Endothelial nitric oxide synthase uncoupling as a key mediator of melanocyte malignant transformation associated with sustained stress conditions

Melanoma cell lines and cells corresponding to premalignant melanocytes were established by our group after subjecting a nontumorigenic murine melanocyte lineage, melan-a, to sequential cycles of anchorage blockade. Previous results showed that in melan-a cells the superoxide level increases after such procedure. Superoxide production during melanocyte de-adhesion was inhibited by L-sepiapterin, the precursor of eNOS cofactor BH(4), and increased by the inhibitor of BH(4) synthesis, DAHP, hence indicating a partial uncoupling state of eNOS. the eNOS uncoupling seems to be maintained in cells derived from melan-a, because they present decreased nitric oxide and increased superoxide levels. the inhibition of superoxide production in Tm5 melanoma cells with L-sepiapterin reinforces their eNOS-uncoupled state. the maintenance of oxidative stress seems to be important in melanoma apoptosis resistance because Mn(III)TBAP, a superoxide scavenger, or L-sepiapterin renders Tm5 cells more sensitive to anoikis and chemotherapy. More importantly, eNOS uncoupling seems to play a pivotal role in melanocyte malignant transformation induced by sustained anchorage impediment, because no malignant transformation was observed when L-NAME-treated melanocytes were subjected to sequential cycles of de-adhesion. Our results show that uncoupled eNOS contributes to superoxide production during melanocyte anchorage impediment, contributing to anoikis resistance and malignant transformation. (C) 2011 Elsevier Inc. All rights reserved.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade Federal de São Paulo, Disciplina Imunol, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Farmacol, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Nefrol, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Imunol, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Farmacol, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Nefrol, BR-04023900 São Paulo, BrazilFAPESP: 06/61293-1FAPESP: 05/60334-3FAPESP: 08/50366FAPESP: 09/03335-8Web of Scienc

Nitric Oxide Activates p21(ras) by S-Nitrosylation during Loss of Integrin-Mediated Cell-Matrix Contact Leading to Increased Superoxide Level and Cell Survival

Universidade Federal de São Paulo, São Paulo, BrazilUniv São Paulo, São Paulo, BrazilUniversidade Federal de São Paulo, São Paulo, BrazilWeb of Scienc

Correction: Ras and Rac1, frequently mutated in melanomas, are activated by superoxide anion, modulate Dnmt1 level and are causally related to melanocyte malignant transformation.

[This corrects the article DOI: 10.1371/journal.pone.0081937.]

Risikobasierte Bewertung von Hochwasserschutzmaßnahmen mit unterschiedlichen Wirkungsweisen auf den Hochwasserschutz

A melanocyte malignant transformation model was developed in our laboratory, in which different melanoma cell lines were obtained after submitting the non-tumorigenic melanocyte lineage melan-a to sequential cycles of anchorage impediment. Our group has already showed that increased superoxide level leads to global DNA hypermemethylation as well increased Dnmt1 expression few hours after melanocyte anchorage blockade. Here, we showed that Ras/Rac1/ERK signaling pathway is activated in melanocytes submitted to anchorage impediment, regulating superoxide levels, global DNA methylation, and Dnmt1 expression. Interestingly, Ras and Rac1 activation is not related to codon mutations, but instead regulated by superoxide. Moreover, the malignant transformation was drastically compromised when melan-a melanocytes were submitted to sequential cycles of anchorage blockage in the presence of a superoxide scavenger. This aberrant signaling pathway associated with a sustained stressful condition, which might be similar to conditions such as UV radiation and inflammation, seems to be an early step in malignant transformation and to contribute to an epigenetic reprogramming and the melanoma development

Transcriptional regulation of dnmt1 by E2F1 in melanoma progression

Universidade Federal de São Paulo, São Paulo, BrazilUniversidade Federal de São Paulo, São Paulo, BrazilWeb of Scienc