Cytochromes p450

Abstract

There are 244 cytochrome P450 genes (and 28 pseudogenes) in the Arabidopsis genome. P450s thus form one of the largest gene families in plants. Contrary to what was initially thought, this family diversification results in very limited functional redundancy and seems to mirror the complexity of plant metabolism. P450s sometimes share less than 20% identity and catalyze extremely diverse reactions leading to the precursors of structural macromolecules such as lignin, cutin, suberin and sporopollenin, or are involved in biosynthesis or catabolism of all hormone and signaling molecules, of pigments, odorants, flavors, antioxidants, allelochemicals and defense compounds, and in the metabolism of xenobiotics. The mechanisms of gene duplication and diversification are getting better understood and together with co-expression data provide leads to functional characterization.

Figure 1.

Naming a P450 protein.

Figure 2.

Conserved structures and sequences in P450 proteins. (A) Map of the signature motifs in the P450 proteins. The Glu and Arg of the K-helix consensus sequence (KETLR) and the Arg in the “PERF” consensus sequence form the E-R-R triad. (B) WebLogo of the P450 heme binding motif constructed from A. thaliana P450s. The letter size is proportional to the degree of amino acid conservation. The F, G and C residues are conserved in all plant P450s. The C residue is universally conserved in all P450s across kingdoms and coordinates the iron in the heme. (C) WebLogos of the other conserved structures and sequences in P450 proteins. AG×DT (I-helix), KETRL (K-helix) and PERF/W, respectively.

Figure 3.

Consensus Matching of 4 P450s as ribbon-structures. P450s: CYP3A4 [1TQN, Homo sapiens], CYP2B4 [1PO5, Oryctolagus cuniculus], CYP2C5 [1DT6, Oryctolagus cuniculus], and CYP2C9 [1R9O, Homo sapiens]. Color key: P450 backbone structures, blue; K-helix consensus, grey; PERF consensus, yellow; heme-binding loop, orange; heme, pink.

Figure 4.

The NADPH-dependent cytochrome P450 reductase (CPR) is a membrane bound protein localized in the ER membrane. CPR is a multidomain protein with three cofactor binding domains (FMN, FAD and NADPH) and a linker domain situated between the FMN and FAD/ NADPH domains. CPR donates electrons from the two-electron donor NADPH to the heme of P450 in a coupled two-step reaction as illustrated in picture on the right. NADPH binds to the NADPH-binding domain and electrons are shuttled from NADPH through FAD and FMN to the P450 heme. The CPR structure is color correlated with the reaction mechanism (http://www.p450.kvl.dk/p450rel.shtml).

Figure 5.

Circular cladogram of 246 full-length cytochrome P450s from A. thaliana. Current software and a complete dataset have allowed resolution of the deeper branches in the non-A type P450s, and provide a clearer view of the relationships among the A type P450s (CYP71 clade). The source multiple sequence alignment was performed in MUSCLE <http://www.drive5.com/muscle/>. Parameters were: gap opening penalty, 11; a gap extension penalty, 0.75; center, 0.0; Gonnet matrix. The alignment was refined once (identical parameters). The Neighbor-Join phylogenetic tree was created using MEGA4 <http://www.megasoftware.net/> with the Jones-Taylor-Thornton matrix-based substitution model and a 1000 bootstrap trial test (data not shown).

Figure 6.

Genetic locations of cytochromes P450 in the Arabidopsis genome. Chromosome Maps were made by BLAST of deduced amino acid sequence against whole chromosome sequences. Arrows indicate the relative gene orientation. Putative pseudogenes are shown in italics.

Figure 7.

Neighbor-join bootstrap tree and intron map of the A-type P450s. Bootstrap values are out of 1000 trials. Phase 0 introns: |; Phase 1 introns: [; Phase 2 introns: ]. Phase 0 introns are positioned between two codons, phase 1 introns are positioned after the first base in the scondon, and phase 2 introns after the second base of the condon. P450s lacking an intronmap are intronless.

Figure 8.

