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ContIron is essential for function of numerous proteins in cells, including proteins involved in oxygen transport and electron transfer. Mammalian cells regulate expression of proteins involved in iron uptake and sequestration to ensure that iron supplies are adequate to meet metabolic needs but not sufficient to cause toxic damage. This section focuses on understanding how cells and organisms regulate iron metabolism at the molecular level. Researchers in this section discovered that mammalian cells contain iron-sensing proteins known as iron regulatory proteins (IRP1 and IRP2). In iron-depleted cells, IRPs bind to a specific class of RNA stem-loops in transcripts of iron metabolism proteins known as iron responsive elements (IREs). When IRPs bind to an IRE at the 5' end of a transcript, translation is repressed. When IRPs bind to IREs in the 3' UTR of transferrin receptor, the transcript is protected from degradation.
The figure below shows the NMR solution structure of a typical IRE. The bulged cytosine (C) and the guanosine (G) at the top of the figure are residues that are critical for IRP binding. Structural features of the IRE, including five base-pairs within the upper stem and C-G base-pair in the loop constitute a "molecular ruler" that determines the appropriate distance and spatial orientation between the bulged C and the unpaired loop G.
IRPs sense cytosolic iron levels by different mechanisms. IRP1 is a bifunctional protein that contains an iron-sulfur cluster and functions as an aconitase in iron-replete cells. When IRP1 lacks an iron-sulfur cluster, it binds to IREs and alters expression of ferritin, transferrin receptor, and several other iron metabolism proteins. IRP2 is iron-dependently ubiquitinated and degraded in iron-replete cells. IRP2 contains a 73 amino acid cysteine rich domain that binds elemental iron and undergoes iron-dependent oxidation in iron-replete cells. Both IRPs are found in mammalian cells.
To elucidate the role of each IRP in iron metabolism, we have genetically ablated each IRP in mice. Mice in which IRP2 is genetically ablated are viable and develop normally, but they develop a progressive neurodegenerative disease in adulthood characterized by increased deposition of ferric iron in discrete locations throughout the brain. IRP2 is well expressed in the central nervous system. Characterization of mice that do not express IRP1 is ongoing. Unlike IRP2-/- mice, IRP1-/- mice do not develop neurodegenerative disease.
There are several major ongoing projects in the laboratory. These include cloning of mammalian iron-sulfur cluster assembly proteins and characterization of their mechanisms of function. In addition, the mechanisms by which IRP2 is ubiquitinated and degraded are under study. We have evidence that numerous proteins in mammalian cells undergo iron-dependent oxidation and degradation, and we believe that recognition of oxidized proteins by ubiquitination enzymes may be a major determinant of the intrinsic half-lives of endogenous proteins. We have purified IRP1 and IRP2 and are working to crystallize each IRP. In addition, we are analyzing the mechanisms that regulate transport of iron into the circulation and central nervous system. The IRP1 and IRP2 knockout mice are important research models that enable us to study how iron metabolism is regulated at both the cellular and organismal level. We are intensively studying the neurodegenerative disease of IRP2 knockout mice.
