STIM1 regulates store-operated Ca2+ entry in oocytes.
The single transmembrane-spanning Ca(2+)-binding protein, STIM1, has been proposed to function as a Ca(2+) sensor that links the endoplasmic reticulum to the activation of store-operated Ca(2+) channels. In this study, the presence, subcellular localization and function of STIM1 in store-operated Ca(2+) entry in oocytes was investigated using the pig as a model. Cloning and sequence analysis revealed the presence of porcine STIM1 with a coding sequence of 2058 bp. In oocytes with full cytoplasmic Ca(2+) stores, STIM1 was localized predominantly in the inner cytoplasm as indicated by immunocytochemistry or overexpression of human STIM1 conjugated to the yellow fluorescent protein. Depletion of the Ca(2+) stores was associated with redistribution of STIM1 along the plasma membrane. Increasing STIM1 expression resulted in enhanced Ca(2+) influx after store depletion and subsequent Ca(2+) add-back; the influx was inhibited when the oocytes were pretreated with lanthanum, a specific inhibitor of store-operated Ca(2+) channels. When STIM1 expression was suppressed using siRNAs, there was no change in cytosolic free Ca(2+) levels in the store-depleted oocytes after Ca(2+) add-back. The findings suggest that in oocytes, STIM1 serves as a sensor of Ca(2+) store content that after store depletion moves to the plasma membrane to stimulate store-operated Ca(2+) entry.
Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606, USA.
Sehwon
S
Koh
Kiho
K
Lee
Chunmin
C
Wang
Ryan A
RA
Cabot
Zoltan
Z
Machaty
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D017931:DNA Primers; D006801:Humans; D007150:Immunohistochemistry; D017136:Ion Transport; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D008845:Microinjections; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism*; D034622:RNA Interference; D017386:Sequence Homology, Amino Acid; D013552:Swine; D014158:Transcription, Genetic
D017931:DNA Primers; D008565:Membrane Proteins; D002118:Calcium