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BDNF-induced recruitment of TrkB receptor into neuronal lipid rafts: roles in synaptic modulation.

Suzuki S, Numakawa T, Shimazu K, Koshimizu H, Hara T, Hatanaka H, Mei L, Lu B, Kojima M - J. Cell Biol. (2004)

Bottom Line: Moreover, disruption of lipid rafts prevented potentiating effects of BDNF on transmitter release in cultured neurons and synaptic response to tetanus in hippocampal slices.In contrast, lipid rafts are not required for BDNF regulation of neuronal survival.Thus, ligand-induced TrkB translocation into lipid rafts may represent a signaling mechanism selective for synaptic modulation by BDNF in the central nervous system.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Cell Engineering, National Institute for Advanced Industrial Science and Technology, Ikeda, Osaka, Japan.

ABSTRACT
Brain-derived neurotrophic factor (BDNF) plays an important role in synaptic plasticity but the underlying signaling mechanisms remain unknown. Here, we show that BDNF rapidly recruits full-length TrkB (TrkB-FL) receptor into cholesterol-rich lipid rafts from nonraft regions of neuronal plasma membranes. Translocation of TrkB-FL was blocked by Trk inhibitors, suggesting a role of TrkB tyrosine kinase in the translocation. Disruption of lipid rafts by depleting cholesterol from cell surface blocked the ligand-induced translocation. Moreover, disruption of lipid rafts prevented potentiating effects of BDNF on transmitter release in cultured neurons and synaptic response to tetanus in hippocampal slices. In contrast, lipid rafts are not required for BDNF regulation of neuronal survival. Thus, ligand-induced TrkB translocation into lipid rafts may represent a signaling mechanism selective for synaptic modulation by BDNF in the central nervous system.

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Differential activation of Erks and Ark in lipid rafts upon BDNF-induced TrkB-FL translocation. Cultured cortical neurons were treated with or without BDNF (200 ng/ml) for 30 min. (A) Western blot analysis of the distribution of major signaling molecules downstream of TrkB before and after BDNF stimulation. Similar results were obtained from two separate experiments. (B and C) Differential activation of Erks and Akt in lipid rafts and nonrafts after a 30-min treatment with BDNF. Fractions 2 and 6 were immunoblotted. (D) Quantitative analysis of BDNF effect on the activation of Erks and Akt in rafts and nonrafts. For each protein, phosphorylation relative to total amount was quantified in fractions 2 and 6, and shown as relative to that of “+BDNF” in fraction 6. *Indicates significantly different from “−BDNF”; t test; P < 0.05. n = 4 independent experiments. In A–C, immunoblots were performed using antibodies specific for indicated proteins and phospho-proteins.
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fig4: Differential activation of Erks and Ark in lipid rafts upon BDNF-induced TrkB-FL translocation. Cultured cortical neurons were treated with or without BDNF (200 ng/ml) for 30 min. (A) Western blot analysis of the distribution of major signaling molecules downstream of TrkB before and after BDNF stimulation. Similar results were obtained from two separate experiments. (B and C) Differential activation of Erks and Akt in lipid rafts and nonrafts after a 30-min treatment with BDNF. Fractions 2 and 6 were immunoblotted. (D) Quantitative analysis of BDNF effect on the activation of Erks and Akt in rafts and nonrafts. For each protein, phosphorylation relative to total amount was quantified in fractions 2 and 6, and shown as relative to that of “+BDNF” in fraction 6. *Indicates significantly different from “−BDNF”; t test; P < 0.05. n = 4 independent experiments. In A–C, immunoblots were performed using antibodies specific for indicated proteins and phospho-proteins.

Mentions: Phosphorylated tyrosine residues on the TrkB intracellular domain form docking sites for the signaling proteins Shc, Grb2, and PLCγ, leading to activation of MAPK (Erks), PLCγ, and PI3-K pathways (Kaplan and Miller, 2000). We tested whether TrkB-FL, upon BDNF stimulation, carries these signaling molecules into lipid rafts during its translocation. In naïve neurons, Erks extended throughout the gradient (fractions 2–5), whereas Shc, Grb2, p85 subunit of PI3-K, Akt, and PLCγ were primarily localized in the bottom, nonraft fraction (Fig. 4 A, left). BDNF application did not increase the amount of any of these proteins in fraction 2 (Fig. 4 A, right), suggesting that TrkB-FL does not carry its associated signaling molecules into lipid rafts during its translocation. When a milder detergent Triton X-165 was used to prepare rafts (Fig. S4 A, available at http://www.jcb.org/cgi/content/full/jcb.200404106/DC1), many of them (Shc, Grb2, Erks, and PLCγ) were extended throughout the gradient whereas p85 subunit of PI3-K and Akt appeared to be still in nonraft fraction. It was notable that BDNF did not stimulate the translocation of these signaling molecules into rafts (Fig. S4 A). Parallel to these, BDNF-induced translocation of TrkB-FL (Fig. 4 B, left) was accompanied by a significant increase in the phosphorylation of Erks (Figs. 4 B, middle and Fig. 4 D), but not that of Akt (Fig. 4 B, right and Fig. 4 D), in lipid rafts. In nonraft regions, however, both Erks and Akt were activated by BDNF application (Fig. 4, C and D). Thus, although TrkB-FL does not move with its associated proteins during translocation, the translocation of TrkB-FL itself into rafts may be a key event in forming TrkB signaling complex, including TrkB-FL and its associated proteins, in rafts, leading to preferential activation of Erks over Akt in neuronal lipid rafts.


