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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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NF-κB activation is modulated by FAIM. (A) PC12 cells were stably transfected with an empty vector (Neo), with the FAIM-S expression plasmid, or with the FAIM RNAi plasmid. Cells were then transiently transfected with an NF-κB–dependent reporter vector (LTR KB) or with the equivalent control vector lacking NF-κB binding sites (LTR deltaKB). After 24 h, the cells were treated with NGF (100 ng/ml) for the indicated times and luciferase activity was measured in the cell lysates using a Luciferase Assay System. Luciferase values were normalized to protein concentration (RLU/μg of protein). (B) Immunocytochemistry was performed to detect the p65 subunit of NF-κB in similar cells as described in A. Representative low and high magnification photomicrographs are shown (arrowheads indicate the selected cells illustrated in the insets). Bar, 25 μm. (C) Graph showing the percentage of nuclear p65 staining in the different experimental conditions. The mean ± SEM of three independent experiments are shown.
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fig6: NF-κB activation is modulated by FAIM. (A) PC12 cells were stably transfected with an empty vector (Neo), with the FAIM-S expression plasmid, or with the FAIM RNAi plasmid. Cells were then transiently transfected with an NF-κB–dependent reporter vector (LTR KB) or with the equivalent control vector lacking NF-κB binding sites (LTR deltaKB). After 24 h, the cells were treated with NGF (100 ng/ml) for the indicated times and luciferase activity was measured in the cell lysates using a Luciferase Assay System. Luciferase values were normalized to protein concentration (RLU/μg of protein). (B) Immunocytochemistry was performed to detect the p65 subunit of NF-κB in similar cells as described in A. Representative low and high magnification photomicrographs are shown (arrowheads indicate the selected cells illustrated in the insets). Bar, 25 μm. (C) Graph showing the percentage of nuclear p65 staining in the different experimental conditions. The mean ± SEM of three independent experiments are shown.

Mentions: NGF signaling through p75NTR and TrkA has been shown to be critical in the regulation of neurite outgrowth (Davies, 2000), and one of the pathways activated by NGF through these receptors is the NF-κB pathway (Wood, 1995; Carter et al., 1996; Yoon et al., 1998; Hamanoue et al., 1999; Foehr et al., 2000). To investigate whether or not FAIM-induced neurite outgrowth was mediated by NF-κB, FAIM stably transfected PC12 cells were transiently transfected with an NF-κB–dependent luciferase reporter construct or a control construct lacking the NF-κB site (Rodriguez et al., 1996) and assayed for luciferase activity after NGF treatment. FAIM-transfected cells showed a marked increase in luciferase activity after 6–9 h of stimulation with NGF compared with the Neo-transfected cells, indicating that NF-κB activation is potentiated by FAIM expression (Fig. 6 A). In cells that were stably transfected with RNAi2, the levels of luciferase activity were found to be significantly decreased at all the time points analyzed (Fig. 6 A).


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)

NF-κB activation is modulated by FAIM. (A) PC12 cells were stably transfected with an empty vector (Neo), with the FAIM-S expression plasmid, or with the FAIM RNAi plasmid. Cells were then transiently transfected with an NF-κB–dependent reporter vector (LTR KB) or with the equivalent control vector lacking NF-κB binding sites (LTR deltaKB). After 24 h, the cells were treated with NGF (100 ng/ml) for the indicated times and luciferase activity was measured in the cell lysates using a Luciferase Assay System. Luciferase values were normalized to protein concentration (RLU/μg of protein). (B) Immunocytochemistry was performed to detect the p65 subunit of NF-κB in similar cells as described in A. Representative low and high magnification photomicrographs are shown (arrowheads indicate the selected cells illustrated in the insets). Bar, 25 μm. (C) Graph showing the percentage of nuclear p65 staining in the different experimental conditions. The mean ± SEM of three independent experiments are shown.
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Related In: Results  -  Collection

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fig6: NF-κB activation is modulated by FAIM. (A) PC12 cells were stably transfected with an empty vector (Neo), with the FAIM-S expression plasmid, or with the FAIM RNAi plasmid. Cells were then transiently transfected with an NF-κB–dependent reporter vector (LTR KB) or with the equivalent control vector lacking NF-κB binding sites (LTR deltaKB). After 24 h, the cells were treated with NGF (100 ng/ml) for the indicated times and luciferase activity was measured in the cell lysates using a Luciferase Assay System. Luciferase values were normalized to protein concentration (RLU/μg of protein). (B) Immunocytochemistry was performed to detect the p65 subunit of NF-κB in similar cells as described in A. Representative low and high magnification photomicrographs are shown (arrowheads indicate the selected cells illustrated in the insets). Bar, 25 μm. (C) Graph showing the percentage of nuclear p65 staining in the different experimental conditions. The mean ± SEM of three independent experiments are shown.
Mentions: NGF signaling through p75NTR and TrkA has been shown to be critical in the regulation of neurite outgrowth (Davies, 2000), and one of the pathways activated by NGF through these receptors is the NF-κB pathway (Wood, 1995; Carter et al., 1996; Yoon et al., 1998; Hamanoue et al., 1999; Foehr et al., 2000). To investigate whether or not FAIM-induced neurite outgrowth was mediated by NF-κB, FAIM stably transfected PC12 cells were transiently transfected with an NF-κB–dependent luciferase reporter construct or a control construct lacking the NF-κB site (Rodriguez et al., 1996) and assayed for luciferase activity after NGF treatment. FAIM-transfected cells showed a marked increase in luciferase activity after 6–9 h of stimulation with NGF compared with the Neo-transfected cells, indicating that NF-κB activation is potentiated by FAIM expression (Fig. 6 A). In cells that were stably transfected with RNAi2, the levels of luciferase activity were found to be significantly decreased at all the time points analyzed (Fig. 6 A).

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