We describe lacerata (lcr) mutants of Arabidopsis, which display various developmental abnormalities, including postgenital organ fusions, and report cloning of the LCR gene by using the maize transposon Enhancer͞Suppressor-mutator (En͞Spm). The pleiotropic mutant phenotype could be rescued by genetic complementation of lcr mutants with the wild-type LCR gene. The LCR gene encodes a cytochrome P450 monooxygenase, CYP86A8, which catalyzes -hydroxylation of fatty acids ranging from C12 to C18:1, as demonstrated by expression of the gene in yeast. Although palmitic and oleic acids were efficient substrates for LCR, 9,10-epoxystearate was not metabolized. Taken together with previous studies, our findings indicate that LCR-dependent -hydroxylation of fatty acids could be implicated in the biosynthesis of cutin in the epidermis and in preventing postgenital organ fusions. Strikingly, the same pathway seems to control trichome differentiation, the establishment of apical dominance, and senescence in plants.T he epidermis of plants is a composite tissue that comprises several cell types. Some of these, such as stoma cells, trichomes, and papilla cells, can be easily distinguished from the predominating pavement cells by their characteristic morphological features. Other cell types are not readily distinguishable, although they apparently perform specific functions. One example of this is given by epidermal cells on the adaxial side of carpels, which exhibit a unique contact response during elaboration of the pistil and are able to adhere and redifferentiate into parenchymatous cells. Another unique feature of these epidermal cells is their ability to adhere to the growing pollen tube and guide it to the embryo sac (1, 2). In contrast to animals, where selectively established cell adhesions are common and play an enormous role in development (3, 4), examples of regular cell adhesions in higher plants are rare, and indeed may be restricted to the processes cited above. In particular mutants in several plant species fusions of organs occur during development of the shoot, in a process that resembles the regular fusion of carpels. It is not yet known whether the same molecular mechanisms underlie all instances of cell fusions. By comparison with the epidermis cells of fused carpels, epidermis cells at sutures in fusion mutants do not alter their normal anticlinal plane of division and do not redifferentiate in response to the adhesion. Cell differentiation, however, is affected in at least two fusion mutants. The epidermis of crinkly4 (cr4) maize plants contains enlarged, occasionally spherical, cells, which can divide periclinally to give rise to multilayered sectors (5). In the fiddlehead ( fdh) mutant of Arabidopsis, the epidermis of rosette leaves displays a 2-fold reduction in the number of trichomes (6). These findings indicate a link between the altered cell differentiation in the epidermis and the fusion of organs in the mutants.By using transposon tagging, FDH and CR4, two genes that result in organ fusions when m...

Suberin composition of various plants including Arabidopsis (Arabidopsis thaliana) has shown the presence of very long chain fatty acid derivatives C20 in addition to the C16 and C18 series. Phylogenetic studies and plant genome mining have led to the identification of putative aliphatic hydroxylases belonging to the CYP86B subfamily of cytochrome P450 monooxygenases. In Arabidopsis, this subfamily is represented by CYP86B1 and CYP86B2, which share about 45% identity with CYP86A1, a fatty acid v-hydroxylase implicated in root suberin monomer synthesis. Here, we show that CYP86B1 is located to the endoplasmic reticulum and is highly expressed in roots. Indeed, CYP86B1 promoter-driven b-glucuronidase expression indicated strong reporter activities at known sites of suberin production such as the endodermis. These observations, together with the fact that proteins of the CYP86B type are widespread among plant species, suggested a role of CYP86B1 in suberin biogenesis. To investigate the involvement of CYP86B1 in suberin biogenesis, we characterized an allelic series of cyp86B1 mutants of which two strong alleles were knockouts and two weak ones were RNA interference-silenced lines. These root aliphatic plant hydroxylase lines had a root and a seed coat aliphatic polyester composition in which C22-and C24-hydroxyacids and a,vdicarboxylic acids were strongly reduced. However, these changes did not affect seed coat permeability and ion content in leaves. The presumed precursors, C22 and C24 fatty acids, accumulated in the suberin polyester. These results demonstrate that CYP86B1 is a very long chain fatty acid hydroxylase specifically involved in polyester monomer biosynthesis during the course of plant development.

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The chemical tagging of a cytochrome P-450-dependent lauric acid omega-hydroxylase from clofibrate-treated Vicia sativa seedlings with [1-14C]11-dodecynoic acid allowed the isolation of a full-length cDNA designated CYP94A1. We describe here the functional expression of this novel P-450 in two Saccharomyces cerevisiae strains overproducing their own NADPH-cytochrome P-450 reductase or a reductase from Arabidopsis thaliana. The results show a much higher efficiency of the yeast strain overproducing the plant reductase compared with the yeast strain overproducing its own reductase for expressing CYP94A1. The methyl end of saturated (from C-10 to C-16) and unsaturated (C18:1, C18:2 and C18:3) fatty acids was mainly oxidized by CYP94A1. Both E/Z and Z/E configurations of 9, 12-octadecadienoic acids were omega-hydroxylated. Lauric, myristic and linolenic acids were oxidized with the highest turnover rate (24 min-1). The strong regioselectivity of CYP94A1 was clearly shifted with sulphur-containing substrates, since both 9- and 11-thia laurate analogues were sulphoxidized. Similar to animal omega-hydroxylases, this plant enzyme was strongly induced by clofibrate treatment. Rapid CYP94A1 transcript accumulation was detected less than 20 min after exposure of seedlings to the hypolipidaemic drug. The involvement of CYP94A1 in the synthesis of cutin monomers and fatty acid detoxification is discussed.