Neighbor-join bootstrap tree and intron map of the non-A type P450s. P450 groups thought to comprise individual clades are colored. Bootstrap values are out of 1000 trials. Phase 0 introns: |; Phase 1 introns: [; Phase 2 introns: ]. Phase 0 introns are positioned between two codons, phase 1 introns are positioned after the first base in the condon, and phase 2 introns after the second base of the codon. P450s lacking an intronmap are intronless.

Figure 9.

The reaction catalyzed by CYP51 genes in higher plants. CYP51 is an obtusifiliol 14α-demethylase and catalyzes three successive oxidations of the carbon 32, leading to C-C rupture and release of the methyl in C-14 as formic acid to form 4αmethyl-5αergosta-8,14,24(24¹)trien-3β ol.

Figure 10.

Activation-tagging identifies an Arabidopsis BR-deficient mutant. The activation-tagged mutant sob7-D is similar to a BR-deficient mutant (det2-1) and the sob7-DR recapitulation line. SOB7 loss-of-function line (sob7-1) is similar to the Col-0 wild type (data not shown). Plants were grown for 9 weeks in short days. Image reprinted from Turk et al. (2005) with permission from Wiley-Blackwell Publishers.

Figure 11.

The reaction catalyzed by CYP73A5.

Figure 12.

In vivo GUS staining in CYP73A5 promoter:GUS transformants. Image reprinted from Bell-Lelong et al. (1997). (A) Ten-day-old seedlings; (B) 10-day-old seedling root; (C) mature leaf; (D) rachis transverse section; (E) flower; (F) mature leaf stained 48h after wounding; (G) mature leaf stained immediately after wounding. A, C, E, F, G, Bar = 500 mm. B, D, Bar= 10 mm. The CYP73A5 promoter was successfully used to drive the expression of the coniferaldehyde 5-hydroxylase (CYP84A1) gene in the vascular tissues from Arabidopsis and tobacco (Franke et al., 2000; Meyer et al., 1998).

Figure 13.

Developmental abnormalities in the ref3 mutants. Plant with strong ref-1 and ref-2 alleles are dwarfed and show changes in apical dominance. Image reprinted from Schillmiller et al. (2009) with permission from Wiley-Blackwell Publishing.

Figure 14.

Reactions catalyzed by CYP74s. CYP74A is an allene oxide synthase (AOS) and dehydrates 13-hydroperoxides of both linoleic and linolenic acids into unstable C18 allene oxides. CYP74B2 catalyzes the cleavage of 13S-hydroperoxy-9(Z), 11(E), 15(Z)-octadecatrienoic acid (linolenic acid 13-hydroperoxide) 10 times more efficiently than that of 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid (linoleic acid 13-hydroperoxide).

Figure 15.

Developmental control of CYP74A. Arabidopsis thaliana (A-I, K) or Nicotiana tabacum (J) plants grown under sterile conditions and harboring the translational CYP74A promoter:uidA fusion. (A) Overview of a 3-week old plant. (B) Leaf base and stipule of a bract of the inflorescence axis. (C) Detached leaves of the plant shown in A arranged in the order of development (from left to right: cotyledons to younger leaves), necrotic areas marked by arrows. (D)(E) Developing flower buds. (F) Flower at the stage of fertilization. (G) Close-up of anther. (H)(I) Abscission zone scars after shedding of flower organs. (J) Pollen grains of transgenic tobacco. (K) Close-up of the bases of floral organs. Image reprinted from Kubigsteltig et al. (1999) with permission from Springer.

Figure 16.

Ribbon diagram of CYP74A1 crystal structure. N-terminus is in blue and C-terminus in red. Hydrophobic tails of two detergent molecules interacting with the membrane-associated nonpolar surface are represented in magenta. Image reprinted from Lee et al. (2008) with permission from Nature Publishing Group.

Figure 17.

CYP75B1 is a flavonoid 3′-hydroxylase. CYP75B1 catalyzes the 3′-hydroxylation of the ring B of naringenin and dihydrokaempferol to form eriodictyol and dihydroquercetin, respectively.

Figure 18.

