Department of Biochemistry
School of Medicine & Biomedical Sciences
Faculty
LAURA GARRICK, Ph.D.
Research Associate Professor
The Role of Divalent Metal Transporter 1 in Iron Uptake
The Belgrade rat has an autosomal recessive mutation (G185R) in divalent metal transporter 1 (DMT1). The mutation inactivates the protein functionally. DMT1 is the major iron transporter in the GI tract and is also involved with iron uptake in most cells of the body. Belgrade rats are severely anemic and are valuable for determining the role of DMT1 in tissues and organs of the whole animal. We breed and supply Belgrade rats to investigators studying the role of DMT1 in iron transport in brain, lung, kidney and central nervous system.
There are four isoforms of DMT1 caused by differential splicing of the mRNA. The N-terminal end of the protein can have either exon 1a or 2 and the C-terminal end can have alternate peptides with an iron response element (+IRE) or without an IRE (-IRE) in the mRNA. Thus the four isoforms are: 1a+IRE, 1a-IRE, 2+IRE and 2-IRE. We have permanently transfected HEK293 cells with 1a+IRE and 2-IRE expression vectors. Cells express the transfected DMT1 in the presence of doxycycline. We are examining the functional characteristics of these two isoforms using 59Fe2+ and 54Mn2+ uptake. Other divalent metals, such as Zn and Co, will also be examined.
We have prepared antibodies that are specific for each of the four isoforms. Using immunofluorescence and confocal microscopy, we have found differential localization of the isoforms in various cell types. In particular, the 2-IRE form is in the nucleus of neuronal and neuronal-like cells. Future experiments will address how this isoform gets to the nucleus, whether the entire protein or a truncated version is in the nucleus, and the function of this membrane protein in the nucleus.
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Click on image to enlarge.
Confocal fluorescence micrographs of primary cultures of rat sympathetic neurons from rat cervical ganglia. The top figure is immunostained with antibodies specific for -IRE DMT1 (red) and transferrin receptor (TfR, green). The bottom figure is immunostained with antibodies specific for +IRE DMT1 (red) and TfR (green). The main images show optical sections with neurites visible. The insets are from a higher plane where the cell body and nucleus are better seen. Note nuclear staining for -IRE DMT1 but not for +IRE DMT1. |
Selected Recent Publications
Garrick, M.D., and Garrick, L.M. (2009) Cellular Iron Transport. Biochim. Biophys. Acta 1790:309-325.
Salazar, J., Mena, N., Hunot, S., Prigent, A., Alvarez-Fischer, D., Arredondo, M., Duyckaerts, C., Sazdovitch, V., Zhao, L., Garrick, L.M., Nuñez, M.T., Garrick, M.D., Raisman-Vozari, R., and Hirsch, E.C. (2008) Divalent Metal Transporter 1 (DMT1) Contributes to Neurodegeneration in Animal Models of Parkinson's Disease. Proc. Natl. Acad. Sci. USA 105:18578-18583.
Collins, J.F., Hu, Z., Ranganathan, P.N., Feng, D., Garrick, L.M., Garrick, M.D., and Browne, R.W. (2008) Induction of Arachidonate 12-Lipoxygenase (Alox15) in Intestine of Iron-Deficient Rats Correlates with the Production of Biologically Active Lipid Mediators. Amer. J. Physiol. GI and Liver Physiol. 294:948-962 doi:10.1152/ajpgi.00274.2007.
Garrick, M.D., and Garrick, L.M. Loss of Rapid Transferrin Receptor Recycling Due to a Mutation in Sec15l1 in hbd Mice. (2007) Biochim. Biophysi. Acta – Molec. Cell Res. 1773:105–108 doi:10.1016/j.bbamcr.2006.09.032.
Ghio, A.J., Turi, J.L., Madden, M.C., Dailey, L.A., Richards, J.D., Stonehuerner, J.G., Morgan, D.L., Singleton, S., Garrick, L.M., and Garrick, M.D. (2007) Lung Injury after Ozone Exposure Is Iron-Dependent. Amer. J. Physiol. Lung Cell. Mol. Physiol. 292: L134-L143 doi:10.1152/ajplung.00534.2005
Garrick, M.D., Kuo, H-C., Vargas, F., Singleton, S.T., Zhao, L., Smith, J.J., Paradkar, P., Roth, J.A., and Garrick, L.M. (2006) Comparison of Mammalian Cell Lines Expressing Distinct Isoforms of Divalent Metal Transporter 1 in a Tetracycline-Regulated Fashion. Biochem. J. 398:539-546 doi: 10.1042/BJ20051987.
Ghio, A.J., Turi, J.L., Yang, F., Garrick, L.M. & Garrick, M.D. (2006) Iron homeostasis in the lung. Biol. Res. 39:67-77.
Garrick, M.D., Singleton, S.T., Vargas, F., Kuo, H-C., Zhao, L., Knöpfel, M., Davidson, T., Costa, M., Paradkar, P., Roth, J.A. & Garrick, L.M. (2006) DMT1: Which metals does it transport? Biol. Res. 39:79-85.
Ghio, A.J.. Wang, X., Dailey, L.A., Stonehuerner, J.D., Piantadosi, C.A., Yang, F., Dolan, K.G., Garrick, L.M., and Garrick, M.D. (2005) DMT1 Decreases Metal-Related Injury in the Lung. Amer. J. Physiol. Lung Cell. Mol. Physiol. 289: L460-L467 doi:10.1152/ajplung.00154.2005.