Specific accumulation of CYP94A1 transcripts after exposure to gaseous benzaldehyde: Induction of lauric acid o-hydroxylase activity in Vicia sativa exposed to atmospheric pollutants P.E. Kastner a,b , S. Le Calve ´ b , L. Diss a,b , V. Sauveplane a , R. Franke c , L. Schreiber c , F. Pinot a,n a Institut de Biologie Mole ´culaire des Plantes, CNRS - Universite´ de Strasbourg IBMP-UPR 2357, De´partement Re´seaux Me´taboliques, 28 rue Goethe, F-67083 Strasbourg Cedex, France b Laboratoire des Mate´riaux, Surfaces et Proce´de´s pour la Catalyse (LMSPC)/CNRS and Universite´ de Strasbourg, 25 rue Becquerel, 67087 Strasbourg Cedex 02, France c Institute of Cellular and Molecular Botany (IZMB), Department of Ecophysiology, University of Bonn, Kirschalle 1, D-53115 Bonn, Germany article info Article history: Received 23 March 2010 Received in revised form 20 July 2010 Accepted 30 September 2010 Keywords: Plant Cytochrome P450 Ozone Induction Pollutant Atmosphere abstract The effects of air pollutants such as aldehydes, ozone, nitrogen dioxide and benzene on fatty acid o- hydroxylase activity in Vicia sativa microsomes have been investigated. Four days old etiolated V. sativa seedlings were exposed to different concentrations of selected pollutants for varying exposure times. Growing etiolated V. sativa seedlings in air containing the gaseous benzaldehyde (150 nM) led to an 8-fold enhancement of lauric acid o-hydroxylase activity in microsomes of treated plants compared to controls grown in pure air (96 710 versus 12 72 pmol/min/mg protein, respectively). The induction increased with increasing gas phase concentrations (10–1300 nM) and the maximum of activity was measured after 48 h of exposure. Northern blot analysis revealed that this induction occurred via transcriptional activation of the gene coding for CYP94A1. The absence of CYP94A2 and CYP94A3 transcription activation together with the missing effect on epoxide hydrolases activities indicate the specificity of CYP94A1 induction by benzaldehyde. Exposure to nitrogen dioxide, ozone and formaldehyde also stimulated lauric acid o-hydroxylases activity while exposure to benzene did not show any effect. & 2010 Elsevier Inc. All rights reserved. 1. Introduction In all organisms, oxidized fatty acids exhibit diverse and fundamental biological activities (Funk, 2001; Blee, 2002; Noverr et al., 2003). Therefore, fatty acid-oxidizing enzymes which generate these compounds have been extensively studied. Cytochrome P450s play a crucial role in these reactions of oxidation. The cytochrome P450 superfamily represents a highly diversified set of heme- containing proteins found in bacteria, fungi, plants and animals (Werck-Reichhart and Feyereisen, 2000). The ‘‘arachidonic cascade’’ in animals leads to the production of hydroxylated and epoxidized derivatives formed mainly in reactions catalyzed by cytochrome P450s and represents one of the most studied biological pathway. In animals, members of the CYP4A subfamily catalyze o-hydroxylation of fatty acids which could represent the first step of their elimination by b-oxidation (Simpson, 1997; Ferdinandusse et al., 2004). However, some data describing biological effects of o-hydroxy derivatives of fatty acids support the hypothesis that o-hydroxylation is not only involved in catabolism (Alonso-Galicia et al., 1998; Birks et al., 1997; Nowicki et al., 1997; Messer-Letienne et al., 1999). In plants fatty acids are also metabolized by cytochrome P450 dependent oxygenases which exhibit diverse catalytic capabilities. We recently characterized CYP77A4 from Arabidopsis thaliana which is the first plant cytochrome P450 able to epoxidize a free fatty acid (Sauveplane et al., 2009). Its physiological role remains to be established. CYP94A5 from tobacco (Le Bouquin et al., 2001) and CYP94C1 from Arabidopsis are able to catalyze the full conversion of a mono to a dicarboxylic fatty acid. Similarly to mammalian cytochrome P450s (Sanders et al., 2005, 2006) these two plant enzymes could be implicated in fatty acid catabolism via b-oxidation of dicarboxylic fatty acids in peroxisomes (Gerhardt, 1992; Poirier et al., 1999). Concerning cytochrome P450 dependent fatty acid hydroxylases (FA hydroxylases), one can distinguish FA o-hydroxylases and FA in-chain hydroxylases which attack the terminal and subterminal position, respectively. So far the majority of the work was performed in the context of plant envelopes cutin and suberin synthesis and has mainly addressed o-hydroxylases represented in CYP86 and CYP94 families (Kandel et al., 2006). The key role of o-hydroxylases in plant envelopes synthesis was demonstrated by reverse genetic: cuticle of A. thaliana exhibits drastic alteration when coding sequence of two fatty acid hydroxylases CYP86A8 and CYP86A2 are modified (Wellesen et al., Contents lists available at ScienceDirect journal homepage: www.elsevier.com/locate/envres Environmental Research 0013-9351/$ - see front matter & 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.envres.2010.09.014 n Corresponding author at: IBMP-CNRS UPR 2357, Institut de Botanique, 28 rue Goethe, F-67083 Strasbourg Cedex, France. E-mail address: franck.pinot@ibmp-cnrs.unistra.fr (F. Pinot). Please cite this article as: Kastner, P.E., et al., Specific accumulation of CYP94A1 transcripts after exposure to gaseous benzaldehyde: Induction of lauric acid o-hydroxylase activity in Vicia sativa exposed to.... Environ. Res. (2010), doi:10.1016/j.envres.2010.09.014 Environmental Research ] (]]]]) ]]]–]]]