Analysis of zinc transporter, hZnT4 ( Slc30A4 ), gene expression in a mammary gland disorder leading to reduced zinc secretion into milk

Abstract

Zinc deficiency, causing impaired growth and development, may have a nutritional or genetic basis. We investigated two cases of inherited zinc deficiency found in breast-fed neonates, caused by low levels of zinc in the maternal milk. This condition is different from acrodermatitis enteropathica but has similarities to the "lethal milk" mouse, where low levels of zinc in the milk of lactating dams leads to zinc deficiency in pups. The mouse disorder has been attributed to a defect in the ZnT4 gene. Little is known about the expression of the human orthologue, hZnT4 (Slc30A4). Sequence analysis of cDNA, real-time PCR and Western blot analysis of hZnT4, carried out on control cells and cells from unrelated mothers of two infants with zinc deficiency, showed no differences. The hZnT4 gene was highly expressed in mouthwash buccal cells compared with lymphoblasts and fibroblasts. The hZnT4 protein did not co-localise with intracellular free zinc pools, suggesting that hZnT4 is not involved in transport of zinc into vesicles destined for secretion into milk. This observation, combined with phenotypic differences between the "lethal milk" mouse and the human disorder, suggests that the "lethal milk" mouse is not the corresponding model for the human zinc deficiency condition.

Altered expression of two zinc transporters, SLC30A5 and SLC30A6, underlies a mammary gland disorder of reduced zinc secretion into milk

“…The following clinical presentation of two cases of zinc deficiency has previously been reported (Michalczyk et al 2003). Infant 1 was born premature at 36 weeks and was breastfed for three months.…”

“…Fibroblast and lymphoblast cell lines from patients (Mother 1 and Mother 2) and respective healthy controls (three lymphoblast and three fibroblast lines) were established as previously described (Michalczyk et al 2003). The human epithelial breast cell line PMC42-LA, a variant of PMC42 cell line, originally derived from a pleural effusion (Whitehead et al 1983).…”

“…Western blots were performed as previously described (Michalczyk et al 2003). Briefly, cell pellets were resuspended in 10 ml of lysis buffer and disrupted by passing through a 25-gauge needle 8-10 times followed by sonication on ice with a Microson Ultrasonic cell disruptor (Misonix Incorporated, NY, USA).…”

“…A study of the human orthologue (SLC30A4) of the murine Slc30a4 gene in two mothers with zinc-deficient milk, including sequence analysis of cDNA, real-time PCR and Western blot analysis, showed no differences between patient and normal control cells, which concluded that unlike the mouse disorder, defects in SLC30A4 were not responsible for the human disorder of reduced secretion of zinc into milk (Michalczyk et al 2003). In these patients, it is likely, therefore, that the maternal mammary gland defect in zinc secretion is different from the 'lethal milk' mouse model and that other candidate SLC30A transporters may underlie the mammary zinc transport defect.…”

The Znt4 mutation inlethal milk mice affects intestinal zinc homeostasis through the expression of other Zn transporters

“…This is especially true for the ZnT transporters (SLC30), most of which localize in intracellular vesicles (Palmiter et al, 1996a;Palmiter et al, 1996b;Murgia et al, 1999;Kambe et al, 2002;Huang et al, 2002;Michalczyk et al, 2003;Kirschke & Huang, 2003;Chimienti et al, 2004). Zinc absorption in the gut is mediated by the uptake activities of the ZIP4 and ZIP5 transporters at the apical and the basolateral side of enterocytes, respectively (DufnerBeattie et al, 2004;Wang et al, 2004;Wang et al, 2002).…”