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Quantitative and kinetic profile of Wnt/β-catenin signaling components during human neural progenitor cell differentiation

Abstract

ReNcell VM is an immortalized human neural progenitor cell line with the ability to differentiate in vitro into astrocytes and neurons, in which the Wnt/β-catenin pathway is known to be involved. However, little is known about kinetic changes of this pathway in human neural progenitor cell differentiation. In the present study, we provide a quantitative profile of Wnt/β-catenin pathway dynamics showing its spatio-temporal regulation during ReNcell VM cell differentiation. We show first that T-cell factor dependent transcription can be activated by stabilized β-catenin. Furthermore, endogenous Wnt ligands, pathway receptors and signaling molecules are temporally controlled, demonstrating changes related to differentiation stages. During the first three hours of differentiation the signaling molecules LRP6, Dvl2 and β-catenin are spatio-temporally regulated between distinct cellular compartments. From 24 h onward, components of the Wnt/β-catenin pathway are strongly activated and regulated as shown by mRNA up-regulation of Wnt ligands (Wnt5a and Wnt7a), receptors including Frizzled-2, -3, -6, -7, and -9, and co-receptors, and target genes including Axin2. This detailed temporal profile of the Wnt/β-catenin pathway is a first step to understand, control and to orientate, in vitro, human neural progenitor cell differentiation.

Abbreviations

APC:

adenomatous polyposis coli

bFGF:

basic fibroblast growth factor

Cdk:

cyclin-dependent kinase

CK1:

casein kinase 1

DAPI:

4′,6-diamidino-2-phenylindole

Dkk1:

Dickkopf 1

DMEM:

Dulbecco’s modified Eagle’s medium

Dvl:

dishevelled

EGF:

epidermal growth factor

Fz:

Frizzled

GAPDH:

glyceraldehyde 3-phosphate dehydrogenase

GFP:

green fluorescent protein

GSK3beta:

glycogen synthase kinase 3

HBSS:

Hank’s buffered salt solution

hNPC:

human neural progenitor cell

LRP6:

low-density lipoprotein receptor-related protein 6

MAP2:

microtubuleassociated protein 2

NPC:

neural progenitor cell

Ror2:

receptor tyrosine kinase-like orphan receptor 2

Ryk:

receptor-like tyrosine kinase

TCF:

T-cell factor

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Correspondence to Arndt Rolfs or Jiankai Luo.

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Both authors contributed equally to this work and should be considered co-first authors

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Mazemondet, O., Hubner, R., Frahm, J. et al. Quantitative and kinetic profile of Wnt/β-catenin signaling components during human neural progenitor cell differentiation. Cell Mol Biol Lett 16, 515 (2011). https://doi.org/10.2478/s11658-011-0021-0

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  • DOI: https://doi.org/10.2478/s11658-011-0021-0

Key words

  • Wnt/β-catenin pathway
  • Spatio-temporal dynamics
  • Quantitative kinetics