Proanthocyanidin deposition in the seed coat of the tt7-3 mutant. Reprinted from Abraham et al. (2002). (A) The tt7-3 mutants defective for CYP75B1 show a characteristic spotted pattern deposition of proanthocyanidin upon staining with p-dimethylaminocinnamaldehyde. (B) Microscopic section of developing seeds at the heart stage showing small round inclusions of proanthocyanidin in the vacuole.

Figure 19.

Conversion of linoleic acid into the corresponding 12, 13-epoxy fatty acid. The epoxy fatty acid product was formed in vitro by incubation of linoleic acid with NADPH and microsomes of yeast expressing recombinant CYP77A4.

Figure 20.

Abaxial epidermal cells of petals in WT and ccyp77a6-1 knockout mutants as seen by scanning electron microscopy. Surface nanoridges are absent in the mutant. Left: wild-type; right: cyp77a6-1 mutant. Abaxial sides of petals are shown. The same ridgeless phenotype was observed on adaxial sides, in sepals and in the cyp77a6-2 mutant. Scale bars: 10 pm. Reprinted from Li-Beisson et al. (2009) with permission from National Academy of Sciences.

Figure 21.

Phenotype of inflorescences in lines under- or over-expressing CYP78A5. (A-E) Inflorescences of (A) Ler control, (B) klu-2, (C) the KLU-overexpressing line klu-2, RLox, (D) Col-0 wild-type, and (E) klu-4. Scale bars are 8 mm. Reprinted from Anastasiou et al. (2007) with permission from Elsevier.

Figure 22.

Phenotype of cyp78a5 and cyp78a7 insertional mutants. (A) T-DNA insertion sites in CYP78A5 and CYP78A7. (B) Rosettes of 30-d-old plants grown in short days. (C) Number of leaves in short-day-grown plants. (D) and (E) Wild-type embryos at bent-cotyledon stage (D) and at maturity (E). (F) and (G) cyp78a5 cyp78a7 embryos at bent-cotyledon stage (F) and at maturity (G). Reprinted from Wang et al. (2008).

Figure 23.

Expression of CYP78A5 compared to cell activity regions in developing petals. pKLU/GUS expression pattern (E to E″) does not occurr in regions of cell division activity (F to F″), which is monitored by the pAtCycB1;1/CDBGUS reporter. Petals in (E) and (F), in (E′) and (F′), and in (E″) and (F″) are from flowers in comparable developmental stages. Scale bar 100 pm. Reprinted from Anastasiou et al. (2007) with permission from Elsevier.

Figure 24.

Phenotype resulting from the over-expression of CYP78A9. Reprinted from lto and Meyerowitz (2000). (A) and (B) Wild-type (left) and mutant flowers (right). The length of the sepals and petals of mutant flowers did not differ from wild-type. However stamens of mutants were approx. 50% shorter than those of the wild type. The single mutant pistils were longer and wider than those of the wild type. (C) Wild type siliques 14 days after emasculation (left), mutant siliques 3 days (middle), and 5 days (right) after anthesis. The siliques of mutants continued to elongate without fertilization. Stigmatic papillae of elongating siliques were still intact. (D) Unpollinated dried mutant silique. The silique of the mutant showed a parthenocarpic phenotype. One carpel was removed to view the inside (right). No seeds were produced. (E) SEM of mutant pistil. Part of one carpel was removed to view the ovules inside. (F) to (H) Close-up view of the ovules. Most of the ovules were shriveled around the region where the embryo sac would be in the wild type (G). However, a few ovules showed normal morphology (H). (I) Self-pollinated wild-type silique (left) and pollinated mutant silique with wild type pollen (middle). Pollinated silique elongated to as much as 18 mm in one extreme case (right).

Figure 25.

Role of CYP79s and CYP83s in the glucosinolate pathway. CYP79 convert a range of aromatic or aliphatic amino acid precursors into the corresponding acetaldoximes. CYP83 catalyze the formation of aci-nitro or nitrile oxide intermediates, which in the presence of thiol compounds (R-SH) form S-alkyl-thiohydroximate adducts. In the absence of a thiol compound, the highly electrophilic product of CYP83 catalysis inactivates the enzyme.

Figure 26.

