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Bone morphogenetic protein signalling in NGF-stimulated PC12 cells
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
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2003 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 307, no 3, 632-639 p.Article in journal (Refereed) Published
Abstract [en]

Bone morphogenetic proteins (BMPs) are shown to potentiate NGF-induced neuronal differentiation in PC12 phaeochromocytoma cells grown on collagen under low-serum conditions. Whereas, cell bodies remained rounded in control medium or with only BMPs present, addition of BMP4 or BMP6 robustly increased the neuritogenic effect of NGF within 2 days. NGF-increased phosphorylation of p44(Erk1) and p42(Erk2) between 2 and 24h was unaffected by addition of BMP6. PC12 cells transfected with the SBE(4x)-luc reporter showed that BMP4 significantly increased receptor-activated Smad activity. Expression of constitutively active BMP receptor ALK2 activating Smad1 and Smad5 resulted in a strong increase in the SBE(4x)-luc reporter response. Adding the inhibitory Smad7 drastically reduced this signal. In contrast to wild-type (wt) Smad5, a Smad5 variant lacking five Erk phosphorylation sites in the linker region (designated Smad5/5SA) showed a strong background transcriptional activity. A fusion construct (Gal4-Smad5/5SA) was also highly transcriptionally active. Addition of the MEK inhibitor U0126 to PC12 cells expressing Gal4-Smad5/wt did not increase background transcriptional activity. However, upon activation by constitutively active ALK2 both Gal4-Smad5/wt and Gal4-Smad5/5SA strongly stimulated transcription. The data show that serine residues of the linker region of Smad5 reduce spontaneous transcriptional activity and that NGF-activated Erk does not antagonise BMP signalling at this site. Hence, NGF and BMP signals are likely to interact further downstream at the transcriptional level in neuronal differentiation of the PC12 cells.

Place, publisher, year, edition, pages
2003. Vol. 307, no 3, 632-639 p.
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-90338DOI: 10.1016/S0006-291X(03)01236-1PubMedID: 12893270OAI: oai:DiVA.org:uu-90338DiVA: diva2:162655
Available from: 2003-04-24 Created: 2003-04-24 Last updated: 2013-07-25Bibliographically approved
In thesis
1. Experimental Studies of BMP Signalling in Neuronal Cells
Open this publication in new window or tab >>Experimental Studies of BMP Signalling in Neuronal Cells
2003 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The developing nervous system depends largely on extracellular cues to shape its complex network of neurons. Classically, neurotrophins are known to be important mediators in this process. More recently, Bone Morphogenetic Proteins (BMPs), belonging to the Transforming Growth Factor beta (TGFβ) superfamily of secreted cytokines, have been shown to exert a wide range of effects, such as cellular growth, differentiation, survival and apoptosis, both in the developing and adult nervous system. They signal via serine/threonine kinase receptor essentially to the Smad pathway, which carries the signal to the nucleus where the transcription of target genes is regulated.

This thesis investigates the functions of BMPs in the nervous system, using a set of different models. Firstly, a targeted deletion of GDF10 (BMP3b) in the mouse was established to evaluate the role of this growth/differentiation factor in the hippocampal formation, a brain area known to be involved in memory processing. Other members of the TGFβ superfamily likely compensate for the lack of GDF10, since no detectable alterations in hippocampal function or gene transcription profile have been found. Secondly, a mouse model was set up, with the aim to study impaired BMP-signalling in dopaminergic neurons. The tyrosine hydroxylase (TH) locus was used to drive the expression of dominant negative BMP receptors by means of bicistronic mRNAs. TH is the rate-limiting enzyme in the biosynthesis of catecholamine and the mice described, show a graded decrease of TH-activity resulting in severe to mild dopamine deficiency. The contribution of the dominant negative BMP receptors to the phenotype is however secondary to the apparent TH hypomorphism. The final theme of this thesis is the potentiating effects of BMPs on neurotrophin-induced neurite outgrowth as studied in explanted ganglia from chick embryos and in the rat phaeochromocytoma cell line PC12. A number of pharmacological inhibitors of intracellular signalling kinases were applied to the cultures in order to reveal the contribution of different pathways to the enhanced neurite outgrowth. We made the unexpected finding that inhibition of MEK signalling mimicked the potentiating effects of BMP stimulation in the chick system. The underlying mechanisms for the synergistic effects, however, are still an enigma.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2003. 75 p.
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1259
Neurosciences, Bone Morphogenetic Protein (BMP), Growth Differentiation Factor 10 (GDF10), Aktivne-receptor Like Kinase 2 (ALK2), Tyrosine Hydroxylase (TH), Neurotrophic Factor, Neurite Outgrowth, Hippocampus, Catecholamine, PC12, Sympathetic Ganglia, Substantia Nigra (SN), Gene Targeting, Neurovetenskap
National Category
Research subject
Developmental Neurosciences
urn:nbn:se:uu:diva-3398 (URN)91-554-5624-3 (ISBN)
Public defence
2003-05-16, B21, BMC, Uppsala, 09:15
Available from: 2003-04-24 Created: 2003-04-24 Last updated: 2013-03-22Bibliographically approved

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