TY - JOUR
T1 - Endogenously activated mGlu5 metabotropic glutamate receptors sustain the increase in c-Myc expression induced by leukaemia inhibitory factor in cultured mouse embryonic stem cells
AU - Spinsanti, Paola
AU - De Vita, Teresa
AU - Di Castro, Sara
AU - Storto, Marianna
AU - Formisano, Pietro
AU - Nicoletti, Ferdinando
AU - Melchiorri, Daniela
PY - 2006/10
Y1 - 2006/10
N2 - We have shown that endogenous activation of type 5 metabotropic glutamate (mGlu5) receptors supports the maintenance of a pluripotent, undifferentiated state in D3 mouse embryonic stem cells cultured in the presence of leukaemia inhibitory factor (LIF). Here, we examined the interaction between LIF and mGlu5 receptors using as a read-out the immediate early gene, c-Myc. The selective mGlu5 receptor antagonist, 2-methyl-6-(phenylenthynyl)pyridine (MPEP; 1 μm), reduced the increase in c-Myc protein levels induced by LIF by enhancing c-Myc ubiquitination. A reduction in c-Myc levels was also observed following small interfering RNA-mediated mGlu5 receptor gene silencing. MPEP reduced glycogen synthase kinase-3β phosphorylation on Ser9, but increased phosphorylation of the phosphatidylinositol-3-kinase (PI-3-K) substrate, AKT. In our hands, activated PI-3-K reduced the stability of c-Myc, because (i) the PI-3-K inhibitor, LY294002, prevented the reduction in c-Myc levels induced by MPEP; and (ii) over-expression of AKT promoted c-Myc ubiquitination. All effects of MPEP were mimicked by protein kinase C (PKC) inhibitors and reversed by the PKC activator, tetradecanoylphorbol-13-acetate. We conclude that endogenous activation of mGlu5 receptors sustains the increase in c-Myc induced by LIF in embryonic stem cells by inhibiting both glycogen synthase kinase-3β and PI-3-K, both effects resulting from the activation of PKC.
AB - We have shown that endogenous activation of type 5 metabotropic glutamate (mGlu5) receptors supports the maintenance of a pluripotent, undifferentiated state in D3 mouse embryonic stem cells cultured in the presence of leukaemia inhibitory factor (LIF). Here, we examined the interaction between LIF and mGlu5 receptors using as a read-out the immediate early gene, c-Myc. The selective mGlu5 receptor antagonist, 2-methyl-6-(phenylenthynyl)pyridine (MPEP; 1 μm), reduced the increase in c-Myc protein levels induced by LIF by enhancing c-Myc ubiquitination. A reduction in c-Myc levels was also observed following small interfering RNA-mediated mGlu5 receptor gene silencing. MPEP reduced glycogen synthase kinase-3β phosphorylation on Ser9, but increased phosphorylation of the phosphatidylinositol-3-kinase (PI-3-K) substrate, AKT. In our hands, activated PI-3-K reduced the stability of c-Myc, because (i) the PI-3-K inhibitor, LY294002, prevented the reduction in c-Myc levels induced by MPEP; and (ii) over-expression of AKT promoted c-Myc ubiquitination. All effects of MPEP were mimicked by protein kinase C (PKC) inhibitors and reversed by the PKC activator, tetradecanoylphorbol-13-acetate. We conclude that endogenous activation of mGlu5 receptors sustains the increase in c-Myc induced by LIF in embryonic stem cells by inhibiting both glycogen synthase kinase-3β and PI-3-K, both effects resulting from the activation of PKC.
KW - c-Myc
KW - Embryonic stem cells
KW - Leukaemia inhibitory factor
KW - Protein kinase C
KW - Self-renewal
KW - Type 5 metabotropic glutamate receptors
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U2 - 10.1111/j.1471-4159.2006.04038.x
DO - 10.1111/j.1471-4159.2006.04038.x
M3 - Article
C2 - 16987252
AN - SCOPUS:33748684592
VL - 99
SP - 299
EP - 307
JO - Journal of Neurochemistry
JF - Journal of Neurochemistry
SN - 0022-3042
IS - 1
ER -