The bus1-1 phenotype induced by En-1 insertion in CYP79F1. Reprinted from Reintanz et al. (2001). The mature bus1-1 mutant exhibits a small bushy phenotype. bus1-1 is shown on the left, wild type on the right. (B) Under normal greenhouse conditions, the bus1-1 mutant has smaller rosette leaves (3 weeks old).

Figure 27.

CYP81F2 catalyzes the conversion of the phytoanticipin indole-3yl-methyl glucosinolate (I3M) into 4-hydroxy-indole-3-yl-methyl glucosinolate (4OH-I3M) in the indole glucosinolate pathway to activate mechanisms of defense against pathogens and insects.

Figure 28.

CYP82C2 promoter-driven GUS expression in A. thaliana. (A) in 6-day-old seedlings; (B) in root vascular tissues of 6-day-old seedlings; (C) in leaf of 14-day-old seedlings; (D) in anthers of open flowers; (E) in silique. Reprinted from Liu et al. (2010) with permission from Nature Publishing Group.

Figure 29.

C-C cleavage reactions for the formation of homoterpenoids from acyclic diterpenoid and sesquiterpenoid substrates by Arabidopsis CYP82G1.

Figure 30.

The rnt1-1 phenotype induced by knock-out of CYP83B1. Reprinted from Bak and Feyereisen (2001). (A) 1 week old rnt1-1 seedlings have increased hypocotyl length and epinastic cotyledons. (B) After two weeks, exfoliation of the hypocotyl begins at the root-hypocotyl junction (-,). Secondary roots initiate from the hypocotyl, and there is an enhanced formation of secondary roots and root hairs. (C) 6 weeks old rnt1-1 plants have increased apical dominance due to elevated IAA levels. Typically, the plants have a reduced height, an increased number of epinastic rosette leaves and a single inflorescence.

Figure 31.

The sin1 (fah1) mutant phenotype. Reprinted from Chappie et al. (1992). Impact of the mutation on the staining of lignin with the Mäule reagent which produces a red coloration in the presence of syringyl units. Upper panel: wild type. Lower panel: sin1 mutant. The position of the pith (p), sclerified parenchyma (s), xylem (x), and epidermis (e) are indicated. Wild-type and sin1 mutant photographed under UV light. Wild-type (right) and sin1 mutant (left) plants were photographed using a 365 nm transillu-minator as a light source and a pale yellow barrier to remove reflected UV light. The green color of the wild-type plants is due to the fluorescence of sinapoyl malate in the leaves' upper epidermis and appears green rather than blue because of the filter employed during photography. The red color of the mutant is due to the UV-induced chlorophyll fluorescence that is revealed in the absence of sinapoyl malate.

Figure 32.

The reactions catalyzed by CYP84A1.

Figure 33.

Simplified route of brassinosteroid biosynthesis and metabolism Location of CYPs on BR synthesis and metabolism pathway, major intermediates are shown. Location of CYP90A1, CYP724A1 and CYP72C1 need to be clarified.

Figure 34.

Knock-out phenotypes of Arabidopsis CYP85A1 and CYP85A2. (A) 24-day-old seedlings of cyp85a1, cyp85a2 and WT. (B) 31-day-old single and double mutant of cyp85a1 and cyp85a2. (C) The effects of BL and CS on the cyp85a1/cyp85a2 double mutant and the cpd mutant. Ten-day-old seedlings were treated on a medium containing either CS (striped bar) or BL (black bar). After 4 days of treatment, the diameters of the rosettes were measured. (D) The cyp85a1-21 cyp85a2-2 mutant treated on a medium containing 100 nM CS or 100 nM BL for 4 days and WT seedlings grown on the medium without BRs. Reprinted from Nomura et al. (2005) with permission from the American Society for Biochemistry and Molecular Biology.

Figure 35.

Features of the phenotype of lcr mutants. (A) Inflorescence of an lcr plant exhibiting organ fusion. The lcr mutant is generally fertile, although partially filled siliques are sometimes observed. (B) An example of the strong leaf fusion that caused tearing of tissues that were not directly implicated in the fusion during growth of the lcr plant. The arrow shows the broken petiole; note that the detached leaf blade did not senesce. Reprinted from Wellesen et al. (2001) with permission from the National Academy of Sciences.

