Short sequence-paper Molecular cloning and expression patterns of three putative functional aldehyde oxidase genes and isolation of two aldehyde oxidase pseudogenes in tomato 1 Xiangjia Min a ; *, Kazunori Okada a , Barbara Brockmann a , Tomokazu Koshiba b , Yuji Kamiya a a Plant Function Laboratory, RIKEN (The Institute of Physical and Chemical Research), Saitama 351-0198, Japan b Department of Biology, Tokyo Metropolitan University, Tokyo 192-0397, Japan Received 6 June 2000; received in revised form 18 July 2000; accepted 27 July 2000 Abstract The final steps in the biosynthesis of the plant hormones abscisic acid (ABA) and indole-3-acetic acid (IAA) have been shown to be catalyzed by aldehyde oxidases (AO). We have cloned three putative functional AO genes (TAO1, TAO2 and TAO3) and two putative AO pseudogenes (TAO4 and TAO5) in tomato. The TAO1 cDNA described here includes the correct amino terminus of the encoded TAO1 protein and is different at the 5P-end from the TAO1 sequence in GenBank (accession number U82558). Northern analysis shows that TAO1 is expressed mainly in vegetative tissues and TAO2 is expressed in both vegetative and reproductive tissues. TAO3 expression was not detectable by Northern hybridization. These results suggest that each AO may play different roles in the regulation of tomato growth and development. ß 2000 Elsevier Science B.V. All rights reserved. Keywords : Aldehyde oxidase ; Abscisic acid ; Indole-3-acetic acid ; (Tomato) The plant hormone abscisic acid (ABA) plays important roles in plant adaptation to variable environments and in seed dormancy and germination. The functions of ABA in the control of leaf stomata opening and closing and seed dormancy are clearly illustrated by ABA de¢cient mutants [1]. In tomato, three ABA de¢cient mutants, namely nota- bilis (not), £acca (£c), and sitiens (sit), have been known for over 40 years [2]. The recent cloning of a putative 9- cis-epoxy-carotenoid dioxygenase (LeNCED1) gene in to- mato and analysis of the not mutant revealed that LeNCED1 was encoded by the NOT gene, and a base- pair deletion in the coding region of the NOT gene re- sulted in a frame-shift mutation in the not mutant [3]. Biochemical characterization of £c and sit showed that both mutants are defective in the last step of ABA biosyn- thesis in which abscisic aldehyde is oxidized to form ABA by a molybdenum cofactor (Moco)-containing aldehyde oxidase (EC 1.2.3.1; AO) [4]. In £c leaves, the activities of both AO and another Moco-containing enzyme xan- thine dehydrogenase (EC 1.1.1.204 ; XD) were not detect- able [5,6]. Thus, the FLC gene is thought to be involved in Moco biosynthesis while SIT might be the structural gene of abscisic aldehyde oxidase [5]. However, the FLC and SIT genes have not been cloned so far. Only recently, one of four known Arabidopsis AO genes, AAO3, was found to encode an abscisic aldehyde oxidase [7]. The biosynthesis of plant hormone indole-3-acetic acid (IAA) was also sug- gested to be catalyzed by an aldehyde oxidase [8^11]. The protein encoded by AAO1 in Arabidopsis, AAOK, with a high a¤nity for indole-3-acetaldehyde, is highly expressed in an IAA overproducing sur1 mutant [10]. At present, only one AO gene from tomato (TAO1) has been reported but it is not likely to be the SIT gene since TAO1 was mapped to chromosome 11 [12] and the SIT gene is known to be on chromosome 1 [13]. In this paper we describe the molecular cloning and characterization of other members of the AO gene family in tomato in an attempt to isolate the SIT gene. 0167-4781 / 00 / $ ^ see front matter ß 2000 Elsevier Science B.V. All rights reserved. PII:S0167-4781(00)00190-1 * Corresponding author. Present address: Department of Forest Sci- ence, University of British Columbia, Vancouver, BC V6T 1Z4, Canada. Fax : +1-604-822-9102 ; E-mail : xiangjia@interchange.ubc.ca 1 The nucleotide sequence data reported will appear in the GenBank under accession numbers: AF258808 (TAO1 cDNA), AF258809 (TAO2 cDNA), AF258810 (TAO3 cDNA), AF258811 (TAO4 cDNA), AF258812 (TAO5 cDNA), AF259793 (TAO1 partial genomic DNA). Biochimica et Biophysica Acta 1493 (2000) 337^341 www.elsevier.com/locate/bba