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Effects of prenyl pyrophosphates on the binding of PKCgamma with RACK1

Yu-Hsun Chen1, Han-Chung Wang, Ching-Yu Lin

  • 1Division of Biochemistry and Molecular Science, Institute of Zoology, Academia Sinica, Nankang 11529, Taipei, Taiwan.

Journal of Experimental Zoology. Part A, Comparative Experimental Biology
|December 31, 2002
PubMed

Insights

Receptors for activated C kinase 1 (RACK1) from shrimp bind to PKCgamma, influencing tubulin polymerization. Low levels of prenyl pyrophosphates enhance this RACK1-PKCgamma interaction, suggesting a signaling role in PKC isoform switching.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Cell Signaling

Background:

  • Receptors for activated C kinase (RACKs) are crucial for protein kinase C (PKC) isoenzyme translocation and function.
  • RACK1 mediates isoform-selective PKC functions, impacting cellular processes.
  • Understanding RACK1 interactions is key to deciphering PKC signaling pathways.

Purpose of the Study:

  • To isolate and characterize RACK1 cDNA from the shrimp Penaeus japonicus.
  • To investigate the binding interaction between shrimp RACK1 and PKCgamma.
  • To explore the role of prenyl pyrophosphates in modulating RACK1-PKCgamma binding and function.

Main Methods:

  • Homology cloning was used to isolate RACK1 cDNA from Penaeus japonicus.
  • Recombinant shrimp RACK1 was expressed and characterized using monoclonal antibodies.
  • Biotinylated shrimp RACK1 was used in co-precipitation assays with purified PKCgamma to assess binding in the presence of geranylgeranyl pyrophosphate and farnesyl pyrophosphate.

Main Results:

  • Shrimp RACK1 cDNA encodes a 318-amino acid polypeptide with 91% homology to human G(beta2)-like proteins.
  • Expressed shrimp RACK1 produced a 45-kDa polypeptide reactive to anti-RACK1 antibodies.
  • Low concentrations of geranylgeranyl pyrophosphate and farnesyl pyrophosphate significantly enhanced RACK1-PKCgamma binding, particularly between species.
  • Both shrimp and mouse PKCgamma inhibited taxol-induced tubulin polymerization.

Conclusions:

  • Depletion of prenyl pyrophosphates may signal RACK1 to switch binding from conventional PKC (cPKC) to novel PKC (nPKC) isoenzymes.
  • A hydrophobic binding pocket for geranylgeranyl pyrophosphate in RACK1 was identified through prenylation.
  • Shrimp RACK1 serves as a functional homolog of mammalian RACK1, participating in PKC-mediated cellular processes.

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