The long-term objective of this application is to elucidate the roles of two lipoxygenase (LOX) enzymes and their essential fatty acid (EFA) substrate in forming the water impermeable barrier of the epidermis, defects in which result in ichthyoses and contribute to atopic dermatitis. 12R-Lipoxygenase (12R-LOX) and Epidermal Lipoxygenase-3 (eLOX3) are the only human lipoxygenases that, if mutated to inactive forms, result in a disease, a form of autosomal recessive congenital ichthyosis (ARCI). The fact that the characteristic scaly skin of ARCI is also manifest in essential fatty acid (EFA) deficiency, suggests a possible substrate-enzyme relationship between EFA and the two LOX enzymes. We will test a new hypothesis that connects EFA, 12R- LOX and eLOX3, and the structure of the epidermal water barrier. We propose that the two lipoxygenases oxygenate a critical form of EFA, linoleate esterified to the omega-hydroxyl of the very long chain fatty acid of the unique epidermal acylceramides. We further propose that this oxygenation is required to facilitate hydrolysis of the (oxidized) linoleate moiety and leave the very long chain fatty acid omega-hydroxyl free for coupling to the cross-linked proteins of the corneocyte envelope, a vital step in sealing the water barrier.
The Specific Aims are (1) To characterize 12R-LOX and eLOX3 activity in the epidermis in vivo using animal models, (2) To assess the efficacy of LOX products and ceramides in rescuing the phenotype in murine LOX-/- models, (3) To characterize 12R-LOX and eLOX3 activity in human epidermis including in ARCI patients, and (4) To characterize the putative oxidant-sensitive step of acylceramide ester hydrolysis. The results of this study wil piece together an important facet of construction of the epidermal water barrier and allow a rational approach to the use of oxidized linoleate and its analogs for future therapeutic interventions in the LOX- dependent classes of ichthyoses. If the physiology of the barrier is properly understood, this constitutes a firm foundation for the rational design of any barrier-related therapeutics, and it is to rationalize the role of these well-known key components of the epidermal water barrier that is the main goal of this project.
The scaly skin diseases of ichthyosis afflict families with inherited mutations in skin enzymes that are normally involved in keeping the skin watertight. This study seeks to clarify how lipid enzymes called lipoxygenases are involved in forming this water barrier. Understanding this will help explain how other enzymes cooperate in the process and allow for rational treatment of the ichthyoses.
|Chiba, Takahito; Thomas, Christopher P; Calcutt, M Wade et al. (2016) The Precise Structures and Stereochemistry of Trihydroxy-linoleates Esterified in Human and Porcine Epidermis and Their Significance in Skin Barrier Function: IMPLICATION OF AN EPOXIDE HYDROLASE IN THE TRANSFORMATIONS OF LINOLEATE. J Biol Chem 291:14540-54|
|Newcomer, Marcia E; Brash, Alan R (2015) The structural basis for specificity in lipoxygenase catalysis. Protein Sci 24:298-309|
|MuÃ±oz-Garcia, AgustÃ; Thomas, Christopher P; Keeney, Diane S et al. (2014) The importance of the lipoxygenase-hepoxilin pathway in the mammalian epidermal barrier. Biochim Biophys Acta 1841:401-8|
|Thomas, Christopher P; Boeglin, William E; Garcia-Diaz, Yoel et al. (2013) Steric analysis of epoxyalcohol and trihydroxy derivatives of 9-hydroperoxy-linoleic acid from hematin and enzymatic synthesis. Chem Phys Lipids 167-168:21-32|
|Zheng, Yuxiang; Yin, Huiyong; Boeglin, William E et al. (2011) Lipoxygenases mediate the effect of essential fatty acid in skin barrier formation: a proposed role in releasing omega-hydroxyceramide for construction of the corneocyte lipid envelope. J Biol Chem 286:24046-56|
|Zheng, Yuxiang; Brash, Alan R (2010) Dioxygenase activity of epidermal lipoxygenase-3 unveiled: typical and atypical features of its catalytic activity with natural and synthetic polyunsaturated fatty acids. J Biol Chem 285:39866-75|
|Zheng, Yuxiang; Brash, Alan R (2010) On the role of molecular oxygen in lipoxygenase activation: comparison and contrast of epidermal lipoxygenase-3 with soybean lipoxygenase-1. J Biol Chem 285:39876-87|
|Zheng, Yuxiang; Brash, Alan R (2010) Formation of a cyclopropyl epoxide via a leukotriene A synthase-related pathway in an anaerobic reaction of soybean lipoxygenase-1 with 15S-hydroperoxyeicosatetraenoic acid: evidence that oxygen access is a determinant of secondary reactions with fatty ac J Biol Chem 285:13427-36|
|Yu, Zheyong; Schneider, Claus; Boeglin, William E et al. (2007) Epidermal lipoxygenase products of the hepoxilin pathway selectively activate the nuclear receptor PPARalpha. Lipids 42:491-7|
|Brash, Alan R; Yu, Zheyong; Boeglin, William E et al. (2007) The hepoxilin connection in the epidermis. FEBS J 274:3494-502|
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