Figure 36.

The null cyp86a2-1 mutant displays altered cuticle membrane ultrastructure. Ultrastructure of the wild-type Col-gl (A) and the cyp86a2-1 mutant (B) cuticle membrane from an epidermal cell. The cuticle (arrowhead) and cell wall (CW) are shown. Note the lower electron density of the cyp86a2-1 cuticle membrane, which was found in all samples examined, compared with the dense, compact cuticle in the wild-type plant. Reprinted from Xia et al. (2004) with permission from the Nature Publishing Group.

Figure 37.

Conversions catalyzed by CYP88 and CYP701 P450s in gibberellin biosynthesis. CYP701A1 the ent-kaurene oxidase (AtKO), catalyzes three successive oxidations at C-19. This is followed by CYP88A3/CYP88A4 ent-kaurenoic acid oxidase (AtKAO) that carryout three successive oxidations on C-7.

Figure 38.

Feedback regulation of CYP90A1 promoter. Developmental activation and brassinolide-mediated inhibition of the CPD promoter during early seedling development. GUS histochemical staining of transgenic seedlings carrying the CPD promoter-uidA reporter construct germinated and grown in the dark (a) and in the light (b). The lower panel in (b) shows seedlings grown in the presence of 1 µM brassinolide. Reprinted from Mathur et al. (1998) with permission from Wiley-Blackwell Publishing.

Figure 39.

Proposed action of CYP96A15/MAH1 in the decarbonylation pathway of wax biosynthesis. Reprinted from Greer et al. (2007). Only the reactions modifying 30:0 acyl CoA are shown.

Figure 40.

Expression of CYP96A15/MAH1 (Greer et al., 2007). MAH1:GFP fusion protein was expressed in the mah1-1 mutant background under the control of the native MAH1 promoter (A, B, C, E) or the 35S promoter (D, F). (A)(D) stems, (B) petioles, (C) sepals, (E) and (F) leaves. Autofluorescence is depicted as red and images are layered signals (autofluorescence and GFP).

Figure 41.

Schematic carotenoid biosynthesis pathways: the alpha- and beta-carotenoid routes to the major metabolites lutein and neoxanthin. Activity of P450 enzymes (CYP97A3, CYP97B3, CYP97C1), deduced from mutant phenotype analysis, heterologously expressed enzymes or biochemical analysis of over-expression lines are indicated at the pathway step (¹Tian et al. (2004); ²Fiore at al. (2006); ³Kim and DellaPenna (2006); ⁴Kim et al. (2010)). ^anon-heme β-carotene-ring hydroxylase.

Figure 42.

Immunolocalization of the expression of CYP98A3 in stems and roots. Hand-cut transversal sections of inflorescence stems and roots were stained with phloroglucinol HCl, a red coloration reflecting lignin content. Adjacent sections were printed onto nitrocellulose and revealed using anti-CYP98A3 polyclonal antibodies. Blue staining is indicative of CYP98A3 expression. In stems, prints were taken at increasing distances from the apical meristem to monitor temporal and developmental expression of CYP98A3 in conjunction with the differentiation of lignified tissues. No blue staining was obtained with preimmune antibodies. (A,C,E), and (G), lignin staining with phloroglucinol; (B,D,F), and (H), immunostaining of CYP98A3. (A) and (B), upper segment of the stem, close to the flower bud; (C) and (D), mid-stem; (E) and (F), lower, well differentiated stem close to the rosette; (G) and (H), root, ep, epidermis; c, cortex; px, protoxylem; mx, metaxylem; ph, phloem; if, interfascicular region; sx, secondary xylem; vc, vascular cambium; sph, secondary phloem; pd, periderm. Reprinted from Schoch et al. (2001) with permission from American Society for Biochemistry and Molecular Biology.

Figure 43.

The reactions catalyzed by CYP98A3. 5-O-(4-Coumaryol)-shikimate is metabolized 4 times more efficiently than 5-O-(4-coumaryol)-D-quinate and is probably the favored substrate In vivo.

