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Toxicity mechanisms of the food contaminant citrinin: application of a quantitative yeast model.

Pascual-Ahuir A, Vanacloig-Pedros E, Proft M - Nutrients (2014)

Bottom Line: Additionally, genes in various multidrug resistance transport systems are functionally involved in the resistance to citrinin.Our study identifies the antioxidant defense as a major physiological response in the case of citrinin.In general, our results show that the use of live cell gene expression reporters in yeast are a powerful tool to identify toxicity targets and detoxification mechanisms of a broad range of food contaminants relevant for human nutrition.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia, Ingeniero Fausto Elio s/n, 46022 Valencia, Spain. apascual@ibmcp.upv.es.

ABSTRACT
Mycotoxins are important food contaminants and a serious threat for human nutrition. However, in many cases the mechanisms of toxicity for this diverse group of metabolites are poorly understood. Here we apply live cell gene expression reporters in yeast as a quantitative model to unravel the cellular defense mechanisms in response to the mycotoxin citrinin. We find that citrinin triggers a fast and dose dependent activation of stress responsive promoters such as GRE2 or SOD2. More specifically, oxidative stress responsive pathways via the transcription factors Yap1 and Skn7 are critically implied in the response to citrinin. Additionally, genes in various multidrug resistance transport systems are functionally involved in the resistance to citrinin. Our study identifies the antioxidant defense as a major physiological response in the case of citrinin. In general, our results show that the use of live cell gene expression reporters in yeast are a powerful tool to identify toxicity targets and detoxification mechanisms of a broad range of food contaminants relevant for human nutrition.

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Related in: MedlinePlus

Citrinin activates gene expression from CRE and AP-1 promoter elements in a dose dependent fashion. Artificial promoter-luciferase constructs were used, which contained multiple repetitions of the same cis-element: CRE, AP-1 or STRE as indicated. The indicated citrinin doses were applied at time point 0 to the yeast cultures and the light emission continuously monitored. Data shown are mean values from three independent biological samples. SD < 15%.
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nutrients-06-02077-f002: Citrinin activates gene expression from CRE and AP-1 promoter elements in a dose dependent fashion. Artificial promoter-luciferase constructs were used, which contained multiple repetitions of the same cis-element: CRE, AP-1 or STRE as indicated. The indicated citrinin doses were applied at time point 0 to the yeast cultures and the light emission continuously monitored. Data shown are mean values from three independent biological samples. SD < 15%.

Mentions: We next used yeast wild type cells, which were transformed with more specific luciferase reporters, for further citrinin studies. The insertion of multiple copies of just one type of transcription factor binding motif into luciferase expression plasmids, has been shown to create very specific reporters which respond to stimuli via just one or few signal transduction pathways [15,16]. Here, we investigated three types of cis elements: STRE (bound by the Msn2/4 factors in response to general stress), CRE (bound by the Sko1 factor in response to osmotic stress and by unknown factors in response to oxidative stress), AP-1 (bound by Yap1 in response to oxidative stress) [16]. As depicted in Figure 2, we found that citrinin exposure activated gene expression from AP-1 and CRE sites, but not from STRE elements. Since yeast AP-1 promoter elements are exclusively activated by oxidative damage, this was a clear indication that citrinin provoked intracellular oxidation, which then activated adaptive gene expression via oxidative stress responsive transcription factors such as Yap1. We further confirmed this by showing that activation of AP1-driven luciferase expression by citrinin was completely absent in a yap1 mutant strain (data not shown).


Toxicity mechanisms of the food contaminant citrinin: application of a quantitative yeast model.

Pascual-Ahuir A, Vanacloig-Pedros E, Proft M - Nutrients (2014)

Citrinin activates gene expression from CRE and AP-1 promoter elements in a dose dependent fashion. Artificial promoter-luciferase constructs were used, which contained multiple repetitions of the same cis-element: CRE, AP-1 or STRE as indicated. The indicated citrinin doses were applied at time point 0 to the yeast cultures and the light emission continuously monitored. Data shown are mean values from three independent biological samples. SD < 15%.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4042565&req=5

nutrients-06-02077-f002: Citrinin activates gene expression from CRE and AP-1 promoter elements in a dose dependent fashion. Artificial promoter-luciferase constructs were used, which contained multiple repetitions of the same cis-element: CRE, AP-1 or STRE as indicated. The indicated citrinin doses were applied at time point 0 to the yeast cultures and the light emission continuously monitored. Data shown are mean values from three independent biological samples. SD < 15%.
Mentions: We next used yeast wild type cells, which were transformed with more specific luciferase reporters, for further citrinin studies. The insertion of multiple copies of just one type of transcription factor binding motif into luciferase expression plasmids, has been shown to create very specific reporters which respond to stimuli via just one or few signal transduction pathways [15,16]. Here, we investigated three types of cis elements: STRE (bound by the Msn2/4 factors in response to general stress), CRE (bound by the Sko1 factor in response to osmotic stress and by unknown factors in response to oxidative stress), AP-1 (bound by Yap1 in response to oxidative stress) [16]. As depicted in Figure 2, we found that citrinin exposure activated gene expression from AP-1 and CRE sites, but not from STRE elements. Since yeast AP-1 promoter elements are exclusively activated by oxidative damage, this was a clear indication that citrinin provoked intracellular oxidation, which then activated adaptive gene expression via oxidative stress responsive transcription factors such as Yap1. We further confirmed this by showing that activation of AP1-driven luciferase expression by citrinin was completely absent in a yap1 mutant strain (data not shown).

Bottom Line: Additionally, genes in various multidrug resistance transport systems are functionally involved in the resistance to citrinin.Our study identifies the antioxidant defense as a major physiological response in the case of citrinin.In general, our results show that the use of live cell gene expression reporters in yeast are a powerful tool to identify toxicity targets and detoxification mechanisms of a broad range of food contaminants relevant for human nutrition.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia, Ingeniero Fausto Elio s/n, 46022 Valencia, Spain. apascual@ibmcp.upv.es.

ABSTRACT
Mycotoxins are important food contaminants and a serious threat for human nutrition. However, in many cases the mechanisms of toxicity for this diverse group of metabolites are poorly understood. Here we apply live cell gene expression reporters in yeast as a quantitative model to unravel the cellular defense mechanisms in response to the mycotoxin citrinin. We find that citrinin triggers a fast and dose dependent activation of stress responsive promoters such as GRE2 or SOD2. More specifically, oxidative stress responsive pathways via the transcription factors Yap1 and Skn7 are critically implied in the response to citrinin. Additionally, genes in various multidrug resistance transport systems are functionally involved in the resistance to citrinin. Our study identifies the antioxidant defense as a major physiological response in the case of citrinin. In general, our results show that the use of live cell gene expression reporters in yeast are a powerful tool to identify toxicity targets and detoxification mechanisms of a broad range of food contaminants relevant for human nutrition.

Show MeSH
Related in: MedlinePlus