Abstract
The regulation of cellular zinc uptake is a key process in the overall mechanism governing mammalian zinc homeostasis and how zinc participates in cellular functions. We analyzed the zinc transporters of the Zip family in both the brain and liver of zinc-deficient animals and found a large, significant increase in Zip10 expression. Additionally, Zip10 expression decreased in response to zinc repletion. Moreover, isolated mouse hepatocytes, AML12 hepatocytes, and Neuro 2A cells also respond differentially to zinc availability in vitro. Measurement of Zip10 hnRNA and actinomycin D inhibition studies indicate that Zip10 was transcriptionally regulated by zinc deficiency. Through luciferase promoter constructs and ChIP analysis, binding of MTF-1 to a metal response element located 17 bp downstream of the transcription start site was shown to be necessary for zinc-induced repression of Zip10. Furthermore, zinc-activated MTF-1 causes down-regulation of Zip10 transcription by physically blocking Pol II movement through the gene. Lastly, ZIP10 is localized to the plasma membrane of hepatocytes and neuro 2A cells. Collectively, these results reveal a novel repressive role for MTF-1 in the regulation of the Zip10 zinc transporter expression by pausing Pol II transcription. ZIP10 may have roles in control of zinc homeostasis in specific sites particularly those of the brain and liver. Within that context ZIP10 may act as an important survival mechanism during periods of zinc inadequacy.
Highlights
The metal response elements (MREs) upon being mutated lost responsiveness to zinc. These results show that the Zip10 promoter as a 3 kb fragment which includes the MRE at +17, responds to zinc as observed in vivo with mouse liver or AML12 hepatocytes, indicating the MTF-1 is necessary for the zinc repression of the reporter
In the present study we demonstrate that MTF-1 is an integral part of ZIP10-related cellular zinc homeostasis in the liver and brain both during zinc restriction and zinc excess
By using intact mice and isolated murine hepatocytes we show that MTF-1 can act as a repressor of Zip10 expression under normal physiologic levels of zinc
Summary
The mechanisms leading to these clinical manifestations of zinc deficiency remain elusive. Tight-control of cellular zinc homeostasis is maintained by proteins that can affect the amount of available zinc. Metal transporters of the ZRT/ IRT-like protein (ZIP) family and zinc transporter (ZnT) family, as well as zinc-binding by metallothioneins (MTs), maintain control of intracellular zinc levels [3,4],. 10 ZnT and 14 ZIP transporters have been identified. The ZnT proteins function in cellular zinc efflux or vesicular storage. ZnT1 was the first zinc transporter to be characterized, and is expressed in all tissues, localizing to the plasma membrane of cells [5]. Subsequent studies revealed zinc-regulated expression of ZnT1 in the intestine [6], and that zinc regulates expression of ZnT1 through activation of the metal response element-binding transcription factor MTF-1 [7]
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