00
PMCR- 2016/04/06
CRDT- 2015/09/10 06:00
PHST- 2015/09/10 06:00 [entrez]
PHST- 2015/09/10 06:00 [pubmed]
PHST- 2016/04/14 06:00 [medline]
PHST- 2016/04/06 00:00 [pmc-release]
AID - 1515606112 [pii]
AID - 201515606 [pii]
AID - 10.1073/pnas.1515606112 [doi]
PST - ppublish
SO  - Proc Natl Acad Sci U S A. 2015 Oct 6;112(40):E5533-42. doi: 
      10.1073/pnas.1515606112. Epub 2015 Sep 8.