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Vitamin D3 signalling in the brain enhances the function of phosphoprotein enriched in astrocytes--15 kD (PEA-15).

Obradovic D, Zanca C, Vogl A, Trümbach D, Deussing J, Condorelli G, Rein T - J. Cell. Mol. Med. (2009)

Bottom Line: PEA-15 (phosphoprotein enriched in astrocytes-15 kD, also known as PED), known to be involved in various anti-proliferative and anti-apoptotic effects, was strongly up-regulated.PEA-15 up-regulation by vitamin D(3) could be confirmed by Western blot in two different cell lines.In a functional test of this novel pathway, we demonstrated that vitamin D(3) was able to rescue cells from TRAIL-induced apoptosis through regulation of the PEA-15 expression and function.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Psychiatry, Munich, Germany.

ABSTRACT
In spite of growing evidence linking vitamin D(3) levels to mental health disorders, little is known about its direct targets in the brain. This study set out to investigate targets of vitamin D(3) in a human brain stem cell line. We employed arrays with antibodies directed against more than 600 structural and signalling proteins, including phospho-variants. Over 180 proteins responded to vitamin D(3), such as cyclin-dependent protein-serine kinase 1/2, epidermal growth factor receptor-tyrosine kinase, protein kinase A, protein-serine kinase Bgamma and protein-serine kinase Calpha. PEA-15 (phosphoprotein enriched in astrocytes-15 kD, also known as PED), known to be involved in various anti-proliferative and anti-apoptotic effects, was strongly up-regulated. In silico promoter analysis revealed conserved binding sites for vitamin D(3) receptor, suggesting a strong vitamin D(3) dependency of the PEA-15 promoter. PEA-15 up-regulation by vitamin D(3) could be confirmed by Western blot in two different cell lines. Analysis of mRNA and protein phosphorylation status of PEA-15 suggests that increased PEA-15 promoter activity and increased protein stabilization contribute to the overall rise of PEA-15 protein. In a functional test of this novel pathway, we demonstrated that vitamin D(3) was able to rescue cells from TRAIL-induced apoptosis through regulation of the PEA-15 expression and function. Summarized, our study presents novel targets of vitamin D(3) relevant for apoptosis and cell proliferation, and thus strongly supports a function of vitamin D(3) in the brain that impacts on processes highly relevant for major neurological disorders.

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Related in: MedlinePlus

Conserved transcription factor binding sites in the promoter of PEA-15 and up-regulation of PEA-15 mRNA and protein. (A) Several transcription factor binding motifs were identified, including those for VDR, NFκB, EGRF, EKLF and SP1. (B) Effect of vitamin D3 on PEA-15 mRNA levels. A549 cells were treated with vitamin D3 (10−7 M), and the mRNA levels were determined by real-time PCR at the times indicated. (C) Verification of induction of total PEA-15 after 12 hrs by Western blot.
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fig04: Conserved transcription factor binding sites in the promoter of PEA-15 and up-regulation of PEA-15 mRNA and protein. (A) Several transcription factor binding motifs were identified, including those for VDR, NFκB, EGRF, EKLF and SP1. (B) Effect of vitamin D3 on PEA-15 mRNA levels. A549 cells were treated with vitamin D3 (10−7 M), and the mRNA levels were determined by real-time PCR at the times indicated. (C) Verification of induction of total PEA-15 after 12 hrs by Western blot.

Mentions: For the Western blots in Figs 4–6, cells were lysed as described earlier. Fifty micrograms of cell extract were resolved on 15% SDS–polyacrylamide and transferred to Hybond-C extra nitrocellulose. Membranes were blocked for 1 hr with 5% non-fat dry milk in TBS containing 0.05% Tween-20 incubated with primary antibodies (directed against: total AKT, pAKT (S473), total PEA-15, pPEA-15 (S116) and β-actin as indicated in the figures) and visualized by chemiluminescence [28].


