Molecular Cloning and Characterization of a Lysophosphatidic Acid Receptor, Edg-7, Expressed in Prostate

  1. Dong-Soon Im1,
  2. Christopher E. Heise1,
  3. Michael A. Harding2,
  4. Susan R. George3,
  5. Brian F. O'Dowd3,
  6. Dan Theodorescu2 and
  7. Kevin R. Lynch1
  1. Departments of 1Pharmacology (D.-S.I., C.E.H., K.R.L.) and 2Urology and Molecular Physiology and Biological Physics (M.A.H., D.T.), University of Virginia School of Medicine, Charlottesville, Virginia; and 3Department of Pharmacology (S.R.G., B.F.O.), University of Toronto, Toronto, Ontario, Canada

    Abstract

    Two G protein-coupled receptors (Edg-2) and (Edg-4) for the lysolipid phosphoric acid mediator lysophosphatidic acid have been described by molecular cloning. However, the calcium-mobilizing receptor Edg-4 is not expressed in some cell lines that exhibit robust calcium responses to this ligand, thus predicting the existence of additional receptor subtypes. We report here on the characterization of a third human lysophosphatidic acid receptor subtype, Edg-7, which mediates lysophosphatidic acid-evoked calcium mobilization. In a rat hepatoma Rh7777 cell line that lacks endogenous responses to lysophosphatidic acid, this lipid mediator, but not others, evokes calcium transients when the cells have been transfected with Edg-7 or Edg-4 DNAs. Furthermore, frog oocytes exhibit a calcium-mediated chloride conductance in response to mammalian-selective lysophosphatidic acid mimetics after injection of Edg-7 mRNA. Edg-7-expressing Rh7777 cells do not show inhibition of forskolin-driven rises in cAMP in response to lysophosphatidic acid. However, membranes from HEK293T cells cotransfected with Edg-7 and Gi2α protein DNAs show lysophosphatidic acid dose-dependent increases in [γ-35S]GTP binding with an EC50 value of 195 nM. When we used this assay to compare various synthetic LPA analogs at Edg-2, Edg-4, and Edg-7 receptors, we found that ethanolamine-based compounds, which are full LPA mimetics at Edg-2 and Edg-4, exhibit little activity at the Edg-7 receptor. Edg-7 RNA was detected in extracts of several rat and human tissues including prostate. Together, our data indicate that Edg-7 is a third lysophosphatidic acid receptor that couples predominantly to Gq/11α proteins.

    Footnotes

    • Send reprint requests to: Dr. Kevin R. Lynch, Department of Pharmacology, University of Virginia Health System, Box 800735, 1300 Jefferson Park Ave., Charlottesville, VA. E-mail: KRL2Z{at}virginia.edu

    • This work was supported by National Institutes of Health Research Grants R01 GM52722 and R21 CA69848. C.E.H. is supported by a National Research Service Award predoctoral traineeship (T32 GM07055).

    • Abbreviations:
      LPA
      1-oleoyl lysophosphatidic acid
      S1P
      sphingosine 1-phosphate
      PTX
      pertussis toxin
      RT-PCR
      reverse transcription-polymerase chain reaction
      RACE
      rapid amplification of cDNA ends
      ORF
      open reading frame
      EST
      expressed sequence tag
      GPCR
      G protein-coupled receptor
      NOEPA
      N-oleoyl ethanolamide phosphoric acid
      NPSPA
      N-palmitoyl serine phosphoric acid
      NAEPA
      N-acyl ethanolamide phosphoric acid
      • Received November 22, 1999.
      • Accepted December 29, 1999.
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    1. Molecular Pharmacology April 1, 2000 vol. 57 no. 4 753-759
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