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Leucine-rich repeat kinase 2 modulates retinoic acid-induced neuronal differentiation of murine embryonic stem cells.

Schulz C, Paus M, Frey K, Schmid R, Kohl Z, Mennerich D, Winkler J, Gillardon F - PLoS ONE (2011)

Bottom Line: LRRK2 is expressed in neural precursor cells suggesting a role in neurodevelopment.By contrast, expression of neurotransmitter receptors and neurotransmitter release was increased in LRRK2+/- cultures indicating that LRRK2 promotes neuronal differentiation.Alterations in phosphorylation of the putative LRRK2 substrates, translation initiation factor 4E binding protein 1 and moesin, do not appear to be involved in altered differentiation, rather there is indirect evidence that a regulatory signaling network comprising retinoic acid receptors, let-7 miRNA and downstream target genes/mRNAs may be affected in LRRK2-deficient stem cells in culture.

View Article: PubMed Central - PubMed

Affiliation: Boehringer Ingelheim Pharma GmbH & Co KG, CNS Research, Biberach an der Riss, Germany.

ABSTRACT

Background: Dominant mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most prevalent cause of Parkinson's disease, however, little is known about the biological function of LRRK2 protein. LRRK2 is expressed in neural precursor cells suggesting a role in neurodevelopment.

Methodology/principal findings: In the present study, differential gene expression profiling revealed a faster silencing of pluripotency-associated genes, like Nanog, Oct4, and Lin28, during retinoic acid-induced neuronal differentiation of LRRK2-deficient mouse embryonic stem cells compared to wildtype cultures. By contrast, expression of neurotransmitter receptors and neurotransmitter release was increased in LRRK2+/- cultures indicating that LRRK2 promotes neuronal differentiation. Consistently, the number of neural progenitor cells was higher in the hippocampal dentate gyrus of adult LRRK2-deficient mice. Alterations in phosphorylation of the putative LRRK2 substrates, translation initiation factor 4E binding protein 1 and moesin, do not appear to be involved in altered differentiation, rather there is indirect evidence that a regulatory signaling network comprising retinoic acid receptors, let-7 miRNA and downstream target genes/mRNAs may be affected in LRRK2-deficient stem cells in culture.

Conclusion/significance: Parkinson's disease-linked LRRK2 mutations that associated with enhanced kinase activity may affect retinoic acid receptor signaling during neurodevelopment and/or neuronal maintenance as has been shown in other mouse models of chronic neurodegenerative diseases.

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Related in: MedlinePlus

Immunocytochemical and morphological analysis of LRRK2+/− (right panels) and wildtype (left panels) ES cells during neuronal differentiation.(A, B) Undifferentiated ES cells on fibroblast feeders are immunoreactive for the pluripotency marker stage-specific embryonic antigen-1. (C, D) Embryoid bodies grown in suspension culture show similar morphology. (E, F) At differentiation day 7, ES cell-derived neurons immunostained for β-III tubulin exhibit similar cell densities and neurite networks.
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pone-0020820-g002: Immunocytochemical and morphological analysis of LRRK2+/− (right panels) and wildtype (left panels) ES cells during neuronal differentiation.(A, B) Undifferentiated ES cells on fibroblast feeders are immunoreactive for the pluripotency marker stage-specific embryonic antigen-1. (C, D) Embryoid bodies grown in suspension culture show similar morphology. (E, F) At differentiation day 7, ES cell-derived neurons immunostained for β-III tubulin exhibit similar cell densities and neurite networks.

Mentions: Pluripotency of LRRK2+/− and wildtype ES cells was confirmed both by detection of high alkaline phosphatase activity [18] and by immunostaining for the surface marker stage-specific embryonic antigen-1 (SSEA-1) (Figure 2 A, B). During all steps of differentiation no obvious differences in cell density or morphology could be observed between LRRK2+/− and wildtype cells by microscopical imaging and digital analysis (Figure 2). Immunostaining for neuron-specific β-III tubulin and nuclear Hoechst 33342 labelling revealed that about 95% of both LRRK2+/− and wildtype cells adopted a neuronal phenotype after retinoic acid treatment for 7 days (Figure 2) [18]. Neuronally-differentiated ES cells cultured for >7 days exhibited neurite blebbing precluding longer observation times.


