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The kinase MSK1 is required for induction of c-fos by lysophosphatidic acid in mouse embryonic stem cells.

Schuck S, Soloaga A, Schratt G, Arthur JS, Nordheim A - BMC Mol. Biol. (2003)

Bottom Line: We propose that MSK1 contributes to ERK-mediated c-fos promoter activation by targeting CRE binding proteins.In addition, they indicate that MSK1 could act through CRE binding proteins to achieve c-fos promoter activation.Thus, they further our understanding of the complex regulation of the model immediate-early gene c-fos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interfakultäres Institut für Zellbiologie, Abteilung Molekularbiologie, Universität Tübingen, Auf der Morgenstelle 15, 72076 Tübingen, Germany. schuck@mpi-cbg.de

ABSTRACT

Background: The regulation of the immediate-early gene c-fos serves as a paradigm for signal-activated gene induction. Lysophosphatidic acid is a potent serum-borne mitogen able to induce c-fos.

Results: Analysing the signalling events following stimulation of mouse embryonic stem cells with serum and lysophosphatidic acid, we show that the extracellular signal-regulated kinase (ERK) pathway is involved in mediating c-fos induction. We demonstrate that the ERK-activated kinase MSK1 is required for full c-fos promoter activation, as well as for the phosphorylation of cAMP-responsive element (CRE) binding proteins. We propose that MSK1 contributes to ERK-mediated c-fos promoter activation by targeting CRE binding proteins.

Conclusion: These results show that MSK1 is an important ERK-activated mediator of mitogen-stimulated c-fos induction. In addition, they indicate that MSK1 could act through CRE binding proteins to achieve c-fos promoter activation. Thus, they further our understanding of the complex regulation of the model immediate-early gene c-fos.

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Induction of c-fos in WT and Msk1(-/-) ES cells. A – WT ES cells and three different Msk1(-/-) ES cell lines (No. 2, No. 4 and No. 5) were serum-starved for 24 h and stimulated with 15% FCS or 20 μM LPA for 30 min, followed by quantitative RT-PCR. Based on relative mRNA levels, fold inductions were calculated from five independent experiments. B – WT and Msk1(-/-) ES cells were serum-starved overnight and stimulated with 20 μM LPA for the times indicated. C-fos mRNA was amplified by semi-quantitative RT-PCR. PCR products were separated on agarose gels and stained with ethidium bromide (EtBr). EtBr intensities were used to calculate c-fos induction.
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Figure 4: Induction of c-fos in WT and Msk1(-/-) ES cells. A – WT ES cells and three different Msk1(-/-) ES cell lines (No. 2, No. 4 and No. 5) were serum-starved for 24 h and stimulated with 15% FCS or 20 μM LPA for 30 min, followed by quantitative RT-PCR. Based on relative mRNA levels, fold inductions were calculated from five independent experiments. B – WT and Msk1(-/-) ES cells were serum-starved overnight and stimulated with 20 μM LPA for the times indicated. C-fos mRNA was amplified by semi-quantitative RT-PCR. PCR products were separated on agarose gels and stained with ethidium bromide (EtBr). EtBr intensities were used to calculate c-fos induction.

Mentions: Msk1(-/-) ES cells [16] allowed direct assessment of the role of MSK1 in c-fos induction. Figure 4a shows that c-fos induction as measured by quantitative RT-PCR was reduced by at least 50% for both stimuli in three different Msk1(-/-) ES cell lines. This was confirmed by RNase protection assays (data not shown). The reduced c-fos induction at 30 min of stimulation could be due to a weaker promoter activation or due to a delayed response. To distinguish between these explanations, induction kinetics of c-fos in wild type and Msk1(-/-) ES cells were measured by semi-quantitative RT-PCR (Figure 4b). The time-course of c-fos induction by LPA in Msk1(-/-) ES cells was identical to that in wild type cells, excluding that disruption of the msk1 gene leads to a delayed c-fos induction. Hence, our data constitute direct genetic evidence that MSK1 is required for the activation of the c-fos promoter.


