An essential role for the SHIP2-dependent negative feedback loop in neuritogenesis of nerve growth factor-stimulated PC12 cells.
The local accumulation of phosphatidylinositol (3,4,5) trisphosphate (PIP(3)) and resulting activation of Rac1/Cdc42 play a critical role in nerve growth factor (NGF)-induced neurite outgrowth. To further explore the mechanism, we visualized PIP(3), phosphatidylinositol (3,4) bisphosphate, and Rac1/Cdc42 activities by fluorescence resonance energy transfer (FRET) imaging in NGF-stimulated PC12 cells. Based on the obtained FRET images, and with the help of in silico kinetic reaction model, we predicted that PI-5-phosphatase negatively regulates PIP(3) upon NGF stimulation. In agreement with this model, depletion of Src homology 2 domain-containing inositol polyphosphate 5-phosphatase 2 (SHIP2) markedly potentiated NGF-induced Rac1/Cdc42 activation and PIP(3) accumulation and considerably increased the number and the length of neurites in phosphate and tensin homologue-depleted PC12 cells. Further refinement of the computational model predicted Rac1 regulation of PI3-kinase and SHIP2, which was also validated experimentally. We propose that the SHIP2-mediated negative feedback on PIP(3) coordinately works with the PI3-kinase-mediated positive feedback to form an initial protrusive pattern and, later, to punctuate the PIP(3) accumulation to maintain proper neurite outgrowth.
Aoki
Kazuhiro
K
Department of Pathology and Biology of Diseases, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Nakamura
Takeshi
T
Inoue
Takanari
T
Meyer
Tobias
T
Matsuda
Michiyuki
M
D000818:Animals; D003198:Computer Simulation; D025461:Feedback, Physiological; D031541:Fluorescence Resonance Energy Transfer; D000072181:Inositol Polyphosphate 5-Phosphatases; D007700:Kinetics; D020932:Nerve Growth Factor / Q000494:pharmacology*; D009415:Nerve Net / Q000254:growth & development; D016501:Neurites / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D016716:PC12 Cells; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism / Q000502:physiology*; D051381:Rats; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
D018129:Phosphatidylinositol Phosphates; C060974:phosphatidylinositol 3,4,5-triphosphate; D020932:Nerve Growth Factor; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; C000606436:Inppl1 protein, rat; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; C499134:Rac1 protein, rat; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein