Proteasomal degradation of eukaryotic elongation factor-2 kinase (EF2K) is regulated by cAMP-PKA signaling and the SCFβTRCP ubiquitin E3 ligase

J Biol Chem. 2013 Jun 14;288(24):17803-11. doi: 10.1074/jbc.M113.477182. Epub 2013 May 2.

Abstract

Protein translation and degradation are critical for proper protein homeostasis, yet it remains unclear how these processes are dynamically regulated, or how they may directly balance or synergize with each other. An important translational control mechanism is the Ca(2+)/calmodulin-dependent phosphorylation of eukaryotic elongation factor-2 (eEF-2) by eukaryotic elongation factor-2 kinase (EF2K), which inhibits elongation of nascent polypeptide chains during translation. We previously described a reduction of EF2K activity in PC12 cells treated with NGF or forskolin. Here, we show that both forskolin- and IGF-1-mediated reductions of EF2K activity in PC12 cells are due to decreased EF2K protein levels, and this is attenuated by application of the proteasome inhibitor, MG132. We further demonstrate that proteasome-mediated degradation of EF2K occurs in response to A2A-type adenosine receptor stimulation, and that activation of protein kinase A (PKA) or phospho-mimetic mutation of the previously characterized PKA site, Ser-499, were sufficient to induce EF2K turnover in PC12 cells. A similar EF2K degradation mechanism was observed in primary neurons and HEK cells. Expression of a dominant-negative form of Cul1 in HEK cells demonstrated that EF2K levels are regulated by an SCF-type ubiquitin E3 ligase. Specifically, EF2K binds to the F-box proteins, βTRCP1 and βTRCP2, and βTRCP regulates EF2K levels and polyubiquitylation. We propose that the proteasomal degradation of EF2K provides a mechanistic link between activity-dependent protein synthesis and degradation.

Keywords: Cyclic AMP (cAMP); E3 Ubiquitin Ligase; Proteasome; Translation Control; Translation Elongation Factors; eEF-2; eEF-2 Kinase; βTRCP.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine / analogs & derivatives
  • Adenosine / pharmacology
  • Adenosine A2 Receptor Agonists / pharmacology
  • Animals
  • Brain-Derived Neurotrophic Factor / pharmacology
  • Brain-Derived Neurotrophic Factor / physiology
  • Colforsin / pharmacology
  • Cullin Proteins / metabolism
  • Cyclic AMP / metabolism*
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Elongation Factor 2 Kinase / metabolism*
  • HEK293 Cells
  • Humans
  • Insulin-Like Growth Factor I / pharmacology
  • Insulin-Like Growth Factor I / physiology
  • Neurons / drug effects
  • Neurons / metabolism
  • PC12 Cells
  • Phenethylamines / pharmacology
  • Primary Cell Culture
  • Proteasome Endopeptidase Complex
  • Protein Binding
  • Protein Stability
  • Proteolysis
  • Rats
  • SKP Cullin F-Box Protein Ligases / metabolism*
  • Second Messenger Systems*
  • Ubiquitination

Substances

  • Adenosine A2 Receptor Agonists
  • Brain-Derived Neurotrophic Factor
  • Cullin 1
  • Cullin Proteins
  • Phenethylamines
  • 2-(4-(2-carboxyethyl)phenethylamino)-5'-N-ethylcarboxamidoadenosine
  • Colforsin
  • Insulin-Like Growth Factor I
  • Cyclic AMP
  • SKP Cullin F-Box Protein Ligases
  • Eef2k protein, rat
  • Cyclic AMP-Dependent Protein Kinases
  • Elongation Factor 2 Kinase
  • Proteasome Endopeptidase Complex
  • Adenosine