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Regulation of ZIP and ZnT zinc transporters in zebrafish gill: zinc repression of ZIP10 transcription by an intronic MRE cluster.
Zheng D
,
Feeney GP
,
Kille P
,
Hogstrand C
.
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Resolving the mechanisms underlying the temporal and spatial profile of zinc transporter expression in response to zinc availability is key to understanding zinc homeostasis. The mRNA expression of seven zinc transporters was studied in zebrafish gills when treated with zinc deficiency/excess over a 14-day period. Of these, ZnT1, ZnT5, ZIP3, and ZIP10 were differentially expressed in response to changed zinc status. The mRNA level of zinc exporter, ZnT1, was upregulated in fish subjected to excess zinc and downregulated by zinc deprivation. This response was similar to that of metallothionein-2 (MT2). Zinc deficiency caused an increased abundance of mRNA for zinc importers ZnT5, ZIP3, and ZIP10. Expression of ZnT5 and ZIP10, but not ZIP3, was inhibited by excess zinc. Zinc influx function of ZIP10 was demonstrated by (65)Zn transport assays in Xenopus oocyte expression experiments, suggesting that the inverse relationship between zinc availability and ZIP10 expression serves to maintain zinc homeostasis. Two distinct transcription start sites (TSS) for ZIP10 were found in gill and kidney. Luciferase assays and mutation/deletion analysis of DNA fragments proximal to the respective TSS revealed that ZIP10 has two alternative promoters (P1 and P2) displaying opposite regulatory control in response to zinc status. Positive as well as negative regulation by zinc involves MRE clusters in the respective promoters. These results provide experimental evidence for MREs functioning as repressor elements, implicating MTF1 involvement in the negative regulation of ZIP10. This is in contrast to the well-established positive regulation by MTF1 of other genes, such as MT2 and ZnT1.
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