Bryan Mackenzie, PhD
Associate Professor
bryan.mackenzie@uc.edu

Iron deficiency is the most prevalent micronutrient deficiency worldwide. Meanwhile iron overload associated with conditions like hereditary hemochromatosis, thalassemia, or sickle-cell disease poses a serious threat to many other individuals. Divalent metal-ion transporter-1 (DMT1) is indispensable for iron homeostasis. It is the front-line, primary route of iron uptake in the intestine. DMT1 is also responsible for mobilization of iron from the endosome to cytosol, a crucial step in the transferrin-associated uptake of iron in erythroid precursor cells. Export of iron from enterocytes and from macrophages (recycling iron from senescent red blood cells) is thought to be mediated by ferroportin (Fpn). Whereas Fpn was identified a decade ago, very little is known about how this transporter works.

We are investigating the molecular mechanisms, substrate selectivity and structure–function relationships of DMT1 and Fpn, and their working relationship with other components of the iron-absorptive machinery. Recently, we found that calcium is a low-affinity noncompetitive inhibitor—but not a transported substrate—of DMT1, explaining in part the effect of high dietary calcium on iron bioavailability [Shawki and Mackenzie, 2010]. We have also characterized the Zrt-, Irt-like protein-14 (Zip14) and found it to be a broad-scope metal-ion transporter that could participate in uptake of nontransferrin-bound iron (NTBI) characteristic of iron-overload disorders [Pinilla-Tenas et al, 2011]. We hope to gain a better understanding of how these transporters function in diverse environments and how they may contribute to the etiology of iron-overload disorders. We expect these studies will help drive the development of novel approaches to improve metal-ion nutrition, and treat iron overload or heavy-metal intoxication.

The approaches we use include the voltage clamp, radiotracer assays and fluorescence-based assays in RNA-injected Xenopus oocytes, together with the use of genetically-modified animal models (including the intestine-specific DMT1 knockout mouse, and intestinal Na⁺/H⁺ exchanger nulls).

DMT1-mediated Fe²⁺ transport is energized by the H⁺ electrochemical gradient, placing DMT1 in a large and important class of membrane proteins we call cotransporters. Mammalian cotransporters use Na⁺ or H⁺ electrochemical gradients as the energy source to drive uphill (concentrative) transport of a broad range of nutrients or solutes. Conventional cotransporter models are deterministic—they comprise a series of ligand-induced conformational changes that result in alternating access to a substrate-binding region, and ion-coupling stoichiometries are fixed. Experimental data for DMT1, however, are difficult to reconcile with such a model. When we expressed wildtype mammalian DMT1 or mutant proteins in Xenopus oocytes, we observed significant uncoupled fluxes ('slippage') both of H⁺ and Fe²⁺ [Mackenzie et al, 2006] and variable stoichiometry [Shawki et al, 2008]. Among our present goals, the development of a new model that can account for such slippage will better direct experimental investigation into the molecular mechanisms of DMT1. Among the other cotransporters we are studying are the Na⁺-coupled ascorbic acid (vitamin C) transporter SVCT1 [Mackenzie et al, 2008] and the System A family of Na⁺-coupled neutral amino acid transporters.

• Jorge J Pinilla-Tenas, Brian K Sparkman, Ali Shawki, Anthony C Illing, Colin J Mitchell, Ningning Zhao, Juan P Liuzzi, Robert J Cousins, Mitchell D Knutson, and Bryan Mackenzie (2011) Zip14 is a complex broad-scope metal-ion transporter whose functional properties support roles in the cellular uptake of zinc and nontransferrin-bound iron. Am J Physiol Cell Physiol, 301, C862–C871.
  View original publication at AJP:Cell Online.
• Ali Shawki and Bryan Mackenzie (2010) Interaction of calcium with the human divalent metal-ion transporter-1. Biochem Biophys Res Comm 393, 471–475.
  View original publication at ScienceDirect.
• Bryan Mackenzie, Anthony C Illing and Matthias A Hediger (2008) Transport model of the human sodium-coupled L-ascorbic acid (vitamin C) transporter SVCT1. Am J Physiol Cell Physiol, 294, C451–C459.
  View original publication at AJP:Cell Online.
• Bryan Mackenzie, Hitomi Takanaga, Nadia Hubert, Andreas Rolfs and Matthias A Hediger (2007) Functional properties of multiple isoforms of human divalent metal-ion transporter 1 (DMT1). Biochem J, 403, 59–69.
  View original publication at PortlandPress.com.
• Bryan Mackenzie, M L Ujwal, Min-Hwang Chang, Michael F Romero and Matthias A Hediger (2006) Divalent metal-ion transporter DMT1 mediates both H⁺-coupled Fe²⁺ transport and uncoupled fluxes. Pflügers Arch Eur J Physiol 451, 544–558.
  View original publication at SpringerLink.com.
• Bryan Mackenzie and Michael D Garrick (2005) Iron Imports. II. Iron uptake at the apical membrane in the intestine. Am J Physiol Gastrointest Liver Physiol 289, G981–G986. [Review]
  View original publication at AJP:GI Online.

• Ali Shawki, Anthony C Illing and Bryan Mackenzie (2008) Molecular impact of divalent metal-ion transporter (DMT1) mutations (V114del and G212V) found in a compound heterozygote with microcytic anemia and hepatic iron overload. FASEB J 22, 1192.3. [Abstract]
  View abstract at FASEB Journal Online.
• Ali Shawki, Anthony C Illing and Bryan Mackenzie (2006) Molecular impact of human divalent metal-ion transporter DMT1 mutations associated with disease phenotypes. FASEB J 20, A1278. [Abstract]
  View abstract at FASEB Journal Online.

• Bryan Mackenzie and Matthias A Hediger (2004) Molecular physiology of the H⁺-coupled metal-ion transporter DMT1, in: The Nramp Family, Mathieu Cellier and Philippe Gros (Eds), Eurekah.com and Kluwer Academic / Plenum Publishers, Georgetown. [Review]
  View electronic article at Eurekah.com.
• Bryan Mackenzie (1999) Selected techniques in membrane transport, ch 11 in: Biomembrane Transport, Lon J Van Winkle (Ed), Academic Press, San Diego, pp 327–342. [Review]
  View pdf version of this article © 1999 Academic Press.
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• Bryan Mackenzie, Aamir Ahmed and Michael J Rennie (1992) Muscle amino acid metabolism and transport, in: Mammalian Amino Acid Transport: Mechanisms and Control, Michael S Kilberg and Dieter Häussinger (Eds), Plenum Press, New York, pp 195–231. [Review]
  Buy the book at Amazon.com.