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The death receptor antagonist FAIM promotes neurite outgrowth by a mechanism that depends on ERK and NF-kapp B signaling.

Sole C, Dolcet X, Segura MF, Gutierrez H, Diaz-Meco MT, Gozzelino R, Sanchis D, Bayascas JR, Gallego C, Moscat J, Davies AM, Comella JX - J. Cell Biol. (2004)

Bottom Line: Whereas FAIM overexpression greatly enhanced neurite outgrowth from PC12 cells and sympathetic neurons grown with nerve growth factor (NGF), reduction of endogenous FAIM levels by RNAi decreased neurite outgrowth in these cells.The effect of FAIM on neurite outgrowth was also blocked by inhibition of the Ras-ERK pathway.These results reveal a new function of FAIM in promoting neurite outgrowth by a mechanism involving activation of the Ras-ERK pathway and NF-kappa B.

View Article: PubMed Central - PubMed

Affiliation: Group Cell Signalling and Apoptosis, Department of Basic Medical Sciences, Universitat de Lleida, 25008 Lleida, Spain.

ABSTRACT
Fas apoptosis inhibitory molecule (FAIM) is a protein identified as an antagonist of Fas-induced cell death. We show that FAIM overexpression fails to rescue neurons from trophic factor deprivation, but exerts a marked neurite growth-promoting action in different neuronal systems. Whereas FAIM overexpression greatly enhanced neurite outgrowth from PC12 cells and sympathetic neurons grown with nerve growth factor (NGF), reduction of endogenous FAIM levels by RNAi decreased neurite outgrowth in these cells. FAIM overexpression promoted NF-kappa B activation, and blocking this activation by using a super-repressor I kappa B alpha or by carrying out experiments using cortical neurons from mice that lack the p65 NF-kappa B subunit prevented FAIM-induced neurite outgrowth. The effect of FAIM on neurite outgrowth was also blocked by inhibition of the Ras-ERK pathway. Finally, we show that FAIM interacts with both Trk and p75 neurotrophin receptor NGF receptors in a ligand-dependent manner. These results reveal a new function of FAIM in promoting neurite outgrowth by a mechanism involving activation of the Ras-ERK pathway and NF-kappa B.

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Effects of FAIM on ERK pathway stimulation. (A) PC12 cells stably transfected with empty vector or with FAIM-S were treated with NGF (100 ng/ml) or with NGF plus 50 μM of PD98059 for 1 d. Histogram shows the neurite length measurements of the cultures transfected with the indicated cDNA. Significant differences are indicated (*, P < 0.05; **, P < 0.001; t test). (B) PC12 cells stably transfected with empty vector (N) or FAIM-S (S) were serum starved for 12 h, pretreated (+) or not (−) with 50 μM PD98059 for 30 min, and stimulated (+) or not (−) with 100 ng/ml NGF for 5 min. Protein extracts were analyzed by Western blotting with an anti–phospho-ERK antibody (top) and stripped and reprobed with anti–pan-ERK and anti-tubulin antibodies (middle) to control the protein loading. Anti-FLAG immunoblotting (bottom) confirmed FAIM-S overexpression in transfected PC12 cells. White lines indicate that intervening lanes have been spliced. (C) PC12 cells stably transfected with empty vector (N), FAIM-S (S), or RNAi2 (R) were serum starved for 12 h and stimulated with 100 ng/ml NGF for the indicated times. Protein extracts were processed as in B.
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fig5: Effects of FAIM on ERK pathway stimulation. (A) PC12 cells stably transfected with empty vector or with FAIM-S were treated with NGF (100 ng/ml) or with NGF plus 50 μM of PD98059 for 1 d. Histogram shows the neurite length measurements of the cultures transfected with the indicated cDNA. Significant differences are indicated (*, P < 0.05; **, P < 0.001; t test). (B) PC12 cells stably transfected with empty vector (N) or FAIM-S (S) were serum starved for 12 h, pretreated (+) or not (−) with 50 μM PD98059 for 30 min, and stimulated (+) or not (−) with 100 ng/ml NGF for 5 min. Protein extracts were analyzed by Western blotting with an anti–phospho-ERK antibody (top) and stripped and reprobed with anti–pan-ERK and anti-tubulin antibodies (middle) to control the protein loading. Anti-FLAG immunoblotting (bottom) confirmed FAIM-S overexpression in transfected PC12 cells. White lines indicate that intervening lanes have been spliced. (C) PC12 cells stably transfected with empty vector (N), FAIM-S (S), or RNAi2 (R) were serum starved for 12 h and stimulated with 100 ng/ml NGF for the indicated times. Protein extracts were processed as in B.

Mentions: Because ERK signaling has been implicated in neurotrophin-induced neurite outgrowth, we investigated the role of this signaling pathway in mediating the neurite outgrowth–inducing effects of FAIM by using pharmacological agents to inhibit ERK signaling. Addition of PD98059, a MEK inhibitor, markedly reduced the extent of neurite outgrowth in FAIM-transfected cells (Fig. 5 A). These results suggest that activation of the ERK pathway is required for the neurite growth–inducing effect of FAIM overexpression. To confirm that the doses of PD98059 completely abolished ERK activation in FAIM-transfected cells as well as in the Neo cells, Western blot analysis was performed using a specific anti–phospho-ERK1/2 antibody. 30-min pretreatment of the stably transfected PC12 cells with 50 μM PD98059 completely abolished ERK1/2 phosphorylation (Fig. 5 B).


