Evaluation of heavy metal toxicity in eukaryotes using a simple functional assay

Abstract

Although carcinogenesis caused by metals has been intensively investigated, the mechanisms of action, especially at the molecular level, are still unclear. This work aimed to investigate Cd(2+), Cu(2+), Ni(2+), Cr(3+), and Zn(2+) mutagenicity and its relationship with oxidative stress. We have applied the Functional Assay for the Separation of Alleles in Yeast (FASAY) with only minor modifications to detect p53 defects caused by metals. In this method, human p53-coding gene (TP53) expressed in Saccharomyces cerevisiae activates transcription of the ADE2 reporter gene. Yeast cells, expressing p53, were exposed to increased concentrations of metals and, then, plated on media supplemented or not with adenine. Yeast colonies containing functional p53 grow independently of adenine supplementation and colonies containing nonfunctional p53 are dependent on this nutrient. Mutations in the TP53 are implicated in the pathogenesis of half of all human tumors. According to our results, Cd(2+) was found to be the most toxic metal and produced the highest oxidative damage to lipids and proteins. At low concentrations (40 μM), this metal decreased viability and completely inhibited cell growth, while higher concentrations were necessary to produce the same toxic effect by Cu(2+), Cr(3+), and Ni(2+). Zn(2+) showed no significant toxicity. Cd(2+) strongly induced damages and altered the function of p53, while Cu(2+), followed by Cr(3+), showed lower percentages of p53-mutant colonies. Our results point towards a relationship between the loss of functional p53 protein and oxidative stress, a mechanism that can be associated with tumor formation induced by heavy metals in mammalian cells. By this adaptation of FASAY developed by us it is possible to easily and rapidly detect mutations caused by metals or other stresses.

Collate literature data on toxicity of Chromium (Cr) and Nickel (Ni) in experimental animals and humans

“…The inhibition of major DNA repair, oxidative DNA damage, deregulation of cell proliferation by induction of signaling pathway or inactivation of growth controls such as tumor suppresor genes for haploid Saccharomyces was reported for chromium (Riger et al, 2011).…”

Copper Acetate–DMSO Promoted Methylthiolation of Arenes and Heteroarenes

“…Copper promoted C-H functionalization either under free-radical or under oxidative conditions has been successfully exploited in a variety of reactions including peroxidation, alkylation, acetoxylation, halogenation, cyanation, and amination, to name a few . In our efforts directed toward development of copper catalyzed C-H functionalization reactions, we discovered copper acetate mediated direct synthesis of aryl and heteroaryl sulfide structural motifs with DMSO as a simple, cheap, and easy-to-handle methylthiolation reagent.…”

Actividad Mutagénica Inducida Por Hidrocarburos Aromáticos Policíclicos en Muestras De PM2.5 en Un Sector Residencial De Villa Del Rosario-Norte De Santander, Colombia

“…La mutagenicidad de las partículas diésel ha sido investigada por algunos autores (Singh et al 2004, Seung-Min y Chung 2006, Riger et al 2011) y se ha demostrado que inducen mutagenicidad en ausencia de enzimas microsomales (Taga et al 2005, Lingzhi et al 2007. Se ha descubierto que una gama más amplia de compuestos aromáticos policíclicos encontrados en las emisiones de combustión de diésel y en la contaminación del aire son a la vez mutagé-nicos y carcinogénicos como los nitroarenos, entre los que se incluyen a las lactonas del nitropireno y nitrofenantreno y a la 3-nitrobenzantrona que es un potente mutágeno encontrado en las partículas de diésel.…”