Inhibition of calcium/calmodulin-dependent protein kinase kinase by protein 14-3-3

Monika Davare, Takeo Saneyoshi, Eric S. Guire, Sean C. Nygaard, Thomas Soderling

Research output: Contribution to journalArticle

41 Citations (Scopus)

Abstract

Intracellular calcium concentrations regulate diverse cellular events including cytoskeletal dynamics, gene transcription, and synaptic plasticity. The calcium signal is transduced in part by the calcium/calmodulin-dependent protein kinase (CaMK) cascade that is comprised of CaMK kinase (CaMKK) and its primary downstream substrates, CaMKI and CaMKIV. The CaMK cascade also participates in cross-talk with other signaling pathways: CaMKK/CaMKI can activate the mitogen-activated protein kinase pathway and cAMP-dependent protein kinase (PKA) can directly phosphorylate two inhibitory sites (Thr108 and Ser458) in CaMKK. Here we report an additional PKA-dependent regulation of CaMKK through its interaction with protein 14-3-3. CaMKK and 14-3-3 co-immunoprecipitated from co-transfected heterologous cells as well as from rat brain homogenate, and site-directed mutagenesis studies identified phospho-Ser74 in CaMKK as the primary 14-3-3 binding site. In cultured rat hippocampal neurons and acute hippocampal slices this interaction was robustly stimulated by activation of PKA through forskoli treatment and was blocked by inhibition of PKA. Interaction of 14-3-3 with CaMKK had two regulatory consequences in vitro. It directly inhibited CaMKK activity, and it also blocked dephosphorylation of Thr108, an inhibitory PKA phosphorylation site. In human embryonic kidney 293 cells transfected with CaMKK and stimulated with forskolin, co-transfection with 14-3-3 prevented dephosphorylation of Thr108 to the same extent as did inhibition of protein phosphatases with okadaic acid. We conclude that binding of 14-3-3 to CaMKK stabilizes its inhibition by PKA-mediated phosphorylation, which may have important consequences in the regulation of CaMKI, CaMKIV, protein kinase B, and ERK signaling pathways.

Original languageEnglish (US)
Pages (from-to)52191-52199
Number of pages9
JournalJournal of Biological Chemistry
Volume279
Issue number50
DOIs
StatePublished - Dec 10 2004

Fingerprint

Calcium-Calmodulin-Dependent Protein Kinase Kinase
14-3-3 Proteins
Phosphotransferases
Protein Kinases
Calcium-Calmodulin-Dependent Protein Kinases
Phosphorylation
Rats
Calcium
Okadaic Acid
Proto-Oncogene Proteins c-akt
Mutagenesis
Neuronal Plasticity
MAP Kinase Signaling System
Phosphoprotein Phosphatases
Colforsin
Transcription
Site-Directed Mutagenesis
Cyclic AMP-Dependent Protein Kinases
Mitogen-Activated Protein Kinases
Neurons

ASJC Scopus subject areas

  • Biochemistry

Cite this

Inhibition of calcium/calmodulin-dependent protein kinase kinase by protein 14-3-3. / Davare, Monika; Saneyoshi, Takeo; Guire, Eric S.; Nygaard, Sean C.; Soderling, Thomas.

In: Journal of Biological Chemistry, Vol. 279, No. 50, 10.12.2004, p. 52191-52199.

Research output: Contribution to journalArticle

Davare, Monika ; Saneyoshi, Takeo ; Guire, Eric S. ; Nygaard, Sean C. ; Soderling, Thomas. / Inhibition of calcium/calmodulin-dependent protein kinase kinase by protein 14-3-3. In: Journal of Biological Chemistry. 2004 ; Vol. 279, No. 50. pp. 52191-52199.
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