Role of the Molybdoflavoenzyme Aldehyde Oxidase Homolog 2 in the Biosynthesis of Retinoic Acid: Generation and Characterization of a Knockout Mouse

Abstract

The mouse aldehyde oxidase AOH2 (aldehyde oxidase homolog 2) is a molybdoflavoenzyme. Harderian glands are the richest source of AOH2, although the protein is detectable also in sebaceous glands, epidermis, and other keratinized epithelia. The levels of AOH2 in the Harderian gland and skin are controlled by genetic background, being maximal in CD1 and C57BL/6 and minimal in DBA/2, CBA, and 129/Sv strains. Testosterone is a negative regulator of AOH2 in Harderian glands. Purified AOH2 oxidizes retinaldehyde into retinoic acid, while it is devoid of pyridoxal-oxidizing activity. Aoh2 ؊/؊ mice, the first aldehyde oxidase knockout animals ever generated, are viable and fertile. The data obtained for this knockout model indicate a significant role of AOH2 in the local synthesis and biodisposition of endogenous retinoids in the Harderian gland and skin. The Harderian gland's transcriptome of knockout mice demonstrates overall downregulation of direct retinoid-dependent genes as well as perturbations in pathways controlling lipid homeostasis and cellular secretion, particularly in sexually immature animals. The skin of knockout mice is characterized by thickening of the epidermis in basal conditions and after UV light exposure. This has correlates in the corresponding transcriptome, which shows enrichment and overall upregulation of genes involved in hypertrophic responses.Aldehyde oxidases (AOXs) (EC 1.2.3.1) are structurally conserved proteins belonging to the family of molybdoflavoenzymes along with xanthine oxidoreductase (XOR), the key enzyme in the catabolism of purines (25,28). In their catalytically active form, both AOXs and XORs are dimers of identical subunits characterized by three conserved domains separated by nonconserved hinge regions (28). The amino-terminal 25-kDa domain contains two nonidentical 2Fe-2S redox centers. The flavin adenine dinucleotide binding region is located in the intermediate 45-kDa domain, while the substrate and the molybdopterin cofactor binding pocket reside in the carboxy-terminal 85-kDa domain (28). AOXs have broad substrate specificity, hydroxylating N-heterocycles or oxidizing aliphatic as well as aromatic aldehydes into the corresponding acids (35,41,42).The primary structures of mammalian AOXs and XORs are more than 40% identical, and the two types of enzymes are evolutionary related. Most of the available data indicate that AOXs evolved from a primordial form of XOR through a series of gene duplication events (28). In mammals, the number of AOX genes is variable and species specific (25,28,37,65). Rodents have four AOX genes (Aox1, Aoh1, Aoh2, and Aoh3) which are the products of asynchronous duplication events and cluster at a short distance from one another on the same chromosome (chromosome 1c1 in mice and chromosome 9 in rats) (Fig. 1). The Aoh1, Aoh2, and Aoh3 genes are also known as Aox3, Aox4, and Aox3l, respectively, and are currently designated with the latter names in the NCBI database.(The designation of the four mouse AOX genes as Aox1, Aox3, Aox4, and Aox3l in...

Aldehyde Oxidase Activity in Fresh Human Skin

“…Observed interindividual variability is likely caused by the differences in the expression levels of human AO or by a number of single nucleotide polymorphisms recently observed in the human AOX1 gene (Hartmann et al, 2012). The relatively robust activity of AO in the human skin, despite the differences in donor demographics and anatomic region of the skin, may be related to the physiologic role of AO in the metabolism of endogenous retinoids (Tomita et al, 1993;Graessler and Fischer, 2007;Terao et al, 2009). A recent study in AO homolog 2 -/ -knockout mice observed thickening of the epidermis in basal conditions and after UV light exposure, probably triggered by local deficiency of all-trans retinoic acid (Terao et al, 2009).…”

“…To our best knowledge, this is the first report of AO enzyme activity in the human skin. Terao et al (2009) recently reported activity of AO homolog 2 in the skin of mice, whereas Ueda et al (2005) investigated AO-catalyzed reduction of nitro-polycyclic aromatic hydrocarbons in skin samples from hamster, rabbit, guinea pig, mouse, and rat. Although the expression of AO was also reported in human adipose tissue (Weigert et al, 2008), the underlying fat was carefully removed from tested human skin samples, thus eliminating the possibility for a misinterpretation of results.…”

“…In addition, as the enzyme name suggests, AO oxidizes aldehydes to carboxylic acids, a role probably related to conversion of endogenous retinaldehyde (retinal) into retinoic acid (Graessler and Fischer, 2007;Terao et al, 2009). Although AO drug metabolism is relevant for pharmacotherapy, many researchers failed to predict high in vivo AO metabolic clearance based on in vitro results, leading to notable drug failures, such as carbazeran (Kaye et al, 1984), BIBX1382 (Dittrich et al, 2002), zoniporide , RO1 (Zhang et al, 2011), SGX523 (Diamond et al, 2010), and FK3453 (Akabane et al, 2011).…”

Prediction of Human Metabolism of FK3453 by Aldehyde Oxidase Using Chimeric Mice Transplanted with Human or Rat Hepatocytes

“…AOX4 is highly presented in the Harderian gland and skin of mice, is absent in humans (Garattini et al, 2009;Terao et al, 2009). Evaluation of extrahepatic metabolism, which shows species differences, may increasingly improve the predictability of AO metabolism in human.…”

Inhibition of Xanthine Oxidase by the Aldehyde Oxidase Inhibitor Raloxifene: Implications for Identifying Molybdopterin Nitrite Reductases

“…Adding to the frustration, XOR tissue-specific conditional knockouts are currently not available while global XOR −/− and XOR +/− mice experience alterations in nutrient absorption and elevated plasma hypoxanthine levels resulting in death from kidney failure before 6 weeks of age [9,10]. As for AO, there is only one report demonstrating successful knockout of one homologue of AO (aldehyde oxidase homologue 2, Aoh2) expressed primarily in the epithelium [11]. The current absence of knockout strategies to interrogate these molybdopterin enzymes has relegated investigators to employ pharmacologic means to conduct proof of principle experimentation regarding contributory roles mediating the effects of treatment.…”