The death receptor antagonist FAIM promotes neurite outgrowth by a mechanism that depends on ERK and NF-kapp B signaling.

Sole C, Dolcet X, Segura MF, Gutierrez H, Diaz-Meco MT, Gozzelino R, Sanchis D, Bayascas JR, Gallego C, Moscat J, Davies AM, Comella JX - J. Cell Biol. (2004)

Effects of FAIM on ERK pathway stimulation. (A) PC12 cells stably transfected with empty vector or with FAIM-S were treated with NGF (100 ng/ml) or with NGF plus 50 μM of PD98059 for 1 d. Histogram shows the neurite length measurements of the cultures transfected with the indicated cDNA. Significant differences are indicated (*, P < 0.05; **, P < 0.001; t test). (B) PC12 cells stably transfected with empty vector (N) or FAIM-S (S) were serum starved for 12 h, pretreated (+) or not (−) with 50 μM PD98059 for 30 min, and stimulated (+) or not (−) with 100 ng/ml NGF for 5 min. Protein extracts were analyzed by Western blotting with an anti–phospho-ERK antibody (top) and stripped and reprobed with anti–pan-ERK and anti-tubulin antibodies (middle) to control the protein loading. Anti-FLAG immunoblotting (bottom) confirmed FAIM-S overexpression in transfected PC12 cells. White lines indicate that intervening lanes have been spliced. (C) PC12 cells stably transfected with empty vector (N), FAIM-S (S), or RNAi2 (R) were serum starved for 12 h and stimulated with 100 ng/ml NGF for the indicated times. Protein extracts were processed as in B.
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fig5: Effects of FAIM on ERK pathway stimulation. (A) PC12 cells stably transfected with empty vector or with FAIM-S were treated with NGF (100 ng/ml) or with NGF plus 50 μM of PD98059 for 1 d. Histogram shows the neurite length measurements of the cultures transfected with the indicated cDNA. Significant differences are indicated (*, P < 0.05; **, P < 0.001; t test). (B) PC12 cells stably transfected with empty vector (N) or FAIM-S (S) were serum starved for 12 h, pretreated (+) or not (−) with 50 μM PD98059 for 30 min, and stimulated (+) or not (−) with 100 ng/ml NGF for 5 min. Protein extracts were analyzed by Western blotting with an anti–phospho-ERK antibody (top) and stripped and reprobed with anti–pan-ERK and anti-tubulin antibodies (middle) to control the protein loading. Anti-FLAG immunoblotting (bottom) confirmed FAIM-S overexpression in transfected PC12 cells. White lines indicate that intervening lanes have been spliced. (C) PC12 cells stably transfected with empty vector (N), FAIM-S (S), or RNAi2 (R) were serum starved for 12 h and stimulated with 100 ng/ml NGF for the indicated times. Protein extracts were processed as in B.
Mentions: Because ERK signaling has been implicated in neurotrophin-induced neurite outgrowth, we investigated the role of this signaling pathway in mediating the neurite outgrowth–inducing effects of FAIM by using pharmacological agents to inhibit ERK signaling. Addition of PD98059, a MEK inhibitor, markedly reduced the extent of neurite outgrowth in FAIM-transfected cells (Fig. 5 A). These results suggest that activation of the ERK pathway is required for the neurite growth–inducing effect of FAIM overexpression. To confirm that the doses of PD98059 completely abolished ERK activation in FAIM-transfected cells as well as in the Neo cells, Western blot analysis was performed using a specific anti–phospho-ERK1/2 antibody. 30-min pretreatment of the stably transfected PC12 cells with 50 μM PD98059 completely abolished ERK1/2 phosphorylation (Fig. 5 B).

Bottom Line: Whereas FAIM overexpression greatly enhanced neurite outgrowth from PC12 cells and sympathetic neurons grown with nerve growth factor (NGF), reduction of endogenous FAIM levels by RNAi decreased neurite outgrowth in these cells.The effect of FAIM on neurite outgrowth was also blocked by inhibition of the Ras-ERK pathway.These results reveal a new function of FAIM in promoting neurite outgrowth by a mechanism involving activation of the Ras-ERK pathway and NF-kappa B.

View Article: PubMed Central - PubMed

Affiliation: Group Cell Signalling and Apoptosis, Department of Basic Medical Sciences, Universitat de Lleida, 25008 Lleida, Spain.

ABSTRACT
Fas apoptosis inhibitory molecule (FAIM) is a protein identified as an antagonist of Fas-induced cell death. We show that FAIM overexpression fails to rescue neurons from trophic factor deprivation, but exerts a marked neurite growth-promoting action in different neuronal systems. Whereas FAIM overexpression greatly enhanced neurite outgrowth from PC12 cells and sympathetic neurons grown with nerve growth factor (NGF), reduction of endogenous FAIM levels by RNAi decreased neurite outgrowth in these cells. FAIM overexpression promoted NF-kappa B activation, and blocking this activation by using a super-repressor I kappa B alpha or by carrying out experiments using cortical neurons from mice that lack the p65 NF-kappa B subunit prevented FAIM-induced neurite outgrowth. The effect of FAIM on neurite outgrowth was also blocked by inhibition of the Ras-ERK pathway. Finally, we show that FAIM interacts with both Trk and p75 neurotrophin receptor NGF receptors in a ligand-dependent manner. These results reveal a new function of FAIM in promoting neurite outgrowth by a mechanism involving activation of the Ras-ERK pathway and NF-kappa B.

Show MeSH
Related in: MedlinePlus