Iron modulates the membrane-binding and the intracellular trafficking of 5-lipoxygenase: functional implications in neurodegeneration

Iron modulates the membrane-binding and the intracellular trafficking of 5-lipoxygenase: functional implications in neurodegeneration Beatrice Dufrusine1, Andrea Di Francesco1, Annalaura Sabatucci1, Clotilde Beatrice Angelucci2, Sergio Oddi2,3, Claudio D’Addario1,3, Dieter Steinhilber4, Mauro Maccarrone3,5* and Enrico Dainese1,3* 1Faculty of Biosciences, University of Teramo, Teramo, 64100, Italy; 2 Faculty of Veterinary Medicine, University of Teramo, Teramo, 64100, Italy; 3 European Center for Brain Research (CERC)/Santa Lucia Foundation, Rome, Italy; 4 Institute of Pharmaceutical Chemistry, Goethe University Frankfurt, Germany; 5 Center of Integrated Research, Campus Bio-Medico University of Rome, Italy. *Equally senior authors. Introduction 5-lipoxygenase (5-LOX) is a non-heme iron-containing enzyme catalyzing the initial steps in the biosynthesis of leukotrienes, inflammatory eicosanoids that are capable of promoting neurodegeneration. Iron accumulation has been demonstrated in Alzheimer’s disease (AD). Here we present the structure of 5-LOX in solution and a new molecular mechanism throughout the iron content of the enzyme is able to influence its membrane binding and subcellular distribution in THP-1 macrophages. Methods 3D structure in solution and membrane binding experiments were done using Small angle X-ray scattering (SAXS) and fluorescence resonance energy transfer (FRET) approaches, as reported [1]. Confocal fluorescence microscopy, subcellular fractionation and western blot (WB) analysis were carried out as already described [1]. Late Onset AD (LOAD) patients and non-demented healthy subjects (CT) were recruited as described [2]. Results SAXS analysis revealed that 5-LOX in solution is mostly organized as a homodimer. Iron removal from the recombinant human 5-LOX altered the catalytic activity of the enzyme, and impaired its membrane-binding ability. THP-1 cells exposed to increasing amounts of iron showed a redistribution of the cytosolic 5-LOX to the nuclear compartment. Additionally, a significant correlation between plasma levels of the 5-LOX end-product LTB4 and hemoglobin was observed in a population of healthy subjects and AD patients. Conclusions These results suggest that exogenous iron modulates 5-LOX activity by increasing its ability to bind to nuclear membranes, further supporting a new role for iron in inflammation-based neurological diseases where its homeostasis is altered. Acknowledgements. M.M. and E.D. wish to thank EU for granting the Biostruct-X project within the FP VII programme. [1] E. Dainese et al., FASEB J. 24 (2010) 1725-1736. [2] A. Di Francesco et al., JAD 37 (2013) 3-8.