Activation of MAPKs by 1α,25(OH)2-Vitamin D3 and 17β-estradiol in skeletal muscle cells leads to phosphorylation of Elk-1 and CREB transcription factors

doi:10.1016/j.jsbmb.2006.11.005

The Journal of Steroid Biochemistry and Molecular Biology

Volume 103, Issues 3–5, March 2007, Pages 462-466

https://doi.org/10.1016/j.jsbmb.2006.11.005 Get rights and content

Abstract

The mitogen activated protein kinases (MAPKs) have been classified into at least six subfamilies, among which ERK1/2, JNK1/2 and p38 MAPK are the most extensively studied. The steroid hormones 1α,25-dihydroxy-Vitamin D₃ and 17β-estradiol promote biological responses through activation of MAPK cascades in various cell types. We previously reported that 1α,25(OH)₂D₃ rapidly (within 1 min) activates p38 MAPK in C2C12 skeletal muscle cells. In this work, using the same muscle cell line, we demonstrate that 1α,25(OH)₂D₃ or 17β-estradiol phosphorylate and activate ERK1/2 and p38 MAPK after longer treatment intervals, maximal effects seen at 90 and 30 min (ERK1/2) and at 60 and 15 min (p38 MAPK) for these hormones, respectively. Hormone-dependent ERK and p38 activation was abolished by MAPK specific inhibitors U0126 and SB203580. 1α,25(OH)₂D₃ and 17β-estradiol also induced the phosphorylation of CREB and Elk-1 transcription factors in an ERK1/2-dependent manner. Simultaneous addition of both hormones potentiated CREB phosphorylation. 1α,25(OH)₂D₃- and 17β-estradiol-induced c-fos expression, which was mediated by p38 phosphorylation. The action of 17β-estradiol on c-fos levels was also dependent on ERK1/2. These results suggest that MAPK signalling pathways play an important role in regulating early gene expression through CREB and Elk-1 activation in skeletal muscle cells.

Introduction

1α,25-Dihydroxy-Vitamin D₃ [1α,25(OH)₂D₃] and 17β-estradiol play an important role in the regulation of cellular proliferation and differentiation [1], [2]. 1α,25(OH)₂D₃ also regulates skeletal muscle contractility and growth [3] and 17β-estradiol modulates multiple signal inputs to prevent cardiomyocyte hypertrophy in vitro [4]. Besides regulating gene expression via the specific intracellular receptors [1], [2], both hormones also exert in their target tissues fast non-transcriptional responses involving stimulation of transmembrane signal transduction pathways [5], [6], [7]. Recent evidence indicates that modulation of various responses to 1α,25-(OH)₂D₃ and 17β-estradiol depend on the fast activation of MAP kinase pathways. Thus, in proliferating cultured myoblasts, 1α,25(OH)₂D₃ rapidly (within 1 min) activates ERK-1/2 [8]. In other cell types, estrogens are able to activate ERK 1/2, JNK and p38 MAPK [9], [10], [11], [12], [13]. Moreover, the Ras-MAPK cascade modulates the activity of the amino-terminal AF-1 of the estrogen receptor (ER) by the phosphorylation of Ser118 [14].

1α,25(OH)₂D₃ plays a role in the regulation of muscle cell proliferation and differentiation [15], stimulating thereby muscle growth. It has been recently established that skeletal muscle contains receptors for 17β-estradiol (ER α and β) [16], [17], congruent with the anabolic action of estrogens in this tissue [18], [19]. In view of these evidences, in the present work we have investigated the effects of 1α,25(OH)₂D₃ and of 17β-estradiol on the ERK1/2 and p38 MAPK cascades using the mouse C2C12 myoblast cell line as an in vitro model for skeletal muscle.

Section snippets

Materials

1α,25(OH)₂D₃ was kindly provided by Dr. Jan-Paul van de Velde from Solvay Pharmaceuticals (Weesp, The Netherlands). 17β-Estradiol, Dulbecco's modified Eagle's medium (DMEM) and fetal bovine serum (FBS) were from Sigma Chemical Co. (St. Louis, MO, USA). Free-phenol red DMEM was from USBiological Inc. (Massachusetts, MA, USA). Anti-p38 α, anti-38 β, anti-phospho p38, anti-phospho CREB, anti-CREB and anti-c-Fos antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Results and discussion

Within the mitogen-activated protein (MAP) kinase family four different subgroups have been described. These include: extracellular signal-regulated kinases (ERKs), c-Jun NH₂-terminal or stress-activated protein kinases (JNK/SAPK), big MAP kinase 1 (BMK1), also called ERK5, and p38 MAP kinases (p38). The ERK, JNK and p38 MAPK cascades consist of three protein kinases (MAPKKK, MAPKK, MAPK) that sequentially activate each other by phosphorylation [21]. The ERK pathway is activated by mitogens and

Acknowledgements

This research was supported by grants from the Agencia Nacional de Promocion Cientifica y Tecnologica, Consejo Nacional de Investigaciones Cientificas y Técnicas (CONICET), and Universidad Nacional del Sur, Argentina.

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Citation Excerpt :
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Activation of MAPKs by 1α,25(OH)2-Vitamin D3 and 17β-estradiol in skeletal muscle cells leads to phosphorylation of Elk-1 and CREB transcription factors

Abstract

Introduction

Section snippets

Materials

Results and discussion

Acknowledgements

J. Biol. Chem.

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Trends Endocrinol. Metab.

Mol. Cell. Endocrinol.

Eur. J. Pharmacol.

Biochem. Biophys. Res. Commun.

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J. Biol. Chem.

J. Invest. Dermatol.

J. Biol. Chem.

Biochem. Biophys. Res. Commun.

Gene

J. Invest. Dermatol.

Mol. Cell. Endocrinol.

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Endocr. Rev.

Activation of MAPKs by 1α,25(OH)₂-Vitamin D₃ and 17β-estradiol in skeletal muscle cells leads to phosphorylation of Elk-1 and CREB transcription factors