Contents
15.1 Plant Secondary Metabolites and Production Routes ................................................. 424
15.1.1 Carotenoids...................................................................................................... 424
15.1.2 Anthocyanins................................................................................................... 425
15.2 Methods to Increase the Yields of Secondary Metabolites in Plant Cell Culture..... 425
15.2.1 Plant Cell Culture (Somaclonal Variation).................................................... 426
15.3 Genetic Transformation of Plant Cells ........................................................................ 427
15.3.1 Methods for Detecting Genetically Transformed Cells ................................ 427
15.3.2 Transfer of Multiple Genes Through Improved Vectors .............................. 428
15.4 Metabolic Engineering ................................................................................................. 429
15.4.1 Strategies for Nutrient Production in Metabolic Engineering ...................... 432
15.4.2 Steps for the Modification of Metabolic Pathways....................................... 433
15.4.3 Classification of Strategies for Modification of Metabolic Pathways.......... 434
15.5 Transcription factors Manipulation.............................................................................. 436
15.5.1 Transcription Factor Expression for the Control
of a Metabolic Pathway.................................................................................. 438
15.5.2 Design of Artificial Transcription Factors..................................................... 438
15.6 Epigenetic Strategies for Pathway Modification......................................................... 439
15.6.1 Genetically Induced Epialleles ....................................................................... 441
15.7 Plant-Based Factories of Bioactive Molecules (Alternative Models) ........................ 442
15.7.1 What to Produce and How? ........................................................................... 442
15.8 Concluding Remarks .................................................................................................... 446
References................................................................................................................................ 447
15.1 Plant Secondary Metabolites and Production Routes
15.1.1 Carotenoids
The carotenoid biosynthetic pathway (Cunningham et al. 1993) takes place within
plastids; the first committed step is the condensation of two geranylgeranyl
diphosphate (GGDP) molecules to produce phytoene, catalyzed by the enzyme
phytoene synthase (PSY). Two enzymes, phytoene desaturase (PDS) and n-carotene
desaturase (ZDS), introduce four double bonds that convert phytoene into lycopene,
the first coloring carotenoid, via phytofluene, n-carotene, and neurosporene. Then,
lycopene may undergo cyclization by two different cyclases, one branch leads to
b-carotene (b,b-carotene) and its derivative xanthophylls zeaxanthin, violaxanthin,
and neoxanthin (Cunningham et al. 1993). The alternative branch leads to carotenoids with one b and one e ring, such as a-carotene and lutein; the latter is a major
xanthophyll in the light harvesting system of higher plants (Cunningham and Gantt
2001).
Carotenoid formation is a highly regulated process; the concentration and
composition of leaf xanthophylls are affected by light intensity (Ruban et al. 1994),
and the accumulation of specific carotenoids in chromoplasts of fruits and flowers
is developmentally regulated (Giuliano et al. 1993).
424
L. A. Cruz-Ramírez et al.
15.1 Plant Secondary Metabolites and Production Routes ................................................. 424
15.1.1 Carotenoids...................................................................................................... 424
15.1.2 Anthocyanins................................................................................................... 425
15.2 Methods to Increase the Yields of Secondary Metabolites in Plant Cell Culture..... 425
15.2.1 Plant Cell Culture (Somaclonal Variation).................................................... 426
15.3 Genetic Transformation of Plant Cells ........................................................................ 427
15.3.1 Methods for Detecting Genetically Transformed Cells ................................ 427
15.3.2 Transfer of Multiple Genes Through Improved Vectors .............................. 428
15.4 Metabolic Engineering ................................................................................................. 429
15.4.1 Strategies for Nutrient Production in Metabolic Engineering ...................... 432
15.4.2 Steps for the Modification of Metabolic Pathways....................................... 433
15.4.3 Classification of Strategies for Modification of Metabolic Pathways.......... 434
15.5 Transcription factors Manipulation.............................................................................. 436
15.5.1 Transcription Factor Expression for the Control
of a Metabolic Pathway.................................................................................. 438
15.5.2 Design of Artificial Transcription Factors..................................................... 438
15.6 Epigenetic Strategies for Pathway Modification......................................................... 439
15.6.1 Genetically Induced Epialleles ....................................................................... 441
15.7 Plant-Based Factories of Bioactive Molecules (Alternative Models) ........................ 442
15.7.1 What to Produce and How? ........................................................................... 442
15.8 Concluding Remarks .................................................................................................... 446
References................................................................................................................................ 447
15.1 Plant Secondary Metabolites and Production Routes
15.1.1 Carotenoids
The carotenoid biosynthetic pathway (Cunningham et al. 1993) takes place within
plastids; the first committed step is the condensation of two geranylgeranyl
diphosphate (GGDP) molecules to produce phytoene, catalyzed by the enzyme
phytoene synthase (PSY). Two enzymes, phytoene desaturase (PDS) and n-carotene
desaturase (ZDS), introduce four double bonds that convert phytoene into lycopene,
the first coloring carotenoid, via phytofluene, n-carotene, and neurosporene. Then,
lycopene may undergo cyclization by two different cyclases, one branch leads to
b-carotene (b,b-carotene) and its derivative xanthophylls zeaxanthin, violaxanthin,
and neoxanthin (Cunningham et al. 1993). The alternative branch leads to carotenoids with one b and one e ring, such as a-carotene and lutein; the latter is a major
xanthophyll in the light harvesting system of higher plants (Cunningham and Gantt
2001).
Carotenoid formation is a highly regulated process; the concentration and
composition of leaf xanthophylls are affected by light intensity (Ruban et al. 1994),
and the accumulation of specific carotenoids in chromoplasts of fruits and flowers
is developmentally regulated (Giuliano et al. 1993).
424
L. A. Cruz-Ramírez et al.