BDNF-induced recruitment of TrkB receptor into neuronal lipid rafts: roles in synaptic modulation.

Suzuki S, Numakawa T, Shimazu K, Koshimizu H, Hara T, Hatanaka H, Mei L, Lu B, Kojima M - J. Cell Biol. (2004)

Differential activation of Erks and Ark in lipid rafts upon BDNF-induced TrkB-FL translocation. Cultured cortical neurons were treated with or without BDNF (200 ng/ml) for 30 min. (A) Western blot analysis of the distribution of major signaling molecules downstream of TrkB before and after BDNF stimulation. Similar results were obtained from two separate experiments. (B and C) Differential activation of Erks and Akt in lipid rafts and nonrafts after a 30-min treatment with BDNF. Fractions 2 and 6 were immunoblotted. (D) Quantitative analysis of BDNF effect on the activation of Erks and Akt in rafts and nonrafts. For each protein, phosphorylation relative to total amount was quantified in fractions 2 and 6, and shown as relative to that of “+BDNF” in fraction 6. *Indicates significantly different from “−BDNF”; t test; P < 0.05. n = 4 independent experiments. In A–C, immunoblots were performed using antibodies specific for indicated proteins and phospho-proteins.
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Related In: Results  -  Collection

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fig4: Differential activation of Erks and Ark in lipid rafts upon BDNF-induced TrkB-FL translocation. Cultured cortical neurons were treated with or without BDNF (200 ng/ml) for 30 min. (A) Western blot analysis of the distribution of major signaling molecules downstream of TrkB before and after BDNF stimulation. Similar results were obtained from two separate experiments. (B and C) Differential activation of Erks and Akt in lipid rafts and nonrafts after a 30-min treatment with BDNF. Fractions 2 and 6 were immunoblotted. (D) Quantitative analysis of BDNF effect on the activation of Erks and Akt in rafts and nonrafts. For each protein, phosphorylation relative to total amount was quantified in fractions 2 and 6, and shown as relative to that of “+BDNF” in fraction 6. *Indicates significantly different from “−BDNF”; t test; P < 0.05. n = 4 independent experiments. In A–C, immunoblots were performed using antibodies specific for indicated proteins and phospho-proteins.
Mentions: Phosphorylated tyrosine residues on the TrkB intracellular domain form docking sites for the signaling proteins Shc, Grb2, and PLCγ, leading to activation of MAPK (Erks), PLCγ, and PI3-K pathways (Kaplan and Miller, 2000). We tested whether TrkB-FL, upon BDNF stimulation, carries these signaling molecules into lipid rafts during its translocation. In naïve neurons, Erks extended throughout the gradient (fractions 2–5), whereas Shc, Grb2, p85 subunit of PI3-K, Akt, and PLCγ were primarily localized in the bottom, nonraft fraction (Fig. 4 A, left). BDNF application did not increase the amount of any of these proteins in fraction 2 (Fig. 4 A, right), suggesting that TrkB-FL does not carry its associated signaling molecules into lipid rafts during its translocation. When a milder detergent Triton X-165 was used to prepare rafts (Fig. S4 A, available at http://www.jcb.org/cgi/content/full/jcb.200404106/DC1), many of them (Shc, Grb2, Erks, and PLCγ) were extended throughout the gradient whereas p85 subunit of PI3-K and Akt appeared to be still in nonraft fraction. It was notable that BDNF did not stimulate the translocation of these signaling molecules into rafts (Fig. S4 A). Parallel to these, BDNF-induced translocation of TrkB-FL (Fig. 4 B, left) was accompanied by a significant increase in the phosphorylation of Erks (Figs. 4 B, middle and Fig. 4 D), but not that of Akt (Fig. 4 B, right and Fig. 4 D), in lipid rafts. In nonraft regions, however, both Erks and Akt were activated by BDNF application (Fig. 4, C and D). Thus, although TrkB-FL does not move with its associated proteins during translocation, the translocation of TrkB-FL itself into rafts may be a key event in forming TrkB signaling complex, including TrkB-FL and its associated proteins, in rafts, leading to preferential activation of Erks over Akt in neuronal lipid rafts.

Bottom Line: Moreover, disruption of lipid rafts prevented potentiating effects of BDNF on transmitter release in cultured neurons and synaptic response to tetanus in hippocampal slices.In contrast, lipid rafts are not required for BDNF regulation of neuronal survival.Thus, ligand-induced TrkB translocation into lipid rafts may represent a signaling mechanism selective for synaptic modulation by BDNF in the central nervous system.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Cell Engineering, National Institute for Advanced Industrial Science and Technology, Ikeda, Osaka, Japan.

ABSTRACT
Brain-derived neurotrophic factor (BDNF) plays an important role in synaptic plasticity but the underlying signaling mechanisms remain unknown. Here, we show that BDNF rapidly recruits full-length TrkB (TrkB-FL) receptor into cholesterol-rich lipid rafts from nonraft regions of neuronal plasma membranes. Translocation of TrkB-FL was blocked by Trk inhibitors, suggesting a role of TrkB tyrosine kinase in the translocation. Disruption of lipid rafts by depleting cholesterol from cell surface blocked the ligand-induced translocation. Moreover, disruption of lipid rafts prevented potentiating effects of BDNF on transmitter release in cultured neurons and synaptic response to tetanus in hippocampal slices. In contrast, lipid rafts are not required for BDNF regulation of neuronal survival. Thus, ligand-induced TrkB translocation into lipid rafts may represent a signaling mechanism selective for synaptic modulation by BDNF in the central nervous system.

Show MeSH
Related in: MedlinePlus