Leucine-rich repeat kinase 2 modulates retinoic acid-induced neuronal differentiation of murine embryonic stem cells.

Schulz C, Paus M, Frey K, Schmid R, Kohl Z, Mennerich D, Winkler J, Gillardon F - PLoS ONE (2011)

Immunocytochemical and morphological analysis of LRRK2+/− (right panels) and wildtype (left panels) ES cells during neuronal differentiation.(A, B) Undifferentiated ES cells on fibroblast feeders are immunoreactive for the pluripotency marker stage-specific embryonic antigen-1. (C, D) Embryoid bodies grown in suspension culture show similar morphology. (E, F) At differentiation day 7, ES cell-derived neurons immunostained for β-III tubulin exhibit similar cell densities and neurite networks.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3111438&req=5

pone-0020820-g002: Immunocytochemical and morphological analysis of LRRK2+/− (right panels) and wildtype (left panels) ES cells during neuronal differentiation.(A, B) Undifferentiated ES cells on fibroblast feeders are immunoreactive for the pluripotency marker stage-specific embryonic antigen-1. (C, D) Embryoid bodies grown in suspension culture show similar morphology. (E, F) At differentiation day 7, ES cell-derived neurons immunostained for β-III tubulin exhibit similar cell densities and neurite networks.
Mentions: Pluripotency of LRRK2+/− and wildtype ES cells was confirmed both by detection of high alkaline phosphatase activity [18] and by immunostaining for the surface marker stage-specific embryonic antigen-1 (SSEA-1) (Figure 2 A, B). During all steps of differentiation no obvious differences in cell density or morphology could be observed between LRRK2+/− and wildtype cells by microscopical imaging and digital analysis (Figure 2). Immunostaining for neuron-specific β-III tubulin and nuclear Hoechst 33342 labelling revealed that about 95% of both LRRK2+/− and wildtype cells adopted a neuronal phenotype after retinoic acid treatment for 7 days (Figure 2) [18]. Neuronally-differentiated ES cells cultured for >7 days exhibited neurite blebbing precluding longer observation times.

Bottom Line: LRRK2 is expressed in neural precursor cells suggesting a role in neurodevelopment.By contrast, expression of neurotransmitter receptors and neurotransmitter release was increased in LRRK2+/- cultures indicating that LRRK2 promotes neuronal differentiation.Alterations in phosphorylation of the putative LRRK2 substrates, translation initiation factor 4E binding protein 1 and moesin, do not appear to be involved in altered differentiation, rather there is indirect evidence that a regulatory signaling network comprising retinoic acid receptors, let-7 miRNA and downstream target genes/mRNAs may be affected in LRRK2-deficient stem cells in culture.

View Article: PubMed Central - PubMed

Affiliation: Boehringer Ingelheim Pharma GmbH & Co KG, CNS Research, Biberach an der Riss, Germany.

ABSTRACT

Background: Dominant mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most prevalent cause of Parkinson's disease, however, little is known about the biological function of LRRK2 protein. LRRK2 is expressed in neural precursor cells suggesting a role in neurodevelopment.

Methodology/principal findings: In the present study, differential gene expression profiling revealed a faster silencing of pluripotency-associated genes, like Nanog, Oct4, and Lin28, during retinoic acid-induced neuronal differentiation of LRRK2-deficient mouse embryonic stem cells compared to wildtype cultures. By contrast, expression of neurotransmitter receptors and neurotransmitter release was increased in LRRK2+/- cultures indicating that LRRK2 promotes neuronal differentiation. Consistently, the number of neural progenitor cells was higher in the hippocampal dentate gyrus of adult LRRK2-deficient mice. Alterations in phosphorylation of the putative LRRK2 substrates, translation initiation factor 4E binding protein 1 and moesin, do not appear to be involved in altered differentiation, rather there is indirect evidence that a regulatory signaling network comprising retinoic acid receptors, let-7 miRNA and downstream target genes/mRNAs may be affected in LRRK2-deficient stem cells in culture.

Conclusion/significance: Parkinson's disease-linked LRRK2 mutations that associated with enhanced kinase activity may affect retinoic acid receptor signaling during neurodevelopment and/or neuronal maintenance as has been shown in other mouse models of chronic neurodegenerative diseases.

Show MeSH
Related in: MedlinePlus