Stimulation of neuropeptide Y gene expression by brain-derived neurotrophic factor requires both the phospholipase Cgamma and Shc binding sites on its receptor, TrkB.
In PC12 cells, it has been previously reported that nerve growth factor stimulates neuropeptide Y (NPY) gene expression. In the current study we examined the signalling pathways involved in this effect by transiently expressing in PC12 cells the receptor (TrkB) for the related neurotrophin, brain-derived neurotrophic factor (BDNF). BDNF caused a 3-fold induction of luciferase expression from a transiently co-transfected plasmid possessing the firefly luciferase gene under the control of the NPY promoter. This effect of BDNF was completely blocked by either a Y484F mutation in TrkB (which blocks high-affinity Shc binding to TrkB) or by a Y785F substitution [which blocks the binding, phosphorylation and activation of phospholipase Cgamma (PLCgamma)]. Activation of the NPY promoter by neurotrophin-3 in PC12 cells overexpressing TrkC was also completely blocked by a naturally occurring kinase insert which prevents the high-affinity binding of Shc and PLCgamma. NPY promoter activation by BDNF was blocked by PD98059, suggesting a role for mitogen-activated protein kinase (MAP kinase). Stimulation of NPY gene expression by PMA, but not by BDNF, was blocked by Ro-31-8220, a protein kinase C inhibitor, excluding a role for this serine/threonine protein kinase in the effect of BDNF. In addition, BDNF did not cause an elevation in cytosolic Ca2+ concentration. Taken together, our results suggest that stimulation of the NPY promoter by BDNF requires the simultaneous activation of two distinct pathways; one involves Shc and MAP kinase, and the other appears to be PLCgamma-independent but requires an intact tyrosine-785 on TrkB and so may involve an effector of TrkB signalling that remains to be identified.
Williams
A G
AG
Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK.
Hargreaves
A C
AC
Gunn-Moore
F J
FJ
Tavaré
J M
JM
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation; D015870:Gene Expression / Q000187:drug effects; D007527:Isoenzymes / Q000378:metabolism*; D009154:Mutation; D009414:Nerve Growth Factors / Q000494:pharmacology*; D009478:Neuropeptide Y / Q000096:biosynthesis*; D016716:PC12 Cells; D051966:Phospholipase C gamma; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D020813:Receptor, trkB; D020812:Receptor, trkC; D017475:Receptors, Nerve Growth Factor / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013268:Stimulation, Chemical; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D019208:Brain-Derived Neurotrophic Factor; D007527:Isoenzymes; D009414:Nerve Growth Factors; D009478:Neuropeptide Y; D011506:Proteins; D017475:Receptors, Nerve Growth Factor; D055765:Shc Signaling Adaptor Proteins; C528035:Shc1 protein, rat; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D020794:Receptor Protein-Tyrosine Kinases; D020813:Receptor, trkB; D020812:Receptor, trkC; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium