Isolation, characterization, and localization of the inositol 1,4,5-trisphosphate receptor protein in Xenopus laevis oocytes

Jan B. Parys, Suzanne W. Sernett, Sylvain DeLisle, Peter M. Snyder, Michael J. Welsh, Kevin P. Campbell

Research output: Contribution to journalArticle

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Abstract

Inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) induces Ca2+ oscillations and waves in Xenopus laevis oocytes. Microsomes from oocytes exhibit high-affinity binding for Ins(1,4,5)P3, and demonstrate Ins(1,4,5)P3-induced Ca2+ release. The Ins(1,4,5)P3 receptor (InsP3R) was purified from oocyte microsomes as a large tetrameric complex and shown to have a monomer molecular mass of 256 kDa, compared with 273 kDa for the brain InsP3R. Binding to the oocyte receptor is highly specific for Ins(1,4,5)P3 and is inhibited by heparin (IC50, 2 μg/ml). Immunoblot analysis revealed that an antibody against the C-terminal sequence of the brain receptor recognized the oocyte receptor. These results, in addition to the difference in pattern obtained after limited proteolysis, suggest that the oocyte InsP3R is a new shorter isoform of the mammalian brain type I InsP3R. Immunofluorescence experiments indicated the presence of the InsP3R in the cortical layer and the perinuclear endoplasmic reticulum of the oocyte. However, immunological and biochemical experiments did not reveal the presence of the ryanodine receptor. The presence of an InsP3R and the absence of a ryanodine receptor support the importance of Ins(1,4,5)P3 in Ca2+ handling by oocytes and particularly in the induction of Ca2+ oscillations and waves.

Original languageEnglish (US)
Pages (from-to)18776-18782
Number of pages7
JournalJournal of Biological Chemistry
Volume267
Issue number26
StatePublished - Sep 15 1992

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Xenopus Proteins
Inositol 1,4,5-Trisphosphate Receptors
Oocytes
Brain
Ryanodine Receptor Calcium Release Channel
Proteolysis
Proteins
Inositol 1,4,5-Trisphosphate
Molecular mass
Heparin
Protein Isoforms
Microsomes
Monomers
Experiments
Antibodies
Xenopus laevis
Endoplasmic Reticulum
Inhibitory Concentration 50
Fluorescent Antibody Technique

ASJC Scopus subject areas

  • Biochemistry

Cite this

Isolation, characterization, and localization of the inositol 1,4,5-trisphosphate receptor protein in Xenopus laevis oocytes. / Parys, Jan B.; Sernett, Suzanne W.; DeLisle, Sylvain; Snyder, Peter M.; Welsh, Michael J.; Campbell, Kevin P.

In: Journal of Biological Chemistry, Vol. 267, No. 26, 15.09.1992, p. 18776-18782.

Research output: Contribution to journalArticle

Parys, Jan B. ; Sernett, Suzanne W. ; DeLisle, Sylvain ; Snyder, Peter M. ; Welsh, Michael J. ; Campbell, Kevin P. / Isolation, characterization, and localization of the inositol 1,4,5-trisphosphate receptor protein in Xenopus laevis oocytes. In: Journal of Biological Chemistry. 1992 ; Vol. 267, No. 26. pp. 18776-18782.
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abstract = "Inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) induces Ca2+ oscillations and waves in Xenopus laevis oocytes. Microsomes from oocytes exhibit high-affinity binding for Ins(1,4,5)P3, and demonstrate Ins(1,4,5)P3-induced Ca2+ release. The Ins(1,4,5)P3 receptor (InsP3R) was purified from oocyte microsomes as a large tetrameric complex and shown to have a monomer molecular mass of 256 kDa, compared with 273 kDa for the brain InsP3R. Binding to the oocyte receptor is highly specific for Ins(1,4,5)P3 and is inhibited by heparin (IC50, 2 μg/ml). Immunoblot analysis revealed that an antibody against the C-terminal sequence of the brain receptor recognized the oocyte receptor. These results, in addition to the difference in pattern obtained after limited proteolysis, suggest that the oocyte InsP3R is a new shorter isoform of the mammalian brain type I InsP3R. Immunofluorescence experiments indicated the presence of the InsP3R in the cortical layer and the perinuclear endoplasmic reticulum of the oocyte. However, immunological and biochemical experiments did not reveal the presence of the ryanodine receptor. The presence of an InsP3R and the absence of a ryanodine receptor support the importance of Ins(1,4,5)P3 in Ca2+ handling by oocytes and particularly in the induction of Ca2+ oscillations and waves.",
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