CYP703 Is an Ancient Cytochrome P450 in Land Plants Catalyzing in-Chain Hydroxylation of Lauric Acid to Provide Building Blocks for Sporopollenin Synthesis in Pollen

Abstract

CYP703 is a cytochrome P450 family specific to land plants. Typically, each plant species contains a single CYP703. Arabidopsis thaliana CYP703A2 is expressed in the anthers of developing flowers. Expression is initiated at the tetrad stage and restricted to microspores and to the tapetum cell layer. Arabidopsis CYP703A2 knockout lines showed impaired pollen development and a partial male-sterile phenotype. Scanning electron and transmission electron microscopy of pollen from the knockout plants showed impaired pollen wall development with absence of exine. The fluorescent layer around the pollen grains ascribed to the presence of phenylpropanoid units in sporopollenin was absent in the CYP703A2 knockout lines. Heterologous expression of CYP703A2 in yeast cells demonstrated that CYP703 catalyzes the conversion of medium-chain saturated fatty acids to the corresponding monohydroxylated fatty acids, with a preferential hydroxylation of lauric acid at the C-7 position. Incubation of recombinant CYP703 with methanol extracts from developing flowers confirmed that lauric acid and in-chain hydroxy lauric acids are the in planta substrate and product, respectively. These data demonstrate that in-chain hydroxy lauric acids are essential building blocks in sporopollenin synthesis and enable the formation of ester and ether linkages with phenylpropanoid units. This study identifies CYP703 as a P450 family specifically involved in pollen development.

The Gibberellin perception system evolved to regulate a pre-existing GAMYB-mediated system during land plant evolution

“…2c-e), where the sporopollenin material is produced for the outer wall development [9][10][11][21][22][23] . Considering that GAMYBs, CYP703As and GA biosynthesis genes in flowering plants are also co-expressed in the tapetal cells and microspores 1,8,12,[24][25][26] , these observations indicate that SmGAMYB activity in these tissues of Selaginella is required to mediate GA signalling during the outer wall formation of microspores.…”

“…In rice, CYP703A3 is an essential enzyme in sporopollenin biosynthesis, and its expression is positively regulated by a transcription factor, GAMYB, which is controlled by the GA signalling pathway 1,7,8 . Previous anatomical and in silico studies suggest that, similar to flowering plants, present-day bryophytes, including mosses, and lycophytes (lycopods) contain sporopollenin-containing outer walls on the surface of their spores and microspores [9][10][11] , and genes encoding CYP703 homologues have been detected in these genomes 12 . Curiously, however, one of the basal vascular plants, Selaginella, contains a functional GA perception system that is mediated by a GA receptor, GIBBERELLIN-INSENSITIVE DWARF1 (GID1), and suppressor proteins (DELLAs), whereas one of the basal land plants, Physcomitrella patens, does not contain this system [13][14][15][16] .…”

The Cytochrome P450 Enzyme CYP96A15 Is the Midchain Alkane Hydroxylase Responsible for Formation of Secondary Alcohols and Ketones in Stem Cuticular Wax of Arabidopsis

“…v-1 or v-2; Whyte et al, 1998;van Beilen et al, 2003;Kandel et al, 2005Kandel et al, , 2006. To date, only one other P450 enzyme with in-chain hydroxylase activity has been described and it prefers C12 fatty acid substrates very different from the VLC alkane substrates of MAH1 (Morant et al, 2007). The ability of MAH1 to catalyze midchain alkane hydroxylation suggests a unique active-site morphology for this enzyme, a unique folding of the alkane substrate, or perhaps a combination of these two conditions.…”

PERSISTENT TAPETAL CELL1Encodes a PHD-Finger Protein That Is Required for Tapetal Cell Death and Pollen Development in Rice

“…Lipidic pollen exine is made of sporopollenin, an extremely resilient material derived from the polymerization of fatty acid metabolites and phenolic acid (Piffanelli et al, 1998;Morant et al, 2007). Even though the pollen grains of rice and Arabidopsis have similar pollen exine layers, rice has a distinct pollen wall ontology compared with Arabidopsis.…”