15,15 -£-phytoene

IPP react with each other, releasiag pyrophosphate to form another allyl pyrophosphate containing 10 carbon atoms. The chain can successively build up by five-carbon units to yield polyisoprenes by head-to-tad condensations alternatively, tad-to-tad condensations of two units can yield squalene, a precursor of sterols. Similar condensation of two C2Q units yields phytoene, a precursor of carotenoids. This information is expected to help ia the development of genetic methods to control the hydrocarbon stmctures and yields. 

A second class of herbicides primarily affects ( -carotene desaturase. These herbicides are apparent feedback inhibitors of PD as well. This class of compounds includes dihydropyrones like LS 80707 [90936-96-2] (56) and 6-methylpyridines (57,58). The third class consists of the ben2oylcyclohexane-diones, eg, 2-(4-chloro-2-nitroben2oyl)-5,5-dimethyl-cyclohexane-I,3-dione. This class of atypical bleaching herbicides induces phytoene accumulation when appHed either pre- or post-emergence. However, it does not inhibit phytoene desaturase activity in vitro (59). Amitrole also has been considered a bleaching herbicide, though its main mode of action is inhibition of amino acid synthesis. 

The desaturation of l5-cis phytoene into lycopene occurs in four stepwise dehydrogenations, yielding phytofluene, ( -carotene, neurosporene and lycopene... 

Isomerisation of 15-c/5-phytoene to the all-/ra x configuration must occur during the desaturation steps, since most desaturated carotenes are in the all-trans form. The CRTI type desaturases appear to be able to carry out this isomerisation themselves (Fraser et al, 1992 Bartley etal, 1999), but mutants of PDS/ZDS-type organisms accumulate cis isomers of unsaturated carotenes, suggesting the presence of a separate isomerase (Clough and Pattenden, 1983 Ernst and Sandmann, 1988). Three recent publications have reported the cloning of a carotene isomerase (CrtlSO) from tomato (Isaacson et al, 2002), Arabidopsis (Park et al, 2002) and Synechocystis 6803 (Breitenbach... 

Clearly, the control of gene expression at the transcriptional level is a key regulatory mechanism controlling carotenogenesis in vivo. However, post-transcriptional regulation of carotenoid biosynthesis enzymes has been found in chromoplasts of the daffodil. The enzymes phytoene synthase (PSY) and phytoene desaturase (PDS) are inactive in the soluble fraction of the plastid, but are active when membrane-bound (Al-Babili et al, 1996 Schledz et al, 1996). The presence of inactive proteins indicates that a post-translational regulation mechanism is present and is linked to the redox state of the membrane-bound electron acceptors. In addition, substrate specificity of the P- and e-lycopene cyclases may control the proportions of the p, P and P, e carotenoids in plants (Cunningham et al, 1996). 

The carotenoid pathway may also be regulated by feedback inhibition from the end products. Inhibition of lycopene cyclisation in leaves of tomato causes increase in the expression of Pds and Psy-1 (Giuliano et al, 1993 Corona et al, 1996). This hypothesis is supported by other studies using carotenoid biosynthesis inhibitors where treated photosynthetic tissues accumulated higher concentrations of carotenoids than untreated tissues (reviewed by Bramley, 1993). The mechanism of this regulation is unknown. A contrary view, however, comes from studies on the phytoene-accumulating immutans mutant of Arabidopsis, where there is no feedback inhibition of phytoene desaturase gene expression (Wetzel and Rodermel, 1998). 

P = phytoene PF = phytofluene -C = -carotene L = lycopene y-C = y-carotene p-C = p-carotene Lut = lutein Viola = violaxanthin Neo = neoxanthin d.p.b. days post breaker. 

Rice Psy cDNA from daffodil CaMV 35S Phytoene accumulation in endosperm Burkhardt et al., 1997... 

A further consideration in the choice of gene relates to those that are members of a gene family. Only one member of such a family may be involved in carotenogenesis in a particular tissue. A good example of this is Psy-1 and-2 of tomato. PSY-1 is responsible for phytoene synthesis in ripening fruit (Fraser et al, 1999) whereas PSY-2 is not functional in chromoplasts, even if the protein is produced. 

AL-BABiLi s, VON LiNTiG J, HAUBRUCK H and BEYER p (1996) A novel, soluble form of phytoene desaturase from Narcissus pseudonarcissus chromoplasts is Hsp70-complexed and competent for flavinylation, membrane association and enzymatic activation . Plant J,9, 601-12. 

POTRYKUS I (1997) Transgenic rice Oryza sativa) endosperm expressing daffodil Narcissuspseudonarcissus) phytoene synthase accumulates phytoene, a key intermediate of provitamin A biosynthesis , Plant J, 11, 1071-78. 

FRASER P D, MISAWA N, LINDEN H, SHIGEYUKI Y, KOBAYASHI K and SANDMANN G (1992) Expression mE. coli, purification and reactivation of a recombinant Erwinia uredovora phytoene desaturase ,Chem, 267, 19891-5. 

FRASER P D, KIANO I W, TRUESDALE M R, SCHUCH W and BRAMLEY P M (1999) PhytOene synthase-2 enzymes activity in tomato does not contribute to carotenoid synthesis in ripening fruit . Plant Mol Biol, 40, 687-98. 

FRASER P D, ROMER S, SHIPTON C A, MILLS P B, KIANO I W, MISAWA N, DRAKE R G, SCHUCH W and BRAMLEY p M (2002) Evaluation of transgenic tomato plants expressing an additional phytoene synthase in a fruit-specific manner , Proc Natl Acad Sci, 99, 1092-7. 

FRAY R, WALLACE A, FRASER P D, VALERO D, HEDDEN P, BRAMLEY P M and GRIERSON D (1995) Constitutive expression of a fruit phytoene synthase gene in transgenic tomatoes causes dwarfism by redirecting metabolites from the gibberellin pathway . Plant J, 8, 696-701. 

Expression of a tomato cDNA coding for phytoene synthase in Escherichia coli, ph)doene formation in vivo and in vitro, and functional analysis of the varions trimcated gene prodncts , J Biochem (Tokyo), 116, 980-85. 

PECKER I, CHAMOVITZ D, LINDEN H, SANDMANN G and HIRSCHBERG Y (1992) A Single polypeptide catalysing the conversion of phytoene to -carotene is transcriptionally regulated during tomato fruit ripening , Proc Natl Acad Sci, 89, 4962-6. 

SCHLEDZ M, AL-BABILI S, VON LINTIG J, HAUBRUCK H, RABBANI S, KLEINIG H and BEYER P (1996) Phytoene synthase fcom Narcissus pseudonarcissus functional expression, galactolipid requirement, topological distribution in chromoplasts and induction during flowering . Plant J, 10, 781-92. 

WETZEL c M and RODERMEL s R (1998) Regulation of phytoene desaturase expression is independent of leaf pigment content in Arabidopsis thaliana , Plant Mol Biol, 37, 1045-53. 

Shewmaker, C.K. et al.. Seed-specific overexpression of phytoene synthase increase in carotenoids and other metabolic effects. Plant J., 20, 401, 1999. 

The first C40 carotenoid, phytoene, is produced by head-to-head condensation of two GGPPs by an enzyme that shares homology to GGPPS and squalene synthases... 

Besides the capacity of CRTI to introduce all four double bonds in the conversion of phytoene to lycopene, the enzyme produces different geometric isomers than does PDS/ZDS (see graphic, side-by-side comparison in Fraser and Bramley ). CRTI produces all-trans isomers. Studies that have examined the function of the paired plant desaturases acting together, from Arabidopsis, and from maize and from... 

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