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Lipoxygenases (LO) are a family of non-heme iron containing proteins that stereospecifically insert molecular oxygen into 1, 4-cis, cis-pentadiene containing polyunsaturated fatty acids’. They are generally classified according to their positional specificity of arachidonic acid oxygenation and cell specific expression pattern2.6. Lipoxygenase activity gives rise to a number of bioactive lipid products including leukotrienes, lipoxins and HETE’s (hydroxyeicosatetraenoic acid’s).7’8The major lipoxygenase forms include the neutrophil 5-LO, platelet 12-LO and reticulocyte 15LO.2’3’6’9 In recent years the lipoxygenase family has expanded by means of molecular cloning studies to include several new members. An e-12(S)LO (epidermal12(S)lipoxygenase) cDNA was cloned and functionally expressed and its gene isolated10’1. Two other lipoxygenases found in skin have also been cloned, a 15-LO in humans distinct from the reticulocyte enzyme4and a related phorbol ester inducible 8-LO enzyme in mouse skins. All of these lipoxygenases generate HETE metabolites of theS-stereo configuration. However, recently mammalian lipoxygenase cDNAs that generated 12(R) -HETE have been cloned and functionally expressed.12-14The epidermal 12(S) and 12-(R) lipoxygenases have yet to be characterized in any great detail.10,12-14Here, we characterize further the epidermal 12(S) and 12(R)-LO’s and attempt to elucidate the enzyme’s primary structural determinants controlling chirality of oxygen insertion.

Mammalian lipoxygenase enzymes (Figure 1) are derived from a multi-gene family, each member consisting of 14 exons (1,2). The 5-lipoxygenase gene is distinct from other lipoxygenase genes in its large size and its location on a separate chromosome (10q1 1.2 in humans and central chromosome 6 in mice) (1-5). A cluster of lipoxygenase genes located on mouse chromosome 11 (5,6) contains genes that encode three distinct 12(S)-lipoxy-genases, referred to as “platelet-type,” “leukocyte-type” and “epidermal-type” (1,7,8). Although substantial information is known about the ability of the various lipoxygenases to oxygenate arachidonic acid at a specific carbon atom position, relatively little is understood regarding the physiological roles of the various metabolites such as 12-hy-dro(pero)xy-eicosatetraenoic acid (12-H(p)ETE) and 15-H(p)ETE. Diverse biological activities have been assigned to lipoxygenase-derived eicosanoids and pathophysiological elevations in their synthesis have been reported (9-11). However, it remains to be determined how relevant these findings are to physiological models in vivo. Thus, we undertook the approach of targeted lipoxygenase gene disruption to elucidate possible functions associated with the various eicosanoid metabolites generated from these pathways. In this review, an effort will be made to emphasize the current status of experimental results using 5-lipoxygenase and 12-lipoxygenase-deficient mice and potential future applications in disease models and other physiological studies.

The lipoxygenases are a family of enzymes that stereospecifically insert molecular oxygen onto defined positions of arachidonic acid giving rise to various eicosanoids including leukotrienes, lipoxins and hydro(pero)xy-eicosatetraenoic (H(P)ETE) acids (1). There are at least four lipoxygenase (LO) forms in mice, each encoded by a separate gene. The precise roles of the various metabolites are poorly understood in many cases. In our laboratory, we have carried out the cloning and inactivation of three murine LO genes (5-LO, leukocyte-type 12-LO and platelet-type 12-LO) to examine potential biological functions of eicosanoids.

Seven years have elapsed since the cloning of the cDNA for the first mammalian lipoxygenase and the deduced primary structure (Matsumoto et al., 1988; Dixon et al., 1988). Here, a brief comparative overview of the molecular properties of human and mouse lipoxygenases will be presented. Although each of the human lipoxygenases were isolated and systematically characterized prior to the corresponding murine homologs it became necessary to evaluate their relatedness in view of our present targeted lipoxygenase gene inactivation experiments in mice.

Arachidonic acid is converted to an array of potent biological mediators such as prostaglandins, leukotrienes and thromboxane in a tissue and cell specific fashion (1,2). The enzymes that catalyze these transformations, leukocyte 5-lipoxygenase, reticulocyte/eosinophil 15-lipoxygenase, platelet 12-lipoxygenase, prostaglandin G/H synthase (cyclooxygenase) and leukotriene A4 hydrolase have been characterized and their sequences have been deduced by cloning of their respective complementary DNAs (3–10). Besides substrate availability, it has become increasingly evident that transcriptional and translational control as well as “suicide-type” inactivation are critical factors that regulate eicosanoid generation (11). We have now established a quantitative polymerase chain reaction (PCR) assay as a means to assess this regulation at the mRNA level.