Marina Gálvez-Peralta, Lei He, Lucia F. Jorge-Nebert, Bin Wang, Marian L. Miller, Bryan L. Eppert, Scott Afton, Daniel W. Nebert

Previously this laboratory characterized Slc39a8-encoded ZIP8 as a Zn(2+)/(HCO(3)(-))(2) symporter; yet, the overall physiological importance of ZIP8 at the whole-organism level remains unclear. Herein we describe the phenotype of the hypomorphic Slc39a8(neo/neo) mouse which has retained the neomycin-resistance gene in intron 3, hence causing significantly decreased ZIP8 mRNA and protein levels in embryo, fetus, placenta, yolk sac, and several tissues of neonates. The Slc39a8(neo) allele is associated with diminished zinc and iron uptake in mouse fetal fibroblast and liver-derived cultures; consequently, Slc39a8(neo/neo) newborns exhibit diminished zinc and iron levels in several tissues. Slc39a8(neo/neo) homozygotes from gestational day(GD)-11.5 onward are pale, growth-stunted, and die between GD18.5 and 48 h postnatally. Defects include: severely hypoplastic spleen; hypoplasia of liver, kidney, lung, and lower limbs. Histologically, Slc39a8(neo/neo) neonates show decreased numbers of hematopoietic islands in yolk sac and liver. Low hemoglobin, hematocrit, red cell count, serum iron, and total iron-binding capacity confirmed severe anemia. Flow cytometry of fetal liver cells revealed the erythroid series strikingly affected in the hypomorph. Zinc-dependent 5-aminolevulinic acid dehydratase, required for heme synthesis, was not different between Slc39a8(+/+) and Slc39a8(neo/neo) offspring. To demonstrate further that the mouse phenotype is due to ZIP8 deficiency, we bred Slc39a8(+/neo) with BAC-transgenic BTZIP8-3 line (carrying three extra copies of the Slc39a8 allele); this cross generated viable Slc39a8(neo/neo)_BTZIP8-3(+/+) pups showing none of the above-mentioned congenital defects-proving Slc39a8(neo/neo) causes the described phenotype. Our study demonstrates that ZIP8-mediated zinc transport plays an unappreciated critical role during in utero and neonatal growth, organ morphogenesis, and hematopoiesis.