The three isoenzymes of human inositol-1,4,5-trisphosphate 3-kinase show specific intracellular localization but comparable Ca2+ responses on transfection in COS-7 cells

Biochem J. 2003 Aug 15;374(Pt 1):41-9. doi: 10.1042/BJ20021963.


Inositol 1,4,5-trisphosphate [Ins(1,4,5) P3] 3-kinase catalyses the phosphorylation of InsP3 to inositol 1,3,4,5-tetrakisphosphate. cDNAs encoding three human isoenzymes of InsP3 3-kinase (A, B and C) have been reported previously [Choi, Kim, Lee, Moon, Sim, Kim, Chung and Rhee (1990) Science 248, 64-66; Dewaste, Pouillon, Moreau, Shears, Takazawa and Erneux (2000) Biochem. J. 352, 343-351; Dewaste, Roymans, Moreau and Erneux (2002) Biochem. Biophys. Res. Commun. 291, 400-405; Takazawa, Perret, Dumont and Erneux (1991) Biochem. Biophys. Res. Commun. 174, 529-535]. The localization of InsP3 3-kinase isoenzymes fused at their N-terminus to the green fluorescent protein has been studied by confocal microscopy. The A isoform appeared to associate with the cytoskeleton, whereas the C isoform was totally cytoplasmic. The B isoform had a more complex localization: it appeared in the plasma membrane, cytoskeleton and in the endoplasmic reticulum. The three human isoenzymes of InsP3 3-kinase can thus be distinguished by their N-terminal sequence, sensitivity to Ca2+/calmodulin and localization on transfection in COS-7 cells. We have compared the cytosolic Ca2+ responses induced by ATP in COS-7 cells transfected with the three isoenzymes. Cells expressing high levels of any of the three isoforms no longer respond to ATP, whereas cells expressing low levels of each enzyme showed a reduced response consisting of one to three Ca2+ spikes in response to 100 microM ATP. These effects were seen only in wild-type InsP3 3-kinase-transfected cells. 3-Kinase-dead mutant cells behaved as vector-transfected cells. The results highlight the potential role of the three isoforms of InsP3 3-kinase as direct InsP3 metabolizing enzymes and direct regulators of Ca2+ responses to extracellular signals.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • CHO Cells
  • COS Cells
  • Calcium / metabolism*
  • Cell Membrane / enzymology
  • Chlorocebus aethiops
  • Cricetinae
  • Cytoskeleton / enzymology*
  • DNA Primers
  • Endoplasmic Reticulum / enzymology*
  • Green Fluorescent Proteins
  • Humans
  • Isoenzymes / genetics*
  • Isoenzymes / metabolism*
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Mutagenesis, Site-Directed
  • Phosphotransferases (Alcohol Group Acceptor) / genetics*
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Subcellular Fractions / enzymology
  • Transfection


  • DNA Primers
  • Isoenzymes
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins
  • Phosphotransferases (Alcohol Group Acceptor)
  • Inositol 1,4,5-trisphosphate 3-kinase
  • Calcium