Phosphorylation of Ago2 and Subsequent Inactivation of let-7a RNP-Specific MicroRNAs Control Differentiation of Mammalian Sympathetic Neurons

MicroRNAs (miRNAs) are small regulatory RNAs that regulate gene expression posttranscriptionally by base pairing to the target mRNAs in animal cells.KRas, an oncogene known to be repressed by let-7a miRNAs, is expressed and needed for the differentiation of mammalian sympathetic neurons and PC12 cells. We documented a loss of let-7a activity during this differentiation process without any significant change in the cellular level of let-7a miRNA. However, the level of Ago2, an essential component that is associated with miRNAs to form RNP-specific miRNA (miRNP) complexes, shows an increase with neuronal differentiation. In this study, differentiation-induced phosphorylation and the subsequent loss of miRNA from Ago2 were noted, and these accounted for the loss of miRNA activity in differentiating neurons. Neuronal differentiation induces the phosphorylation of mitogen-activated protein kinase p38 and the downstream kinase mitogen- and stress-activated protein kinase 1 (MSK1). This in turn upregulates the phosphorylation of Ago2 and ensures the dissociation of miRNA from Ago2 in neuronal cells. MSK1-mediated miRNP inactivation is a prerequisite for the differentiation of neuronal cells, where let-7a miRNA gets unloaded from Ago2 to ensure the upregulation ofKRas, a target of let-7a. We noted that the inactivation of let-7a is both necessary and sufficient for the differentiation of sympathetic neurons.

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Initiation Factor 2B Activity Is Regulated by Protein Phosphatase 1, Which Is Activated by the Mitogen-activated Protein Kinase-dependent Pathway in Insulin-like Growth Factor 1-stimulated Neuronal Cells

Journal of Biological Chemistry ◽

10.1074/jbc.m212936200 ◽

2003 ◽

Vol 278 (19) ◽

pp. 16579-16586 ◽

Cited By ~ 20

Author(s):

Celia Quevedo ◽

Matilde Salinas ◽

Alberto Alcázar

Keyword(s):

Growth Factor ◽

Protein Kinase ◽

Protein Phosphatase ◽

Neuronal Cells ◽

Mitogen Activated Protein Kinase ◽

Protein Phosphatase 1 ◽

Initiation Factor ◽

Insulin Like Growth Factor ◽

Mitogen Activated Protein ◽

Dependent Pathway

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Involvement of Ras/MAP Kinase in the Regulation of Ca2+ Channels in Adult Bullfrog Sympathetic Neurons by Nerve Growth Factor

Journal of Neurophysiology ◽

10.1152/jn.1998.80.3.1352 ◽

1998 ◽

Vol 80 (3) ◽

pp. 1352-1361 ◽

Cited By ~ 21

Author(s):

Saobo Lei ◽

William F. Dryden ◽

Peter A. Smith

Keyword(s):

Nerve Growth Factor ◽

Growth Factor ◽

Protein Kinase ◽

Map Kinase ◽

Kinase Inhibitors ◽

Sympathetic Neurons ◽

Mitogen Activated Protein Kinase ◽

Nerve Growth ◽

Channel Current ◽

Mitogen Activated Protein

Lei, Saobo, William F. Dryden, and Peter A. Smith. Involvement of Ras/MAP kinase in the regulation of Ca2+ channels in adult bullfrog sympathetic neurons by nerve growth factor. J. Neurophysiol. 80: 1352–1361, 1998. The cellular mechanisms that underlie nerve growth factor (NGF) induced increase in Ca2+-channel current in adult bullfrog sympathetic B-neurons were examined by whole cell recording techniques. Cells were maintained at low density in neuron-enriched, defined-medium, serum-free tissue culture for 6 days in the presence or absence of NGF (200 ng/ml). The increase in Ba2+ current ( I Ba) density induced by NGF was attenuated by the RNA synthesis inhibitor cordycepin (20 μM), by the DNA transcription inhibitor actinomycin D (0.01 μg/ml), by inhibitors of Ras isoprenylation (perillic acid 0.1–1.0 mM or α-hydroxyfarnesylphosphonic acid 10–100 μM), by tyrosine kinase inhibitors genistein (20 μM) or lavendustin A (1 μM), and by PD98059 (10–100 μM), an inhibitor of mitogen-activated protein kinase kinase. Inhibitors of the phosphatidylinositol 3-kinase (PI3K) pathway (wortmannin, 100 nM, or LY29400, 100 μM) were ineffective as were inhibitors of phospholipase Cγ (U73122 or neomycin, both 100 μM). The effect of NGF persisted in Ca2+-free medium that contained 1.8 mM Mg2+ and 2 mM ethylene glycol-bis(β-aminoethyl ether)- N, N, N′, N′-tetraacetic acid. It was mimicked by a Trk antibody that was capable of inducing neurite outgrowth in explant cultures of bullfrog sympathetic ganglion. Antibodies raised against the low-affinity p75 neurotrophin receptor were ineffective in blocking the effect of NGF on I Ba. These results suggest that NGF-induced increase in Ca2+ channel current in adult sympathetic neurons results, at least in part, from new channel synthesis after Trk activation of Ras and mitogen activated protein kinase by a mechanism that is independent of extracellular Ca2+.

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Identification of Cdc37 as a Novel Regulator of the Stress-Responsive Mitogen-Activated Protein Kinase

Molecular and Cellular Biology ◽

10.1128/mcb.23.15.5132-5142.2003 ◽

2003 ◽

Vol 23 (15) ◽

pp. 5132-5142 ◽

Cited By ~ 34

Author(s):

Hisashi Tatebe ◽

Kazuhiro Shiozaki

Keyword(s):

Protein Kinase ◽

Protein Kinases ◽

Inflammatory Responses ◽

Cellular Level ◽

Mitogen Activated Protein Kinase ◽

Cellular Survival ◽

Evolutionarily Conserved ◽

Mitogen Activated Protein ◽

Extracellular Stimuli ◽

The Stability

ABSTRACT Eukaryotic cells utilize multiple mitogen-activated protein kinases (MAPKs) to transmit various extracellular stimuli to the nucleus. A subfamily of MAPKs that mediates environmental stress stimuli is also called stress-activated protein kinase (SAPK), which has crucial roles in cellular survival under stress conditions as well as inflammatory responses. Here we report that Cdc37, an evolutionarily conserved kinase-specific chaperone, is a positive regulator of Spc1 SAPK in the fission yeast Schizosaccharomyces pombe. Through a genetic screen, we have identified cdc37 as a mutation that compromises signaling through Spc1 SAPK. The Cdc37 protein physically interacts with Spc1, and the cdc37 mutation affects both the cellular level of the Spc1 protein and stress-induced Spc1 phosphorylation by Wis1 MAPK kinase (MAPKK). Consistently, expression of the stress response genes regulated by the Spc1 pathway is compromised in cdc37 mutant cells. On the other hand, a mutation in Hsp90, which often cooperates with Cdc37 in chaperoning protein kinases, does not affect Spc1 SAPK. These results suggest that Spc1 SAPK is a novel client protein for the Cdc37 chaperone, and the Cdc37 function is important to maintain the stability of the Spc1 protein and to facilitate stress signaling from Wis1 MAPKK to Spc1 SAPK.

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