Figure 44.

Phenotype of the ref8 mutant. UV and size phenotypes of wild type and ref8, a CYP98A3 null mutant. Rosette leaves of 3-week-old plants were photographed under 365 nm UV light using a yellow barrier filter. The blue-green color of the wild-type rosettes is due to the fluorescence of sinapoylmalate. The red fluorescence of the ref8 plant is due to chlorophyll fluorescence that is revealed in the absence of sinapate ester fluorescence. Reprinted from Franke et al. (2002a) with permission from Wiley-Blackwell Publishing.

Figure 45.

Phenotypes associated with CYP98 co-suppression. Arabidopsis Col-0 plants were transformed with a CaMV 35S:CYP98A3 construct (Abdulrazzak et al., 2006). Approximately 10% of primary transformants showed cosuppression of CYP98A3. The phenotypes of selected transformants are shown in A to E (roman numbers identify individual transformants). (A) Three-weeks old T1 plants grown on vertical agar plates. Wild-type (Col-0) plants are shown on both sides. (B) to (E), Different 10-week-old cosuppressed T1 lines grown on soil (B). Moderately cosuppressed plants arrested at different stages of bolting stem development (C–E). When plants bolted, inflorescences showed purple stems and cauline leaves; they were limp, in most cases male sterile, and rarely produced viable seeds (C) and (E).

Figure 46.

Phylogeny of the CYP98 proteins

Figure 47.

The reactions catalyzed by CYP98A8 and CYP98A9 in the phenolamide pathway and formation of major pollen coat constituents. SHT : spermidine hydroxycinnamoyl transferase AtSM1 : cation-dependent O-methyltransferase-like protein CCoAMT1 : caffeoyl CoA O-methyltransferase 1

Figure 48.

Evolution of CYP98A8 and CYP98A9.

Figure 49.

The ga3 mutant phenotype and complementation of the mutation with CYP701A3. (I–III) Three kanamycin resistant plants from the transformation of ga3-2 with a genomic CYP701A3 clone. (ttg ga3-2): The mutant ga3-2 line. (L.er): The wild-type Landsberg erecta line. Reprinted from Helliwell et al. (1998) with permission from the National Academy of Sciences.

Figure 50.

Confocal images of tobacco leaf cells following microprojectile bombardment showing chloroplast localization of CYP701 A3. Images were taken 24 h after bombardment of tobacco leaves with tungsten particles coated with individual plasmid constructs. The cells expressing the GFP construct are epidermal cells with the exception of Ei-iii which show a mesophyll cell. The images are dual GFP (green) and chlorophyll (red) channels, except those labelled (i) and (iii) which show separate chlorophyll and GFP channels, respectively. The lengths of scale bars are given in µm in each panel. (A) TPCPS—GFP (copalyl diphosphate synthase transit peptide-GFP fusion: plastidial control), (B) TPKS—GFP (kaurene synthase transit peptide-GFP fusion: plastidial control), (C) RbcS—GFP, (D) smGFP, (E) TPCYP701A3—GFP, (F) CYP701A3—GFP, (G) TPCYP88A3—GFP, (H) TPCYP88A4—GFP, (I) mGFP5, (J) 20ox2—GFP Images reprinted from Heliwell et al. (2001) with permission from Wiley-Blackwell.

Figure 51.

Comparison of pollen from CYP703A2 knockout plants and wild-type pollen. (A) to (D) Surface structure of Arabidopsis pollen from wild-type Col-0 plants (A) and (B) and the CYP703A2 SLAT N56842 knockout line (C) and (D) after fixation of the tissues (A) and (C) or without fixation (B) and (D), monitored by scanning electron microscopy. Images modified from Morant et al. (2007).

Figure 52.

The reactions catalyzed by yeast-expressed CYP703A2. Yeast-expressed CYP703A2 catalyzes a single hydroxylation of medium chain fatty acids, mainly C12:0 and C10:0, in the chain with C7 as the main but not exclusive position of hydroxylation. Hydroxylated products obtained decrease with the distance from C7 as indicated by the size of the arrows on the top of the chain that roughly reflect reaction efficiency. A more specific reaction might occur in vivo.

