Abstract
Endosperm carotenoid content in wheat is a primary determinant of flour colour and this affects both the nutritional value of the grain and its utility for different applications. Utilising wheat rice synteny two genes, ε-cyclase (ε-LCY) and phytoene synthase (Psy-A1), were identified as candidate genes for two of the QTL affecting lutein content in wheat endosperm. Analysis of the sequence changes in ε-LCY and Psy-A1 revealed possible causal mechanisms for both QTL. A point mutation in ε−LCY results in the substitution of a conserved amino acid in the high lutein allele. This substitution has been observed in high lutein-accumulating species from the Gentiales order. In Psy-A1, a sequence duplication at the end of exon 2 creates a new splice site and causes alternative splicing of the transcript and activation of a cryptic exon, resulting in four different transcripts: a wild-type transcript, two transcripts with early terminations and a transcript that would produce an in-frame, albeit longer protein. Only the wild-type splice variant produced an enzymatically active protein and its mRNA abundance was reduced by titration with the other splice variants. This reduction in wild-type mRNA is argued to result in a reduction in PSY protein and thus carotenoid content in wheat.
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Acknowledgements
We would like to thank Abby Cuttriss for assistance with the HPLC analysis, and Ibrahim Kutty for the flour b* measurements from the Chara × Glenlea population. The Chara × Glenlea population was milled at the Bread Research Institute, Sydney Australia. The flour b* data for the Sunco × Tasman population was generated by the Australian Winter Cereals Molecular Marker Program. pAC-Phyt and pAC-Lyc were kindly provided by F.X. Cunningham. JLM was the recipient of a CSIRO/ANU postgraduate award. BJP is grateful for support from Australian Research Council Centre of Excellence in Plant Energy Biology (CE0561495).
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Supplementary Table 1
Primers used in this study (DOC 11 kb)
Supplementary Table 2
Synteny mapping of genes involved in carotenoid biosynthesis and cleavage (DOC 35 kb)
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Howitt, C.A., Cavanagh, C.R., Bowerman, A.F. et al. Alternative splicing, activation of cryptic exons and amino acid substitutions in carotenoid biosynthetic genes are associated with lutein accumulation in wheat endosperm. Funct Integr Genomics 9, 363–376 (2009). https://doi.org/10.1007/s10142-009-0121-3
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DOI: https://doi.org/10.1007/s10142-009-0121-3