Early specification of dopaminergic phenotype during ES cell differentiation.
BACKGROUND
Understanding how lineage choices are made during embryonic stem (ES) cell differentiation is critical for harnessing strategies for controlled production of therapeutic somatic cell types for cell transplantation and pharmaceutical drug screens. The in vitro generation of dopaminergic neurons, the type of cells lost in Parkinson's disease patients' brains, requires the inductive molecules sonic hedgehog and FGF8, or an unknown stromal cell derived inducing activity (SDIA). However, the exact identity of the responding cells and the timing of inductive activity that specify a dopaminergic fate in neural stem/progenitors still remain elusive.


RESULTS
Using ES cells carrying a neuroepithelial cell specific vital reporter (Sox1-GFP) and FACS purification of Sox1-GFP neural progenitors, we have investigated the temporal aspect of SDIA mediated dopaminergic neuron specification during ES cell differentiation. Our results establish that SDIA induces a dopaminergic neuron fate in nascent neural stem or progenitor cells at, or prior to, Sox1 expression and does not appear to have further instructive role or neurotrophic activity during late neuronal differentiation of neural precursors. Furthermore, we show that dopaminergic neurons could be produced efficiently in a monolayer differentiation paradigm independent of SDIA activity or exogenous signalling molecules. In this case, the competence for dopaminergic neuron differentiation is also established at the level of Sox1 expression.


CONCLUSIONS
Dopaminergic neurons are specified early during mouse ES cell differentiation. The subtype specification seems to be tightly linked with the acquisition of a pan neuroectoderm fate.
Parmar
Malin
M
Institute for Stem Cell Research, University of Edinburgh, Edinburgh, UK. Malin.Parmar@med.lu.se
Li
Meng
M
D000818:Animals; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D019070:Cell Lineage; D002469:Cell Separation; D018920:Coculture Techniques; D004268:DNA-Binding Proteins / Q000235:genetics; D004298:Dopamine / Q000502:physiology*; D053595:Embryonic Stem Cells / Q000166:cytology*; D005434:Flow Cytometry; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D005819:Genetic Markers; D049452:Green Fluorescent Proteins / Q000235:genetics; D006609:High Mobility Group Proteins / Q000235:genetics; D006801:Humans; D051379:Mice; D009474:Neurons / Q000166:cytology*; D055748:SOXB1 Transcription Factors
D004268:DNA-Binding Proteins; D005819:Genetic Markers; D006609:High Mobility Group Proteins; D055748:SOXB1 Transcription Factors; C098693:Sox1 protein, mouse; D049452:Green Fluorescent Proteins; D004298:Dopamine