Molecular cloning of mouse type 2 and type 3 inositol 1,4,5-trisphosphate receptors and identification of a novel type 2 receptor splice variant

J Biol Chem. 2005 Mar 18;280(11):10305-17. doi: 10.1074/jbc.M413824200. Epub 2005 Jan 4.

Abstract

We isolated cDNAs encoding type 2 and type 3 inositol 1,4,5-trisphosphate (IP(3)) receptors (IP(3)R2 and IP(3)R3, respectively) from mouse lung and found a novel alternative splicing segment, SI(m2), at 176-208 of IP(3)R2. The long form (IP(3)R2 SI(m2)(+)) was dominant, but the short form (IP(3)R2 SI(m2)(-)) was detected in all tissues examined. IP(3)R2 SI(m2)(-) has neither IP(3) binding activity nor Ca(2+) releasing activity. In addition to its reticular distribution, IP(3)R2 SI(m2)(+) is present in the form of clusters in the endoplasmic reticulum of resting COS-7 cells, and after ATP or Ca(2+) ionophore stimulation, most of the IP(3)R2 SI(m2)(+) is in clusters. IP(3)R3 is localized uniformly on the endoplasmic reticulum of resting cells and forms clusters after ATP or Ca(2+) ionophore stimulation. IP(3)R2 SI(m2)(-) does not form clusters in either resting or stimulated cells. IP(3) binding-deficient site-directed mutants of IP(3)R2 SI(m2)(+) and IP(3)R3 fail to form clusters, indicating that IP(3) binding is involved in the cluster formation by these isoforms. Coexpression of IP(3)R2 SI(m2)(-) prevents stimulus-induced IP(3)R clustering, suggesting that IP(3)R2 SI(m2)(-) functions as a negative coordinator of stimulus-induced IP(3)R clustering. Expression of IP(3)R2 SI(m2)(-) in CHO-K1 cells significantly reduced ATP-induced Ca(2+) entry, but not Ca(2+) release, suggesting that the novel splice variant of IP(3)R2 specifically influences the dynamics of the sustained phase of Ca(2+) signals.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / chemistry
  • Alternative Splicing*
  • Amino Acid Sequence
  • Animals
  • Blotting, Western
  • CHO Cells
  • COS Cells
  • Calcium / metabolism
  • Calcium Channels / chemistry
  • Calcium Channels / genetics*
  • Cell Line
  • Cloning, Molecular
  • Cricetinae
  • Cytoplasm / metabolism
  • DNA, Complementary / metabolism
  • Endoplasmic Reticulum / metabolism
  • Green Fluorescent Proteins / metabolism
  • Immunoprecipitation
  • Inositol 1,4,5-Trisphosphate Receptors
  • Insecta
  • Ionophores / pharmacology
  • Kinetics
  • Lung / metabolism
  • Mice
  • Mice, Inbred C57BL
  • Microscopy, Fluorescence
  • Microsomes / metabolism
  • Molecular Sequence Data
  • Multigene Family
  • Mutagenesis, Site-Directed
  • Rats
  • Receptors, Cytoplasmic and Nuclear / chemistry
  • Receptors, Cytoplasmic and Nuclear / genetics*
  • Recombinant Proteins / chemistry
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Homology, Amino Acid
  • Time Factors
  • Tissue Distribution

Substances

  • Calcium Channels
  • DNA, Complementary
  • Inositol 1,4,5-Trisphosphate Receptors
  • Ionophores
  • Receptors, Cytoplasmic and Nuclear
  • Recombinant Proteins
  • Green Fluorescent Proteins
  • Adenosine Triphosphate
  • Calcium

Associated data

  • GENBANK/AB182288
  • GENBANK/AB182289
  • GENBANK/AB182290