Recruitment of active glycogen synthase kinase-3 into neuronal lipid rafts

doi:10.1016/j.bbrc.2006.05.087

Biochemical and Biophysical Research Communications

Volume 345, Issue 4, 14 July 2006, Pages 1643-1648

https://doi.org/10.1016/j.bbrc.2006.05.087 Get rights and content

Abstract

Glycogen synthase kinase (GSK)-3β has emerged as a key molecule that regulates neuronal apoptosis. To examine the molecular mechanism(s) through which GSK-3β regulates this process, we studied the subcellular localization of GSK-3β following exposure of the cells to well-characterized apoptotic stimuli. Here, we report that the induction of apoptosis by withdrawal of serum and potassium triggers dephosphorylation of GSK-3β at serine 9 and subsequent translocation of these molecules into neuronal lipid raft microdomains. Inhibition of GSK-3β by small molecule inhibitors blocks specific phosphorylation of lipid raft associated protein Tau. Consistent with the notion that the lipid raft domains may serve as a platform for the cellular signaling complexes, disruption of lipid rafts protected neurons from apoptosis induced by withdrawal of serum and potassium as well as by HIV-1 Tat. Our observations reveal novel interaction of GSK-3β and raft domains, and suggest that such interaction could contribute to neuronal apoptosis.

Section snippets

Materials and methods

Neuronal cell cultures. Primary cultures of cerebellar granule cells (CGNs) were prepared from 7 to 8-day-old Sprague–Dawley rat (Charles River) cerebella, as previously described [14]. Cells were cultured for 6–7 days in Basal Eagle’s medium (BME) supplemented with 10% heat-inactivated fetal calf serum, 20 mM KCl, 2 mM glutamine, and 100 μg/ml gentamicin (K25 + S; BME contains 5 mM KCl). Cytosine-β-arabinofuranoside (10 μM) was added 24 h after plating to limit the number of non-neuronal cells to less

Results and discussion

Isolation of lipid rafts from cells using relative insolubility of these microdomains in Triton X-100 and their separation by discontinuous (40/30/5%) sucrose gradient has been widely used to characterize dynamic assemblies of proteins on rafts and concurrent regulation of various signaling pathways [16], [18]. We used this well-characterized method to purify lipid rafts from PC12 cells. As shown in Fig. 1, we collected 11 gradient fractions, fractions 3–5 at the 5%–30% interface contained the

Acknowledgment

This research was supported by the National Institutes of Health.

References (29)

J.R. Woodgett
Recent advances in the protein kinase B signaling pathway
Curr. Opin. Cell Biol.
(2005)
N. Goode et al.
Differential regulation of glycogen synthase kinase-3 β by protein kinase C isotypes
J. Biol. Chem.
(1992)
H. Eldar-Finkelman et al.
Inactivation of glycogen synthase kinase-3 by epidermal growth factor is mediated by mitogen-activated protein kinase/p90 ribosomal protein S6 kinase signaling pathway in NIH/3T3 cells
J. Biol. Chem.
(1995)
N. Jin et al.
Opposite effects of lithium and valproic acid on trophic factor deprivation-induced glycogen synthase kinase-3 activation, c-Jun expression and neuronal cell death
Neuropharmacology
(2005)
A. Takashima et al.
Exposure of rat hippocampal neurons to amyloid β peptide (25–35) induces the inactivation of phosphatidyl inositol-3 kinase and the activation of tau protein kinase I/glycogen synthase kinase-3 β
Neurosci. Lett.
(1996)
C.A. Grimes et al.
The multifaceted roles of glycogen synthase kinase 3β in cellular signaling
Prog. Neurobiol.
(2001)
R.S. Jope et al.
The glamour and gloom of glycogen synthase kinase-3
Trends Biochem. Sci.
(2004)
B.A. Tsui-Pierchala et al.
Lipid rafts in neuronal signaling and function
Trends Neurosci.
(2002)
A.D. Kovacs et al.
Selective and AMPA receptor-dependent astrocyte death following prolonged blockade of glutamate reuptake in rat cerebellar cultures
Exp. Neurol.
(2002)
A. Mukherjee et al.
Lipid-dependent recruitment of neuronal Src to lipid rafts in the brain
J. Biol. Chem.
(2003)

H. Cui et al.

Inhibition of glycogen synthase kinase 3beta activity regulates proliferation of cultured cerebellar granule cells

Brain Res. Dev. Brain Res.

(1998)

C. Tanji et al.

A-kinase anchoring protein AKAP220 binds to glycogen synthase kinase-3β (GSK-3β) and mediates protein kinase A-dependent inhibition of GSK-3β

J. Biol. Chem.

(2002)

N. Reinton et al.

Localization of a novel human A-kinase-anchoring protein, hAKAP220, during spermatogenesis

Dev. Biol.

(2000)

H.S. Kim et al.

Regulation of angiogenesis by glycogen synthase kinase-3β

J. Biol. Chem.

(2002)

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Recruitment of active glycogen synthase kinase-3 into neuronal lipid rafts

Abstract

Section snippets

Materials and methods

Results and discussion

Acknowledgment

Curr. Opin. Cell Biol.

J. Biol. Chem.

J. Biol. Chem.

Neuropharmacology

Neurosci. Lett.

Prog. Neurobiol.

Trends Biochem. Sci.

Trends Neurosci.

Exp. Neurol.

J. Biol. Chem.

Brain Res. Dev. Brain Res.

J. Biol. Chem.

Dev. Biol.

J. Biol. Chem.