Intestinal Absorption of Minerals II Iron and other micro-metals

Handling the metal internally Expelling the metal from the enterocyte Serosal Side Ferritin (Fe) Metallothionein (Zn, Cu, Cd) Efflux system for export

A metal ion upon entering the enterocytes is either delayed in transit or released quickly from antiluminal (basolateral) surface into the system Binding to internal stationary factors such as proteins and vesicles allows the metal to be sequestered for safety purposes and for regulating the flow of that metal into the system. The above scenario plays out for Fe, Zn and Cu, where storage and transport proteins have been identified. Metallothionein: Binds Cu, Zn, and Cd. Will not bind Fe Ferritin: Binds Fe exclusively Binding to Internal Factors

Ferritin Gold are gated pores that control iron release from the inner core

24 subunits, with 2 distinct isoforms: H ferritin and L ferritin To release iron, the Fe 3+ must first be reduced to Fe 2+. H: predominates in heart L: predominates in liver To store iron, the Fe2+ must first be oxidized to Fe3+

b. Iron is stored in ferritin as Fe(III) in the mineral [FeO(OH)] 8 [FeO(H 2 PO 4 )]. This mineral can be represented by ferrihydrite, FeO(OH) (shown above). Note: the name "ferrihydrite" is used for both [FeO(OH)] 8 [FeO(H 2 PO 4 )] and FeO(OH). Note: Iron (III) ions are shown in brown, and oxygen (II) ions are shown in red. Ferrihydrite

Iron Export from Mucosal Cells

Duodenal LumenDuodenal MucosaPlasma Heme- Protein Heme + Polypeptides Mucin (gastroferrin) Fe 3+ B2-microglobulin HFE Biliverdin Bilirubin Bilirubin CO Heme Oxygenase Heme Fe 2+ Fe 3+ DCT-1 B 3 integrin paraferrin Mobilferrin (vesicles) Fe 2+ Fe 3+ Iron Absorption (heme and non-heme) Ferroportin Ferritin Fe 3+ FR Fe 3+ Haephestin Transferrin

Ferroportin, the only way iron can escape from the cell. Adriana Donovan and Nancy Andrews were the first to show the importance of ferroportin

Knockout mouse cells lacking the gene for ferroportin are unable to release iron (blue)

Ferroportin appears to be the portal for releasing iron from the cell. Also called IREG1 and MPT1 FP Ferroportin Hc Hepcidin (Hc) downregulates the surface concentration of ferroportin thereby controlling the concentration of the iron exported from the cell. Hc Considered the master regulator of cellular iron export

Hephaestin (a Cu protein): Iron exported from the enterocyte at the basal surface is primarily Fe2+. In order to transfer Fe2+ to transferrin it must be oxidized to Fe3+. Oxidation of Fe2+ to Fe3+ on the serosal side of the intestine is catalyzed by a copper protein, haephestin. Haephestin (name after the Greek god of metals) was isolated as the gene product of a genetic defect in mice called Sex-Linked Anemia (SLA). Mice with the defect were iron deficient causing a pronounced anemia

Regulation of Iron Uptake Exogenous dietary factors Role of IRPs (Iron regulatory proteins)

Dietary Factors that Influence Iron Absorption Amino AcidsOrganic AcidsSugars Cysteine Glycine Histidine Lysine Methionine Ascorbate Citrate Lactate Malate tartaric Fructose Sorbitol Facilitate Inhibit Bran Hemicellulose Cellulose Pectin Guar gum Fibers Polyphenolics Flavonoids Anthrocyanins Isoflavonoids Others Oxalates Carbonate Phosvitin (iron- binding protein in egg yolk)

Iron, a “one-way” metal Excretion of iron from the body is not regulated Iron status of the system is controlled only at the absorption stage

Intestinal iron transport is influenced by the amount of iron in the diet. Sequestering by ferritin when iron is in abundant supply is one mechanism. Another is regulating the surface density of the importing and exporting factors. DMT-1 and ferroprotin 1(FPN1), respectively. Both are subject to mobilizing effects in response to iron. When iron is low DMT1 is rich on the cell surface primed to input more iron. FPN1 in low iron is localize to the cytosol. When iron is enriched DMT1 is down-regulated, meaning it shifts to the cytosol. The opposite occurs with FPN1 where movement in response to iron is to the membrane preparing the cell to release more iron.

How do you interpret these observations? (iron-dependent enzyme in the cytosol) Iron storage protein No iron, no ferritin

Regulation at the level of transferrin receptor and ferritin mRNA Iron regulatory protein (IRP) Iron regulatory protein (IRP) Iron response elements Transferrin receptor is believed to be the factor that tunes iron status of the body to iron absorption in the intestine

Addendum IRPs control the expression of DMT-1 (DCT-1, Nramp2) mRNA and protein and are critically important to secure physiological levels of ferroportin, the iron export protein. IRPs thus coordinate the synthesis of key iron metabolism proteins in the duodenum. Galy et al, 2008

Vesicle Transport of Metals Zn Cu

Metallothionein A small metal binding protein with an unusually high amount of cysteine residues. One third of the residues are cysteine Binding sites for Zn2+ and Cu+ in different locations of the protein Primary role was considered to be detoxification, not nutrition

Metallothionein 6 copper atoms inside bound to cysteines Cu Cysteine

Storage and Release of Metals from Metallothionein Reduced glutathione controls the storage of copper and zinc by glutathione; oxidized controls the release Reduced

Absorption Mucosa Serosa Zn++ NSBP MTI MTI-Zn CRIP CRIP-Zn Zn++ Zn++-Albumin Albumin CRIP=cysteine-rich intestinal protein; MTI=metallothionine; NSBP, non-specfic binding protein Non-saturable = Passive Diffusion Saturable = Bound to form transport ligand Zn++-Albumin

A 13-year-old girl presented with a history of red scaly plaques involving the chest, arms and legs beginning in infancy. Punch biopsy revealed psoriasiform hyperplasia and pallor of the epidermis. The patient's serum zinc level was 36 μg/dl [nl μg/dl]. A diagnosis of acrodermatitis enteropathica was established and the patient responded well to zinc replacement therapy. Acrodermatitis enteropathica is a rare autosomal recessive disorder caused by mutations in SLC39A4, which encodes the tissue-specific zinc transporter ZIP4

Cellular access protein Glucose, amino acid transporters K + channel protein, Na/K ATPase CaT1 DCT-1 (DMT-1)? Cytosol storage protein None VesiclesNone Cytosol trafficking protein None CalbindinNone Basal export protein Na/K ATPaseNoneCa-ATPaseNone Plasma trafficking protein None Albumin Sodium Potassium Calcium Magnesium Proteins Involved in the Absorption and Transport of Macro-metals Process

Cellular access protein Mobilferrin DCT-1 (DMT-1) Zip4Ctr1 Cytosol storage protein FerritinMetallothionein Cytosol trafficking protein MobilferrinZip4-containing vesicles, CRIP Atox1 Basal export protein FerroportinZnT1ATP7A Plasma trafficking protein Transferrin  2 - macroglobulin albumin Albumin, transcuprein, ceruloplasmin ProcessIron ZincCopper Proteins Involved in the Absorption and Transport of Micro-metals