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Biochemical and Molecular Mechanisms
Underlying Colour Retention in
Capsicum annuum.
Alexandra Holden1, Harriet Berry1, Daniel Rickett2, Paul Fraser1.
1Department of Biological Sciences, Royal Holloway, University of London, Egham, Surrey, UK | email: [email protected]
2Syngenta, Jealott’s Hill International Research Centre, Bracknell, Berkshire, RG42 6EY.
Background
Capsicum annuum
• Chilli pepper (Capsicum annuum) is the most widely grown spice
product.
• Colour retention is a key quality trait.
• Colour is lost over time during storage, leading to less valuable crop.
Carotenoids
• Carotenoids are coloured lipophilic tetraterpenes, synthesised by
photosynthetic plants (Fig. 1).
• Carotenoid quantity and composition, and sub-chromoplast
sequestration mechanisms, are directly linked to red colour
phenotype.
• Capsanthin (Fig. 2), and its esters, are primarily responsible for red
colour of C. annuum.
Objective 1: Metabolic profiling of a C.
annuum population
x
Spec
Assay
HPLC
Metabolic profiling of a C. annuum
population displaying variation in colour
retention phenotype will identify differences
in carotenoid profile, along with other
metabolites, in high and low retention lines.
GC-MS
This data will then be used in
QTL analysis for locating
genomic regions containing
candidate genes responsible
for carotenoid degradation.
Fig 3. Carotenoid profile
HPLC chromatograms.
Fig 1. Carotenoid
modification pathway,
from phytoene1.
Fig 2. Capsanthin structure.
Objective 2: Identification of candidate
genes involved in carotenoid degradation
Using QTL data, loci in the C. annuum genome containing candidate genes
involved in colour loss mechanisms will be identified. Candidate genes
will be identified through bioinformatic data mining using the Capsicum
genome sequence.
We hypothesise that candidate genes may
include carotenoid cleavage dioxygenases, or
genes involved in carotenoid sequestration and
storage. Gene expression techniques, such as
RNA-seq and qPCR, will determine the gene
expression profile in high retention lines.
Protein expression will determine function.
Objective 3: Subchromoplast structural
study
Objective 4: Characterisation of lipid
composition and peroxidation
Carotenoids are sequestered in organelles,
known as chromoplasts; plastoglobules, within
chromoplasts, are lipoproteins associated with
carotenoids2.
Plastoglobule structure is
dependent on the carotenoids present. Fibrillin
is a protein commonly found associated with
carotenoids in plastoglobules in C. annuum3.
Lipid peroxidation is thought to cause carotenoid degradation. Carotenoids
protect lipids from oxidation by scavenging ROS5. The antioxidative role of
carotenoids is a mechanism for carotenoid degradation.
Lipid peroxidation rate will be assessed in different lines to determine
whether a correlation exists.
Variation in lipid composition in sub-
Fig 4. Chromoplast4.
Understanding of the storage mechanisms will highlight the role that such
structures play in carotenoid degradation, and hence, colour loss. Electron
microscopy will be used to determine sub-chromoplast structure, and
enzyme assays will determine enzyme localisation.
Fig 5. Waxy cuticle structure.
chromoplast fractions of different
lines will be characterised by GC-MS.
Fruit waxy cuticle structure will be
observed by electron microscopy to
determine
whether
cuticle
ultrastructure correlates with colour
retention.
Expected outcomes and impact
Understanding colour loss and carotenoid degradation mechanisms in C. annuum will:
•
•
•
•
Result in the breeding of high colour retention chilli pepper varieties leading to greater economic return for growers
Increase understanding of colour loss mechanisms in a wide array of species
Allow the nutritional benefit of chilli peppers to be exploited, as carotenoids may be degraded slower
Provide a basis on which genome editing may be carried out, following identification of genes involved in colour loss
Acknowledgements and References
I would like to thank members of the Fraser Group at Royal Holloway for help and support with this project. This PhD studentship is funded by BBSRC CASE studentships in collaboration with Syngenta.
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Fraser PD, Bramley PM (2004). Progress in Lipid Research 43 (3):228-265.
Austin JR, Frost E, Vidi P-A, Kessler F, Staehelin LA (2006). The Plant Cell 18 (7):1693-1703.
Deruère J, Römer S, d'Harlingue A, Backhaus RA, Kuntz M, Camara B (1994). The Plant Cell 6 (1):119-133.
Botte CY, Marechal E (2013). Trends in Plant Science 19 (2):71-78.
Sharma P, Jha AB, Dubey RS, Pessarakli M (2012). Journal of Botany 2012:26.