The Mitogen-activated Protein Kinase Kinase MEK1 Stimulates a Pattern of Gene Expression Typical of the Hypertrophic Phenotype in Rat Ventricular Cardiomyocytes

The Raf protein kinases are major effectors of Ras GTPases and key components of the transcriptional response to serum factors, acting at least in part through the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway. It has recently been suggested that Raf also may trigger other as yet uncharacterized signaling pathways. Here, we have used cDNA microarrays to dissect changes in gene expression induced by activation of inducible c-Raf-1 constructs in human mammary epithelial and ovarian epithelial cells. The majority of Raf-induced transcriptional responses are shown to be blocked by pharmacological inhibition of the Raf substrate mitogen-activated protein kinase kinase, indicating that potential mitogen-activated protein kinase kinase-independent Raf signaling pathways have no significant influence on gene expression. In addition, we used epidermal growth factor receptor inhibitory drugs to address the contribution of autocrine signaling by Raf-induced EGF family proteins to the Raf transcriptional response. At least one-half of the transcription induced by Raf activation requires epidermal growth factor (EGF) receptor function The EGF receptor-independent component of the Raf transcriptional response is entirely up-regulation of gene expression, whereas the EGF receptor-dependent component is an equal mixture of up- and down-regulation. The use of transcriptional profiling in this way allows detailed analysis of the architecture of signaling pathways to be undertaken.

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Inhibition of a signaling pathway in cardiac muscle cells by active mitogen-activated protein kinase kinase.

Molecular Biology of the Cell ◽

10.1091/mbc.6.11.1479 ◽

1995 ◽

Vol 6 (11) ◽

pp. 1479-1490 ◽

Cited By ~ 56

Author(s):

J Thorburn ◽

M Carlson ◽

S J Mansour ◽

K R Chien ◽

N G Ahn ◽

...

Keyword(s):

Gene Expression ◽

Protein Kinase ◽

Cardiac Muscle ◽

Map Kinases ◽

Mitogen Activated Protein Kinase ◽

Cardiac Muscle Cells ◽

Erk Activation ◽

Mitogen Activated Protein ◽

Protein Kinase Kinase ◽

Constitutively Active

Signaling via the Ras pathway involves sequential activation of Ras, Raf-1, mitogen-activated protein kinase kinase (MKK), and the extracellular signal-regulated (ERK) group of mitogen-activated protein (MAP) kinases. Expression from the c-Fos, atrial natriuretic factor (ANF), and myosin light chain-2 (MLC-2) promoters during phenylephrine-induced cardiac muscle cell hypertrophy requires activation of this pathway. Furthermore, constitutively active Ras or Raf-1 can mimic the action of phenylephrine in inducing expression from these promoters. In this study, we tested whether constitutively active MKK, the molecule immediately downstream of Raf, was sufficient to induce expression. Expression of constitutively active MKK induce ERK2 kinase activity and caused expression from the c-Fos promoter, but did not significantly activate expression of reporter genes under the control of either the ANF or MLC-2 promoters. Expression of CL100, a phosphatase that inactivates ERKs, prevented expression from all of the promoters. Taken together, these data suggest that ERK activation is required for expression from the Fos, ANF, and MLC-2 promoters but MKK and ERK activation is sufficient for expression only from the Fos promoter. Constitutively active MKK synergized with phenylephrine to increase expression from a c-Fos- or an AP1-driven reporter. However, active MKK inhibited phenylephrine- and Raf-1-induced expression from the ANF and MLC-2 promoters. A DNA sequence in the MLC-2 promoter that is a target for inhibition by active MKK, but not CL100, was mapped to a previously characterized DNA element (HF1) that is responsible for cardiac specificity. Thus, activation of cardiac gene expression during phenylephrine-induced hypertrophy requires ERK activation but constitutive activation by MKK can inhibit expression by targeting a DNA element that controls the cardiac specificity of gene expression.

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