The Acrodermatitis Enteropathica Gene ZIP4 Encodes a Tissue-specific, Zinc-regulated Zinc Transporter in Mice

Abstract

The human ZIP4 gene (SLC39A4) is a candidate for the genetic disorder of zinc metabolism acrodermatitis enteropathica. To understand its role in zinc homeostasis, we examined the function and expression of mouse ZIP4. This gene encodes a well conserved eight-transmembrane protein that can specifically increase the influx of zinc into transfected cells. Expression of this gene is robust in tissues involved in nutrient uptake, such as the intestines and embryonic visceral yolk sac, and is dynamically regulated by zinc. Dietary zinc deficiency causes a marked increase in the accumulation of ZIP4 mRNA in these tissues, whereas injection of zinc or increasing zinc content of the diet rapidly reduces its abundance. Zinc can also regulate the accumulation of ZIP4 protein at the apical surface of enterocytes and visceral endoderm cells. These results provide compelling evidence that ZIP4 is a zinc transporter that plays an important role in zinc homeostasis, a process that is defective in acrodermatitis enteropathica in humans.A long recognized disease of zinc metabolism is the human genetic disorder acrodermatitis enteropathica (AE) 1 (1, 2). This autosomal recessive disorder causes classic symptoms of zinc deficiency (3), such as dermatological lesions, changes in the gastric mucosa associated with digestive system problems, lack of weight gain, and immune and reproductive problems (4 -8). Remarkably, these symptoms can be ameliorated by dietary zinc supplement (4, 6, 9 -11), consistent with the finding of reduced, but not eliminated, uptake of 65 Zn by the intestine from AE patients (12, 13), and the reduced uptake and total content of zinc in AE fibroblasts (14,15). Recent genetic mapping localized the AE gene to chromosome 8q24.3 (16) and led to its identification as a member of the ZIP superfamily (17, 18). That gene was named hZIP4 (the Human Genome Organization Nomenclature Committee named this gene SLC39A4).ZIP4 was found to be expressed in enterocytes and to reside in the plasma membrane. Mutations in hZIP4 were detected in AE patients (17, 18), strongly suggesting that they cause this genetic disorder.The recently identified ZIP superfamily of metal ion uptake transporters (19,20) are found in all eukaryotes, and many of its members mediate zinc uptake. In yeast, ZRT1 encodes the high affinity zinc transporter, and ZRT2 encodes the low affinity zinc transport system. The Arabidopsis iron-regulated transporter gene (IRT1) encodes a metal transporter that has remarkable sequence similarity with the yeast ZRTs and with other Arabidopsis zinc transporters (19, 21). Thus, the acronym ZIP was adopted to reflect ZRT/IRT-related proteins. Many members of the ZIP gene superfamily have now been detected based on sequence homology with yeast and Arabidopsis ZIP genes (22, 23). The ZIP proteins typically have eight membrane-spanning domains, and spanning domain four contains fully conserved histidyl and glycyl residues in an amphipathic ␣-helix. These proteins also often have a histidine-rich intracellular loop between s...

Novel zinc-responsive post-transcriptional mechanisms reciprocally regulate expression of the mouse Slc39a4 and Slc39a5 zinc transporters (Zip4 and Zip5)

“…Embryos in pregnant mice fed a ZnD diet beginning on d8 of pregnancy have a greatly increased risk of abnormal development (Dufner-Beattie et al, 2005. As noted previously (Dufner-Beattie et al, 2003b, Zip4 mRNA abundance was significantly increased in the maternal intestine and fetal VYS in mice fed the ZnD diet, while Zip5 and Zip1 mRNA abundance were unaltered (Figure 3). In a previous study, we noted that Zip4 mRNA levels in the VYS decline by 9 h after an intraperitoneal injection of zinc; earlier time points were not examined in that study.…”

“…Therefore, we used these cells as a model system in which to study regulation of the endogenous Zip4 gene. In the presence of 10% FBS in the culture medium, the addition of 100 to 150 μM zinc maximally repressed Zip4 mRNA (Dufner-Beattie et al, 2003b) and maximally induced metallothioneins-I (MT-I) mRNA in Hepa cells ( Figure 1B,C). Expression of the mouse MT-I gene is regulated by the zinc-sensing transcription factor MTF-1 (Andrews et al, 2001) and MT-I functions to chelate excess zinc and provide a biologically important pool of labile zinc when zinc becomes limiting (Dalton et al, 1996).…”

“…The mouse Zip4 gene is essential early during development of the post-implantation embryo and heterozygosity hypersensitizes mice to the effects of dietary zinc deficiency . Although the physiological functions of ZIP5 are unknown, it is a closely related member of the LIV-1 subfamily that can also function as a zinc transporter (Dufner-Beattie et al, 2003bWang et al, 2004a) and is also expressed in tissues involved in zinc homeostasis (intestine, pancreas and visceral endoderm).…”

“…Where ZIP4 is important for zinc acquisition from the diet (or mother), whereas ZIP5 functions to remove excess zinc from the blood when zinc is replete. Zip4 mRNA abundance is increased in the adult and neonatal intestine and embryonic visceral yolk sac during periods of dietary zinc deficiency (Dufner-Beattie et al, 2003b;Huang et al, 2006) leading to the intense localization of ZIP4 protein on the apical membranes of enterocytes and visceral endoderm cells and increased amounts of this protein . Studies of transfected cells revealed that zinc can stimulate the endocytosis and degradation of ZIP4 (Kim et al, 2004((•• a or b? ••)); Mao et al, 2007), and ZIP4 is removed from the apical surfaces of enterocytes and endoderm cells several hours after an injection of zinc into the peritoneum of zinc deficient mice (Dufner-Beattie et al, 2003b).…”

“…Zip4 mRNA abundance is increased in the adult and neonatal intestine and embryonic visceral yolk sac during periods of dietary zinc deficiency (Dufner-Beattie et al, 2003b;Huang et al, 2006) leading to the intense localization of ZIP4 protein on the apical membranes of enterocytes and visceral endoderm cells and increased amounts of this protein . Studies of transfected cells revealed that zinc can stimulate the endocytosis and degradation of ZIP4 (Kim et al, 2004((•• a or b? ••)); Mao et al, 2007), and ZIP4 is removed from the apical surfaces of enterocytes and endoderm cells several hours after an injection of zinc into the peritoneum of zinc deficient mice (Dufner-Beattie et al, 2003b). Our recent studies of mouse ZIP5 revealed that this protein is localized on basolateral membranes of intestinal enterocytes, visceral endoderm cells and pancreatic acinar cells when dietary zinc is not limiting (Dufner-Beattie et al, 2004), but in contrast to ZIP4, this protein is removed from these basolateral membranes of each of these cell types during periods of dietary zinc deficiency.…”