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Oncogene activation in melanocytes links reactive oxygen to multinucleated phenotype and senescence

Abstract

Contrary to malignant melanoma, nevi are a benign form of melanocytic hyperproliferation. They are frequently observed as precursor lesions of melanoma, but they also feature biochemical markers of senescence. In particular, evidence for oncogene-induced melanocyte senescence as natural means to prevent tumorigenesis has been obtained in nevi with mutated B-RafV600E. Here, we demonstrate that strong oncogenic growth factor receptor signalling drives melanocytes into senescence, whereas weaker signals keep them in the proliferative state. Activation of oncogene-induced senescence also produces multinucleated giant cells, a long known histological feature of nevus cells. The protein levels of the senescence mediators, p53 and pRB, and their upstream activators do not correlate with senescence. However, strong oncogene signalling leads to pronounced reactive oxygen stress, and scavenging of reactive oxygen species (ROS) efficiently prevents the formation of multinucleated cells and senescence. Similarly, expression of oncogenic N-RAS results in ROS generation, DNA damage and the same multinuclear senescent phenotype. Hence, we identified oncogenic signalling-dependent ROS production as critical mediator of the melanocytic multinuclear phenotype and senescence, both of them being hallmarks of human nevus cells.

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Abbreviations

ATM:

ataxia telangiectasia mutated

ARF:

alternative reading frame

CDKN2A:

cyclin-dependent kinase inhibitor 2A

Chk2:

checkpoint kinase 2

CKI:

cyclin-dependent kinase inhibitor

DCF:

2′,7′-dichlorofluorescein

EGF:

epidermal growth factor

EGFR:

epidermal growth factor receptor

Hm:

HERmrk

INK4A:

inhibitor of cyclin-dependent kinase 4A

INK4B:

inhibitor of cyclin-dependent kinase 4B

NAC:

N-acetylcysteine

ROS:

reactive oxygen species

RTK:

receptor tyrosine kinase

SA-β-gal:

senescence-associated β-galactosidase

Xmrk:

Xiphophorus melanoma receptor kinase

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Acknowledgements

We thank Dr Thorsten Stiewe for kindly providing the PAB 122 hybridoma supernatant and Professor Jürgen Hoppe for his help with the time lapse experiment. In addition, we thank Johannes Haydn for generating the melan a pWHE459 cell lines and Toni Wagner for great technical support. This work was supported by the Deutsche Forschungsgesellschaft, Transregio 17 (‘RAS-dependent pathways in human cancer’). CL was supported by a grant of the German Excellence Initiative to the Graduate School of Life Sciences, University of Wurzburg.

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Correspondence to S Meierjohann.

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Leikam, C., Hufnagel, A., Schartl, M. et al. Oncogene activation in melanocytes links reactive oxygen to multinucleated phenotype and senescence. Oncogene 27, 7070–7082 (2008). https://doi.org/10.1038/onc.2008.323

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