Vitamin D3 signalling in the brain enhances the function of phosphoprotein enriched in astrocytes--15 kD (PEA-15).

Obradovic D, Zanca C, Vogl A, Trümbach D, Deussing J, Condorelli G, Rein T - J. Cell. Mol. Med. (2009)

Conserved transcription factor binding sites in the promoter of PEA-15 and up-regulation of PEA-15 mRNA and protein. (A) Several transcription factor binding motifs were identified, including those for VDR, NFκB, EGRF, EKLF and SP1. (B) Effect of vitamin D3 on PEA-15 mRNA levels. A549 cells were treated with vitamin D3 (10−7 M), and the mRNA levels were determined by real-time PCR at the times indicated. (C) Verification of induction of total PEA-15 after 12 hrs by Western blot.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4516488&req=5

fig04: Conserved transcription factor binding sites in the promoter of PEA-15 and up-regulation of PEA-15 mRNA and protein. (A) Several transcription factor binding motifs were identified, including those for VDR, NFκB, EGRF, EKLF and SP1. (B) Effect of vitamin D3 on PEA-15 mRNA levels. A549 cells were treated with vitamin D3 (10−7 M), and the mRNA levels were determined by real-time PCR at the times indicated. (C) Verification of induction of total PEA-15 after 12 hrs by Western blot.
Mentions: For the Western blots in Figs 4–6, cells were lysed as described earlier. Fifty micrograms of cell extract were resolved on 15% SDS–polyacrylamide and transferred to Hybond-C extra nitrocellulose. Membranes were blocked for 1 hr with 5% non-fat dry milk in TBS containing 0.05% Tween-20 incubated with primary antibodies (directed against: total AKT, pAKT (S473), total PEA-15, pPEA-15 (S116) and β-actin as indicated in the figures) and visualized by chemiluminescence [28].

Bottom Line: PEA-15 (phosphoprotein enriched in astrocytes-15 kD, also known as PED), known to be involved in various anti-proliferative and anti-apoptotic effects, was strongly up-regulated.PEA-15 up-regulation by vitamin D(3) could be confirmed by Western blot in two different cell lines.In a functional test of this novel pathway, we demonstrated that vitamin D(3) was able to rescue cells from TRAIL-induced apoptosis through regulation of the PEA-15 expression and function.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Psychiatry, Munich, Germany.

ABSTRACT
In spite of growing evidence linking vitamin D(3) levels to mental health disorders, little is known about its direct targets in the brain. This study set out to investigate targets of vitamin D(3) in a human brain stem cell line. We employed arrays with antibodies directed against more than 600 structural and signalling proteins, including phospho-variants. Over 180 proteins responded to vitamin D(3), such as cyclin-dependent protein-serine kinase 1/2, epidermal growth factor receptor-tyrosine kinase, protein kinase A, protein-serine kinase Bgamma and protein-serine kinase Calpha. PEA-15 (phosphoprotein enriched in astrocytes-15 kD, also known as PED), known to be involved in various anti-proliferative and anti-apoptotic effects, was strongly up-regulated. In silico promoter analysis revealed conserved binding sites for vitamin D(3) receptor, suggesting a strong vitamin D(3) dependency of the PEA-15 promoter. PEA-15 up-regulation by vitamin D(3) could be confirmed by Western blot in two different cell lines. Analysis of mRNA and protein phosphorylation status of PEA-15 suggests that increased PEA-15 promoter activity and increased protein stabilization contribute to the overall rise of PEA-15 protein. In a functional test of this novel pathway, we demonstrated that vitamin D(3) was able to rescue cells from TRAIL-induced apoptosis through regulation of the PEA-15 expression and function. Summarized, our study presents novel targets of vitamin D(3) relevant for apoptosis and cell proliferation, and thus strongly supports a function of vitamin D(3) in the brain that impacts on processes highly relevant for major neurological disorders.

Show MeSH
Related in: MedlinePlus