Funk C D, Keeney D S, Oliw E H, Boeglin W E, Brash A R
Department of Pharmacology, Vanderbilt University, Nashville, Tennessee 37232, USA.
J Biol Chem. 1996 Sep 20;271(38):23338-44. doi: 10.1074/jbc.271.38.23338.
Three distinct murine lipoxygenase genes have been functionally characterized: 5-lipoxygenase (Chen, X.-S., Naumann, T. A., Kurre, U. , Jenkins, N. A., Copeland, N. G., and Funk, C. D. (1995) J. Biol. Chem. 270, 17993-17999), platelet-type 12-lipoxygenase and leukocyte-type 12-lipoxygenase (Chen, X.-S., Kurre, U., Jenkins, N. A., Copeland, N. G., and Funk, C. D. (1994) J. Biol. Chem. 269, 13979-13987). Here, we describe the cloning and functional characterization of a fourth lipoxygenase gene in mice. Using a polymerase chain reaction-based approach together with partial sequence information from a genomic clone, we isolated a novel lipoxygenase cDNA from the RNA of 3-6-day-old mouse epidermis. The open reading frame predicts a 662-amino acid lipoxygenase that displays 60% identity with both murine 12-lipoxygenase isozymes and 40% identity to 5-lipoxygenase; the sequence is identical to a genomic sequence reported recently (van Dijk, K. W., Steketee, K., Havekes, L., Frants, R., and Hofker, M. (1995) Biochim. Biophys. Acta 1259, 4-8). A full-length clone was expressed in human embryonic kidney 293 cells and homogenates from disrupted cells produced 12-hydroxyeicosatetraenoic acid (12-HETE) and minor amounts of 15-HETE from arachidonic acid. Chiral phase analysis indicated that the 12-HETE is exclusively the 12S enantiomer. In situ hybridization revealed highly specific expression of epidermal lipoxygenase in differentiated keratinocytes of the epidermis and in restricted regions of the root sheath and bulb of hair follicles. High expression was also detected in conjunctiva of the eyelid and in cells of Meibomian and preputial (sebaceous) glands. A 2. 4-kilobase mRNA was detected in mouse epidermis by Northern blot analysis and its abundance was not affected by phorbol ester treatment. The epidermal lipoxygenase gene (Aloxe) resides on mouse chromosome 11 closely linked with the two 12-lipoxygenase genes (Alox12p and Alox12l).
5-脂氧合酶（陈，X.-S.，瑙曼，T.A.，库雷，U.，詹金斯，N.A.，科普兰，N.G.，和芬克，C.D.（1995年）《生物化学杂志》270，17993 - 17999）、血小板型12-脂氧合酶和白细胞型12-脂氧合酶（陈，X.-S.，库雷，U.，詹金斯，N.A.，科普兰，N.G.，和芬克，C.D.（1994年）《生物化学杂志》269，13979 - 13987）。在此，我们描述了小鼠中第四个脂氧合酶基因的克隆及功能鉴定。利用基于聚合酶链反应的方法并结合基因组克隆的部分序列信息，我们从小鼠3至6日龄表皮的RNA中分离出一个新的脂氧合酶cDNA。开放阅读框预测一个662个氨基酸的脂氧合酶，它与小鼠的两种12-脂氧合酶同工酶具有60%的同一性，与5-脂氧合酶具有40%的同一性；该序列与最近报道的一个基因组序列相同（范·迪克，K.W.，斯特克泰，K.，哈韦克斯，L.，弗兰茨，R.，和霍夫克，M.（1995年）《生物化学与生物物理学报》1259，4 - 8）。一个全长克隆在人胚肾293细胞中表达，破碎细胞的匀浆从花生四烯酸产生12-羟基二十碳四烯酸（12-HETE）和少量的15-HETE。手性相分析表明12-HETE完全是12S对映体。原位杂交显示表皮脂氧合酶在表皮分化的角质形成细胞以及毛囊根鞘和毛球的特定区域高度特异性表达。在眼睑结膜以及睑板腺和包皮（皮脂腺）细胞中也检测到高表达。通过Northern印迹分析在小鼠表皮中检测到一个2.4千碱基的mRNA，其丰度不受佛波酯处理的影响。表皮脂氧合酶基因（Aloxe）位于小鼠11号染色体上，与两个12-脂氧合酶基因（Alox12p和Alox12l）紧密连锁。
