The ADH2 gene codes for the Arabidopsis glutathione-dependent formaldehyde dehydrogenase (FALDH), an enzyme involved in formaldehyde metabolism in eukaryotes. In the present work, we have investigated the potential role of FALDH in detoxification of exogenous formaldehyde. We have generated a yeast (Saccharomyces cerevisiae) mutant strain (sfa1⌬) by in vivo deletion of the SFA1 gene that codes for the endogenous FALDH. Overexpression of Arabidopsis FALDH in this mutant confers high resistance to formaldehyde added exogenously, which demonstrates the functional conservation of the enzyme through evolution and supports its essential role in formaldehyde metabolism. To investigate the role of the enzyme in plants, we have generated Arabidopsis transgenic lines with modified levels of FALDH. Plants overexpressing the enzyme show a 25% increase in their efficiency to take up exogenous formaldehyde, whereas plants with reduced levels of FALDH (due to either a cosuppression phenotype or to the expression of an antisense construct) show a marked slower rate and reduced ability for formaldehyde detoxification as compared with the wild-type Arabidopsis. These results show that the capacity to take up and detoxify high concentrations of formaldehyde is proportionally related to the FALDH activity in the plant, revealing the essential role of this enzyme in formaldehyde detoxification.

A glutathione-dependent formaldehyde dehydrogenase (class I11 alcohol dehydrogenase) has been characterized from Arubidopsis thulium. This plant enzyme exhibits kinetic and molecular properties in common with the class I11 forms from mammals, with a K,, for S-hydroxymethylglutathione of 1.4 pM, an anodic electrophoretic mobility (PI: 5.3-5.6) and a cross-reaction with anti-(rat class I11 alcohol dehydrogenase) antibodies. The enzyme structure, deduced from the cDNA sequence, fits into the complex system of alcohol dehydrogenases and shows that all life forms share the class I11 protein type. The corresponding mRNA is 1.4 kb and present in all plant organs; a single copy of the gene is found in the genome. The class I11 structural variability is different from that of the ethanol-active enzyme types in both vertebrates (class I) and plants (class P), although class P conserves more of the class 111 properties than class I does. Also the enzymatic properties differ between the two ethanol-active classes. Activesite variability and exchanges at essential residues (Leu/Gly57, AsplArgll5) may explain the distinct kinetics. These patterns are consistent with two different metabolic roles for the ethanol-active enzymes, a more constant function, reduction of acetaldehyde during hypoxia, for class P, and a more variable function, the detoxication of alcohols and participation in metabolic conversions, for class I. A sequence motif, Pro-Xaa-IleNal-Xaa-Gly-His-Glu-Xaa-Xaa-Gly, common to all medium-chain alcohol dehydrogenases is defined.

It has recently been discovered that glutathione-dependent formaldehyde dehydrogenase (FALDH) exhibits a strong S-nitrosoglutathione reductase activity. Plants use NO and S-nitrosothiols as signaling molecules to activate defense mechanisms. Therefore, it is interesting to investigate the regulation of FALDH by mechanical wounding and plant hormones involved in signal transduction. Our results show that the gene encoding FALDH in Arabidopsis (ADH2) is down-regulated by wounding and activated by salicylic acid (SA). In tobacco, FALDH levels and enzymatic activity decreased after jasmonate treatment, and increased in response to SA. This is the ¢rst time that regulation of FALDH in response to signals associated with plant defense has been demonstrated. ß

The glutathione-dependent formaldehyde dehydrogenase (FALDH) is the main enzyme of the formaldehyde detoxification system in eukaryotes. In Arabidopsis FALDH is coded by a single gene, which is constitutively expressed [1]. By immunolocalization experiments on Arabidopsis root and leaf sections, we demonstrate that the pattern of expression of the enzyme is cell specific. By using tobacco BY-2 cell cultures we show that FALDH co-localizes with tubulin on the cortical microtubules and the microtubules figures (preprophase band, mitotic spindle and phragmoplast), which suggests a role for FALDH in some plant-specific function during cell division. Overexpression of FALDH in Arabidopsis plants results in a 25% increase in the efficiency of elimination of exogenous formaldehyde, whereas plants with reduced levels of FALDH, bearing antisense constructs, show a reduced ability and slower rate in formaldehyde elimination [2]. These results confirm the central role of FALDH in formaldehyde metabolism in plants and have important implications in the phytoremediation of environmental formaldehyde.The importance of FALDH has been greatly increased by the discovery of its potent activity toward S-nitrosoglutathione, the condensation product of glutathione and nitric oxide (NO) [3][4][5]. NO and NO-related metabolites, such as S-nitrosothiols (SNOs) play a central role in signal transduction and host defense [6]. We have investigated the gene response to mechanical wounding and plant hormones involved in the signal transduction pathway, showing that the gene is down-regulated by wounding in a JA-dependent pathway, and that it is transcriptionally activated by salicylic acid [7]. This is the first time that regulation of FALDH in response to signals associated with plant defense has been demonstrated. References

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