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The stimulation of mitogenic signaling pathways by N-POMC peptides.

Pepper DJ, Bicknell AB - Mol. Cell. Endocrinol. (2008)

Bottom Line: However, little is known about the molecular mechanisms by which mitogenesis is stimulated, although it has been shown that N-POMC(1-28) stimulates the ERK pathway in human H295R cells.Similar results were observed for both MEK and c-Raf phosphorylation, although N-POMC(1-49) stimulated the phosphorylation of Akt more robustly than N-POMC(1-28).Sequencing of 114 clones from each cDNA population revealed the expression of a number of receptors, several of which have not been described previously in the adrenal.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, The University of Reading, Whiteknights, PO Box 228, Reading, Berkshire RG6 6AJ, UK.

ABSTRACT
The N-terminal fragment of pro-opiomelancortin (POMC) has been shown previously to act as an adrenal mitogen. However, little is known about the molecular mechanisms by which mitogenesis is stimulated, although it has been shown that N-POMC(1-28) stimulates the ERK pathway in human H295R cells. We have investigated signaling stimulated by N-POMC(1-28) and N-POMC(1-49) in the mouse Y1 cell line and found that both peptides stimulate ERK phosphorylation with maximal stimulation being achieved within 5min. Similar results were observed for both MEK and c-Raf phosphorylation, although N-POMC(1-49) stimulated the phosphorylation of Akt more robustly than N-POMC(1-28). We also investigated the expression of tyrosine kinase receptors in adrenal cells. PCR utilizing degenerate primers was performed on cDNA from both Y1 cells and rat adrenal tissue. Sequencing of 114 clones from each cDNA population revealed the expression of a number of receptors, several of which have not been described previously in the adrenal.

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Serum starved Y1 cells were stimulated with either 1 nM of N-POMC1–28 (A and C) or N-POMC1–49 (B and D) for various times. Cell lysates were analyzed by immunoblotting with antibodies against phospho MEK1/2 (A and B) and phospho c-RAF (C and D) ERK. Band intensities are normalized to those of the total amount of p44 ERK and expressed as the mean (±S.E.M.) of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 relative to value at 0 min.
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fig2: Serum starved Y1 cells were stimulated with either 1 nM of N-POMC1–28 (A and C) or N-POMC1–49 (B and D) for various times. Cell lysates were analyzed by immunoblotting with antibodies against phospho MEK1/2 (A and B) and phospho c-RAF (C and D) ERK. Band intensities are normalized to those of the total amount of p44 ERK and expressed as the mean (±S.E.M.) of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 relative to value at 0 min.

Mentions: As both peptides could clearly stimulate the phosphorylation of ERK we next investigated if they could also phosphorylate the upstream regulators of ERK: MEK and c-RAF. Using immunoblotting together with specific phospho antibodies, both peptides were found to stimulate the phosphorylation of both MEK and c-RAF (Fig. 2). We also investigated if N-POMC1–28 and N-POMC1–49 were capable of stimulating the phosphorylation of Akt, a key molecule in the PI3 kinase pathway. N-POMC1–49 was found to stimulate a robust phosphorylation of Akt, while in contrast the response to N-POMC1–28 was weaker (Fig. 3).


The stimulation of mitogenic signaling pathways by N-POMC peptides.

Pepper DJ, Bicknell AB - Mol. Cell. Endocrinol. (2008)

Serum starved Y1 cells were stimulated with either 1 nM of N-POMC1–28 (A and C) or N-POMC1–49 (B and D) for various times. Cell lysates were analyzed by immunoblotting with antibodies against phospho MEK1/2 (A and B) and phospho c-RAF (C and D) ERK. Band intensities are normalized to those of the total amount of p44 ERK and expressed as the mean (±S.E.M.) of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 relative to value at 0 min.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651482&req=5

fig2: Serum starved Y1 cells were stimulated with either 1 nM of N-POMC1–28 (A and C) or N-POMC1–49 (B and D) for various times. Cell lysates were analyzed by immunoblotting with antibodies against phospho MEK1/2 (A and B) and phospho c-RAF (C and D) ERK. Band intensities are normalized to those of the total amount of p44 ERK and expressed as the mean (±S.E.M.) of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 relative to value at 0 min.
Mentions: As both peptides could clearly stimulate the phosphorylation of ERK we next investigated if they could also phosphorylate the upstream regulators of ERK: MEK and c-RAF. Using immunoblotting together with specific phospho antibodies, both peptides were found to stimulate the phosphorylation of both MEK and c-RAF (Fig. 2). We also investigated if N-POMC1–28 and N-POMC1–49 were capable of stimulating the phosphorylation of Akt, a key molecule in the PI3 kinase pathway. N-POMC1–49 was found to stimulate a robust phosphorylation of Akt, while in contrast the response to N-POMC1–28 was weaker (Fig. 3).

Bottom Line: However, little is known about the molecular mechanisms by which mitogenesis is stimulated, although it has been shown that N-POMC(1-28) stimulates the ERK pathway in human H295R cells.Similar results were observed for both MEK and c-Raf phosphorylation, although N-POMC(1-49) stimulated the phosphorylation of Akt more robustly than N-POMC(1-28).Sequencing of 114 clones from each cDNA population revealed the expression of a number of receptors, several of which have not been described previously in the adrenal.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, The University of Reading, Whiteknights, PO Box 228, Reading, Berkshire RG6 6AJ, UK.

ABSTRACT
The N-terminal fragment of pro-opiomelancortin (POMC) has been shown previously to act as an adrenal mitogen. However, little is known about the molecular mechanisms by which mitogenesis is stimulated, although it has been shown that N-POMC(1-28) stimulates the ERK pathway in human H295R cells. We have investigated signaling stimulated by N-POMC(1-28) and N-POMC(1-49) in the mouse Y1 cell line and found that both peptides stimulate ERK phosphorylation with maximal stimulation being achieved within 5min. Similar results were observed for both MEK and c-Raf phosphorylation, although N-POMC(1-49) stimulated the phosphorylation of Akt more robustly than N-POMC(1-28). We also investigated the expression of tyrosine kinase receptors in adrenal cells. PCR utilizing degenerate primers was performed on cDNA from both Y1 cells and rat adrenal tissue. Sequencing of 114 clones from each cDNA population revealed the expression of a number of receptors, several of which have not been described previously in the adrenal.

Show MeSH
Related in: MedlinePlus