Figure 53.

zebra mutants have severe defects in exine structure. Scanning electron micrographs of the surface structure of pollen grains from the wild type (A) and the cyp704b1 mutant (allele SAIL_1149_B03; B). Samples were prepared without fixation. Reprinted from Dobritsa et al., (2009).

Figure 54.

Phylogenetic tree of the Arabidopsis CYP705 family. The remotely related subfamily CYP712 was used as outgroup and the broken line indicates greater distance than shown. Suffix P is used for pseudogenes. The number in brackets indicates location in one of five gene clusters: At1g50520 At1g50560 (1), At2g27000 At2g27010 (2), At3g20080 At3g20083 At3g20090 At3g20100 At3g20110 At3g20120 At3g20130 At3g20140 (3a), At3g20940 At3g20950 At3g20960 AT3G20935 (3b) and At4g15330 At4g15350 At4g15360 At4g15380 (4). The chevron highlights genes that were likely re-located after, or throughout duplication. The scale bar represents 0.1 amino acid changes. Highlighted the thalianol dehydrogenase. [Method: Maximum likelihood phylogeny using PhyML on manally curated alignment (Dialign2) of the deduced protein sequences (www.p450.kvl.dk), JTT Model of amino acids substitution, discrete gamma model with 4 rate categories (gamma shape parameter : 1.434), 100 bootstrap repetitions. Asterisks indicate bootstrap support of 70% and greater.]

Figure 55.

The thalianol biosynthetic gene cluster. (A) The position of the cluster on chromosome 5 is given as yellow bar and the distribution of the genes encoding the thalianol cluster is shown in the second panel. (B) Scheme of the pathway to modified thalianol derivatives. Position of the introduced hydroxyl function is unclear, and several compounds detected in Arabidopsis possibly represent positional isomers. Drawn after Field and Osbourn (2008).

Figure 56.

Hydroxylation of ABA catalyzed by the CYP707A enzymes. CYP707A catalyses the 8′ hydroxylation of ABA.

Figure 57.

Spatial expression patterns of CYP707A1 and CYP707A3 in response to high humidity. Expression of CYP707A1 promoter:GUS before (A) and after transferring to high humidity conditions (B–D). Expression of promoter CYP707A3:GUS before (E) and after transferring to high humidity conditions (F–H). Approximately 3-week-old plants were grown on soil in pots under RH 60% at 22°C condition. High humidity treatment (from RH 60% to 90% at 22°C) was performed for 2 h. Reprinted from Okamoto et al. (2009).

Figure 58.

Common backbone structure of non-hormonal bulk sterol lipids and the desaturation reaction catalysed by Arabidopsis CYP710A. Functional evidence through in vitro enzyme assay ¹Morikawa et al., (2006); over-expression in planta ²Morikawa et al., (2006) and Arnqvist et al., (2008); null mutant allele ³Morikawa et al., (2006) and Griebel and Zeier, (2010).

Figure 59.

The possible role(s) of CYP711 in the strigolacone pathway. (A) Common backbone structure (in red) and various oxidative decorations leading to naturally occurring strigolactones. (B) Hypothesized biosynthetic pathway leading to strigolactones based hormones and signaling molecules. Asterisks indicate strigolactones detected in Arabidopsis. Shown in grey are functions not found in Arabidopsis.

Figure 60.

CYP735A members catalyze hydroxylation of cytokinins. CYP735A1 and CYP735A2 can hydroxylate the side chain of cytokinins in the synthesis of trans-zeatin. They can convert the following mono-, di- and tri- phosphates forms into their respective hydroxylated products; iPRMP, isopentenyladenine riboside 5′-monophosphate to tZRMP, trans-zeatin riboside 5′-monophosphate; iPRDP, isopentenyladenine riboside 5′-diphosphate to tZRDP, trans-zeatin riboside 5′-diphosphate; iPRTP, isopentenyladenine riboside 5′-triphosphate to tZRTP, trans-zeatin riboside 5′-triphosphate.