Abstract
Atrazine, a member of the 2-chloro-s-triazine family of herbicides, is the most widely used pesticide in the world and often detected in agriculture watersheds. Although it was generally considered as an endocrine disruptor, posing a potential threat to human health, the molecular mechanisms of atrazine effects remain unclear. Using two-dimensional gel electrophoresis, we identified a panel of differentially expressed phosphoproteins and total proteins in human breast epithelial MCF-10A cells after being exposed to environmentally relevant concentrations of atrazine. Atrazine treatments for 6 h resulted in differential expression of 4 phosphoproteins and 8 total-proteins as compared to the control cells (>1.5-fold, p < 0.05). MALDI-TOF MS/MS analysis revealed that the differentially expressed proteins belong to various cellular compartments (nucleus, cytosol, membrane) and varied in function, including those regulating the stress response such as peroxiredoxin I, HSP70 and HSP27; structural proteins such as tropomyosin and profilin 1; and oncogenesis proteins such as ANP32A. Six of the 12 identified proteins were verified by quantitative PCR for their transcript levels. The most up-regulated phosphoprotein by atrazine treatment, ANP32A, was further analyzed for its expression, distribution and cellular localization using Western blot and immunocytochemical approaches. The results revealed that ANP32 expression after atrazine treatment increased dose and time dependently and was primarily located in the nucleus. This study may provide new evidence on the potential toxicity of atrazine in human cells.
Highlights
Atrazine, a member of the 2-chloro-s-triazine family of herbicides, is the most widely used pesticide in the world [1,2]
To further examine the atrazine effects on differential expressed total protein, the Pro-Q stained 2D gels were subjected to Coomassie
The gel images from the samples after 6 h atrazine treatment were shown since they represented the early events of differential phosphoprotein and total protein expression under the current experimental conditions (Figure 1)
Summary
A member of the 2-chloro-s-triazine family of herbicides, is the most widely used pesticide in the world [1,2] It is the most common pesticide present in ground and surface water [3,4,5,6,7,8,9] and chemically persistent in the environment [10]. Human breast epithelial MCF-10A cells were used as a model because atrazine has been reported to increase the incidence of mammary tumors in female Sprague-Dawley rats [29] and human breast cancers [27]. This research was to identify and verify the differentially expressed proteins induced by atrazine at environmental relevant concentrations in MCF-10A cells using proteomic, molecular and immunological methods. The data presented in this study would provide insights of the toxicological mechanisms of atrazine on human cells
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