Abstract
Slc39a8 encodes ZIP8, a divalent cation/bicarbonate symporter expressed in pluripotent mouse embryonic stem cells, and therefore ubiquitous in adult tissues; ZIP8 influxes Zn2+, Mn2+ and Fe2+. Slc39a8(neo/neo) knockdown mice exhibit 10–15% of wild-type ZIP8 mRNA and protein levels, and show pleiotropic phenotype of stunted growth, neonatal lethality, multi-organ dysmorphogenesis, and dysregulated hematopoiesis manifested as severe anemia. Herein we performed RNA-seq analysis of gestational day (GD)13.5 yolk sac and placenta, and GD16.5 liver, kidney, lung, heart and cerebellum, comparing Slc39a8(neo/neo) with Slc39a8(+/+) wild-type. Meta-data analysis of differentially-expressed genes revealed 29 unique genes from all tissues — having enriched GO categories associated with hematopoiesis and hypoxia and KEGG categories of complement, response to infection, and coagulation cascade — consistent with dysregulated hematopoietic stem cell fate. Based on transcription factor (TF) profiles in the JASPAR database, and searching for TF-binding sites enriched by Pscan, we identified numerous genes encoding zinc-finger and other TFs associated with hematopoietic stem cell functions. We conclude that, in this mouse model, deficient ZIP8-mediated divalent cation transport affects zinc-finger (e.g. GATA proteins) and other TFs interacting with GATA proteins (e.g. TAL1), predominantly in yolk sac. These data strongly support the phenotype of dysmorphogenesis and anemia seen in Slc39a8(neo/neo) mice in utero.
Highlights
Starting with a Cd2+-induced testicular necrosis phenotype and genetically “sensitive” vs “resistant” inbred mouse strains, this laboratory identified Slc39a8 as the major gene responsible for this trait[1,2,3]
ZIP8 is expressed in mouse visceral endoderm at gestational day (GD)7.5 [ref.9], in the gastrula stage[10], and in pluripotent embryonic stem cells[11]
The Slc39a8(neo/neo) mouse has been extensively characterized: pleiotropic effects include stunted growth, neonatal lethality, shortened limbs and deformed skull, severe anemia, dysregulation of hematopoiesis, and multi-organ dysmorphogenesis; consistent with the severe anemia — striking decreases in size of placenta, and size and number of hematopoietic islands are seen in Slc39a8(neo/neo) GD13.5 yolk sac and placenta, as well as in GD16.5 liver[13]
Summary
Starting with a Cd2+-induced testicular necrosis phenotype and genetically “sensitive” vs “resistant” inbred mouse strains, this laboratory identified Slc39a8 (encoding the ZIP8 transporter) as the major gene responsible for this trait[1,2,3]. ZIP8 is expressed in mouse visceral endoderm at gestational day (GD)7.5 [ref.9], in the gastrula stage[10], and in pluripotent embryonic stem cells[11]. These findings are compatible with the observation of early embryolethality, seen in Slc39a8(−/−) knockout mice when the gene is globally 100% ablated[12]. A Slc39a8 inducible-global knockout and a Slc39a8 hepatocyte-specific conditional knockout, created independently, exhibited striking decreases in Mn2+ concentrations in multiple organs tested; authors found lowered levels of two Mn2+-dependent enzymes – arginase and β-1,4-galactosyltransferase activities – and evidence that hepatic ZIP8 rescues Mn2+ from bile and regulates whole-body Mn2+ homeostasis[14]. The Slc39a8 global knockout shows striking cardiac extracellular matrix accumulation, suggesting that Slc39a8 expression is essential for development of ventricular compaction[15]
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