Zn-Responsive Proteome Profiling and Time-Dependent Expression of Proteins Regulated by MTF-1 in A549 Cells

Abstract

Zinc plays a critical role in many biological processes. However, it is toxic at high concentrations and its homeostasis is strictly regulated by metal-responsive transcription factor 1 (MTF-1) together with many other proteins to protect cells against metal toxicity and oxidative stresses. In this paper, we used high-resolution two-dimensional gel electrophoresis (2DE) to profile global changes of the whole soluble proteome in human lung adenocarcinoma (A549) cells in response to exogenous zinc treatment for 24 h. Eighteen differentially expressed proteins were identified by MALDI TOF/TOF and MASCOT search. In addition, we used Western blotting and RT-PCR to examine the time-dependent changes in expression of proteins regulated by MTF-1 in response to Zn treatment, including the metal binding protein MT-1, the zinc efflux protein ZnT-1, and the zinc influx regulator ZIP-1. The results indicated that variations in their mRNA and protein levels were consistent with their functions in maintaining the homeostasis of zinc. However, the accumulation of ZIP-1 transcripts was down-regulated while the protein level was up-regulated during the same time period. This may be due to the complex regulatory mechanism of ZIP-1, which is involved in multiple signaling pathways. Maximal changes in protein abundance were observed at 10 h following Zn treatment, but only slight changes in protein or mRNA levels were observed at 24 h, which was the time-point frequently used for 2DE analyses. Therefore, further study of the time-dependent Zn-response of A549 cells would help to understand the dynamic nature of the cellular response to Zn stress. Our findings provide the basis for further study into zinc-regulated cellular signaling pathways.

The phosphoinositide 3-kinase pathway and glycogen synthase kinase-3 positively regulate the activity of metal-responsive transcription factor-1 in response to zinc ions

“…Notably, the Mtf-1 gene is essential, as Mtf-1 knock-out (KO) mice die in utero due to liver degeneration and oedema (Günes et al 1998). MTF-1 regulates the expression of several genes, including those encoding placenta growth factor (Green et al 2001), several zinc transporters (Zhao et al 2014), the human prion protein (Bellingham et al 2009), and mir132 miRNA (Lee et al 2007). Knocking-down MTF-1 with siRNAs changes the transcriptional response to zinc for over 1000 genes (Hogstrand et al 2008).…”

Time‐dependent response of A549 cells upon exposure to cadmium

“…The proteins Hsp90α and hnRNPA1 were found to respond to metal exposure (Barque, Abahamid, Chacun, & Bonaly, ; Padmini & Rani, ; Zhao et al, ; Zhao et al, ). The kinetic responses and differential expression patterns of Hsp90α and hnRNPA1 were observed when A549 cells were incubated with CdSO 4 for 1, 3, 6, 9, 12 or 24 hours.…”

“…In previous studies, we have examined the time‐dependent changes in the expression of proteins and the proteome involved in Zn homeostasis, and we elucidated the mechanism of the process. Our results showed that A549 cells present a kinetic response to exogenous zinc sulfate (ZnSO 4 ) stress in four conservatively time‐dependent manners and that exogenous ZnSO 4 more predominantly reduced the expression of proteins in cells after 24 hours than 9 hours (Zhao et al, ; Zhao et al, ). In the present work, to unclose further the difference of response mechanisms of different cells to various metals ions, we profiled the toxicokinetic response of the proteome in A549 cells exposed to extracellular CdSO 4 at different time points using 2DE coupled with silver staining.…”

The crosstalk between trace elements with DNA damage response, repair, and oxidative stress in cancer

“…In addition, the metabolism of As 3+ to As 5+ affects the reduction of H 2 O, which generates H 2 O 2 , and some metabolic intermediates, such as dimethyl arsine peroxyl radical. 124 OS contributes to As 3+ -induced kinase activation that leads to S21 phosphorylation of the enhancer of zeste homolog 2 (EZH2) protein, and is present in the direct interaction of the activated Akt and EZH2. Furthermore, OS appears to be able to induce cytoplasmic localization of the phosphorylated and the nonphosphorylated EZH2 protein in As 3+ -treated cells.…”

“…These proteins show maximum changes in their expression by Zn treatment, and the proteome of A549 cells possibly exhibits significant changes to exogenous Zn ions exposure. 124 Zn and the ionophore compound pyrrolidinedithiocarbamate (PDTC) treatment provoke apoptosis, which is associated with ROS-dependent nuclear translocation of the mitochondrial factor AIF. P53 degradation plays an important role in fibroblast resistance to Zn/PDTC.…”