The kinase MSK1 is required for induction of c-fos by lysophosphatidic acid in mouse embryonic stem cells.

Schuck S, Soloaga A, Schratt G, Arthur JS, Nordheim A - BMC Mol. Biol. (2003)

Induction of c-fos in WT and Msk1(-/-) ES cells. A – WT ES cells and three different Msk1(-/-) ES cell lines (No. 2, No. 4 and No. 5) were serum-starved for 24 h and stimulated with 15% FCS or 20 μM LPA for 30 min, followed by quantitative RT-PCR. Based on relative mRNA levels, fold inductions were calculated from five independent experiments. B – WT and Msk1(-/-) ES cells were serum-starved overnight and stimulated with 20 μM LPA for the times indicated. C-fos mRNA was amplified by semi-quantitative RT-PCR. PCR products were separated on agarose gels and stained with ethidium bromide (EtBr). EtBr intensities were used to calculate c-fos induction.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC161794&req=5

Figure 4: Induction of c-fos in WT and Msk1(-/-) ES cells. A – WT ES cells and three different Msk1(-/-) ES cell lines (No. 2, No. 4 and No. 5) were serum-starved for 24 h and stimulated with 15% FCS or 20 μM LPA for 30 min, followed by quantitative RT-PCR. Based on relative mRNA levels, fold inductions were calculated from five independent experiments. B – WT and Msk1(-/-) ES cells were serum-starved overnight and stimulated with 20 μM LPA for the times indicated. C-fos mRNA was amplified by semi-quantitative RT-PCR. PCR products were separated on agarose gels and stained with ethidium bromide (EtBr). EtBr intensities were used to calculate c-fos induction.
Mentions: Msk1(-/-) ES cells [16] allowed direct assessment of the role of MSK1 in c-fos induction. Figure 4a shows that c-fos induction as measured by quantitative RT-PCR was reduced by at least 50% for both stimuli in three different Msk1(-/-) ES cell lines. This was confirmed by RNase protection assays (data not shown). The reduced c-fos induction at 30 min of stimulation could be due to a weaker promoter activation or due to a delayed response. To distinguish between these explanations, induction kinetics of c-fos in wild type and Msk1(-/-) ES cells were measured by semi-quantitative RT-PCR (Figure 4b). The time-course of c-fos induction by LPA in Msk1(-/-) ES cells was identical to that in wild type cells, excluding that disruption of the msk1 gene leads to a delayed c-fos induction. Hence, our data constitute direct genetic evidence that MSK1 is required for the activation of the c-fos promoter.

Bottom Line: We propose that MSK1 contributes to ERK-mediated c-fos promoter activation by targeting CRE binding proteins.In addition, they indicate that MSK1 could act through CRE binding proteins to achieve c-fos promoter activation.Thus, they further our understanding of the complex regulation of the model immediate-early gene c-fos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Interfakultäres Institut für Zellbiologie, Abteilung Molekularbiologie, Universität Tübingen, Auf der Morgenstelle 15, 72076 Tübingen, Germany. schuck@mpi-cbg.de

ABSTRACT

Background: The regulation of the immediate-early gene c-fos serves as a paradigm for signal-activated gene induction. Lysophosphatidic acid is a potent serum-borne mitogen able to induce c-fos.

Results: Analysing the signalling events following stimulation of mouse embryonic stem cells with serum and lysophosphatidic acid, we show that the extracellular signal-regulated kinase (ERK) pathway is involved in mediating c-fos induction. We demonstrate that the ERK-activated kinase MSK1 is required for full c-fos promoter activation, as well as for the phosphorylation of cAMP-responsive element (CRE) binding proteins. We propose that MSK1 contributes to ERK-mediated c-fos promoter activation by targeting CRE binding proteins.

Conclusion: These results show that MSK1 is an important ERK-activated mediator of mitogen-stimulated c-fos induction. In addition, they indicate that MSK1 could act through CRE binding proteins to achieve c-fos promoter activation. Thus, they further our understanding of the complex regulation of the model immediate-early gene c-fos.

Show MeSH