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Spatio-Temporal Manipulation of Small GTPase Activity at Subcellular Level and on Timescale of Seconds in Living Cells
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Novel functional interaction between the plasma membrane Ca2+ pump 4b and the proapoptotic tumor suppressor

Angel L Armesilla1, Judith C Williams, Mamta H Buch

  • 1Division of Cardiology, University of Manchester, Manchester M13 9PT, United Kingdom.

The Journal of Biological Chemistry
|May 18, 2004
PubMed

Insights

Plasma membrane calcium ATPases (PMCAs) interact with tumor suppressor RASSF1, modulating Ras signaling. This novel interaction, distinct from the C-terminus, reveals PMCA4b

Area of Science:

  • Cellular Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Plasma membrane calcium ATPases (PMCAs) are key in calcium extrusion.
  • Previous studies identified PMCA interactions with PDZ proteins at the C-terminus, suggesting roles in signal transduction.
  • The existence of other intracellular regions prompted investigation into additional protein interactions.

Purpose of the Study:

  • To identify novel protein interaction partners for domains of PMCA other than the C-terminus.
  • To investigate the functional consequences of PMCA4b interaction with the tumor suppressor RASSF1.
  • To elucidate the role of PMCA4b in organizing macromolecular protein complexes and modulating signaling pathways.

Main Methods:

  • Two-hybrid screen using human PMCA4b (residues 652-840) against a human fetal heart cDNA library.
  • Immunofluorescence co-localization, immunoprecipitation, and glutathione S-transferase pull-down assays.
  • Functional assays involving co-expression of PMCA4b and RASSF1, and inhibition studies using a RASSF1 peptide.

Main Results:

  • A novel interaction was discovered between human PMCA4b and the tumor suppressor RASSF1.
  • The interaction domains were mapped to specific regions within PMCA4b (652-748) and RASSF1 (74-123 for RASSF1C, 144-193 for RASSF1A).
  • Co-expression of PMCA4b and RASSF1 inhibited epidermal growth factor-induced Erk pathway activation, an effect reversible by disrupting their association.

Conclusions:

  • PMCA4b interacts with RASSF1 via a domain distinct from its C-terminus.
  • PMCA4b functions as an organizer of macromolecular complexes, recruiting diverse proteins through different domains.
  • The interaction with RASSF1 highlights a role for PMCA4b in modulating Ras-mediated signaling pathways, including H-Ras-induced apoptosis.

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