Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis

Li Tian, Maria Magallanes-Lundback, Valeria Musetti, Dean DellaPenna

Research output: Contribution to journalArticle

87 Citations (Scopus)

Abstract

Lutein and zeaxanthin are dihydroxy xanthophylls that are produced from their corresponding carotene precursors by the action of β- and ε-ring carotenoid hydroxylases. Two genes that encode β-ring hydroxylases (β-hydroxylases 1 and 2) have been identified in the Arabidopsis genome and are highly active toward β-rings but only weakly active toward ε-rings. A third distinct activity required for ε-ring hydroxylation has been defined by mutation of the LUTEIN1 (LUT1) locus, but LUT1 has not yet been cloned. To address the individual and overlapping functions of the three Arabidopsis carotenoid hydroxylase activities in vivo, T-DNA knockout mutants corresponding to β-hydroxylases 1 and 2 (b1 and b2, respectively) were isolated and all possible hydroxylase mutant combinations were generated. β-Hydroxylase single mutants do not exhibit obvious growth defects and have limited impact on carotenoid composition relative to the wild type, suggesting that the encoded proteins have a significant degree of functional redundancy in vivo. Surprisingly, the b1 b2 double mutant, which lacks both known β-hydroxylase enzymes, still contains significant levels of β-carotene-derived xanthophylls, suggesting that additional β-ring hydroxylation activity exists in vivo. The phenotype of double and triple hydroxylase mutants indicates that at least a portion of this activity resides in the LUT1 gene product. Despite the severe reduction of β-carotene-derived xanthophylls (up to 90% in the lut1 b1 b2 triple mutant), the double and triple hydroxylase mutants still contain at least 50% of the wild-type amount of hydroxylated β-rings. This finding suggests that it is the presence of minimal amounts of hydroxylated β-rings, rather than minimal amounts of specific β-carotene-derived xanthophylls, that are essential for light-harvesting complex II assembly and function in vivo. The carotenoid profiles in wild-type seeds and the effect of single and multiple hydroxylase mutations are distinct from those in photosynthetic tissues, indicating that the activities of each gene product differ in the two tissues. Overall, the hydroxylase mutants provide insight into the unexpected overlapping activity of carotenoid hydroxylases in vivo.

Original languageEnglish (US)
Pages (from-to)1320-1332
Number of pages13
JournalPlant Cell
Volume15
Issue number6
StatePublished - Jun 1 2003
Externally publishedYes

Fingerprint

Functional analysis
Carotenoids
Mixed Function Oxygenases
Arabidopsis
carotenoids
xanthophylls
mutants
carotenes
Xanthophylls
hydroxylation
Genes
Hydroxylation
mutation
knockout mutants
genes
light harvesting complex
zeaxanthin
lutein
Tissue
Lutein

ASJC Scopus subject areas

  • Plant Science
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Cell Biology

Cite this

Tian, L., Magallanes-Lundback, M., Musetti, V., & DellaPenna, D. (2003). Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis. Plant Cell, 15(6), 1320-1332.

Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis. / Tian, Li; Magallanes-Lundback, Maria; Musetti, Valeria; DellaPenna, Dean.

In: Plant Cell, Vol. 15, No. 6, 01.06.2003, p. 1320-1332.

Research output: Contribution to journalArticle

Tian, L, Magallanes-Lundback, M, Musetti, V & DellaPenna, D 2003, 'Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis', Plant Cell, vol. 15, no. 6, pp. 1320-1332.
Tian L, Magallanes-Lundback M, Musetti V, DellaPenna D. Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis. Plant Cell. 2003 Jun 1;15(6):1320-1332.
Tian, Li ; Magallanes-Lundback, Maria ; Musetti, Valeria ; DellaPenna, Dean. / Functional analysis of β- and ε-ring carotenoid hydroxylases in arabidopsis. In: Plant Cell. 2003 ; Vol. 15, No. 6. pp. 1320-1332.
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abstract = "Lutein and zeaxanthin are dihydroxy xanthophylls that are produced from their corresponding carotene precursors by the action of β- and ε-ring carotenoid hydroxylases. Two genes that encode β-ring hydroxylases (β-hydroxylases 1 and 2) have been identified in the Arabidopsis genome and are highly active toward β-rings but only weakly active toward ε-rings. A third distinct activity required for ε-ring hydroxylation has been defined by mutation of the LUTEIN1 (LUT1) locus, but LUT1 has not yet been cloned. To address the individual and overlapping functions of the three Arabidopsis carotenoid hydroxylase activities in vivo, T-DNA knockout mutants corresponding to β-hydroxylases 1 and 2 (b1 and b2, respectively) were isolated and all possible hydroxylase mutant combinations were generated. β-Hydroxylase single mutants do not exhibit obvious growth defects and have limited impact on carotenoid composition relative to the wild type, suggesting that the encoded proteins have a significant degree of functional redundancy in vivo. Surprisingly, the b1 b2 double mutant, which lacks both known β-hydroxylase enzymes, still contains significant levels of β-carotene-derived xanthophylls, suggesting that additional β-ring hydroxylation activity exists in vivo. The phenotype of double and triple hydroxylase mutants indicates that at least a portion of this activity resides in the LUT1 gene product. Despite the severe reduction of β-carotene-derived xanthophylls (up to 90{\%} in the lut1 b1 b2 triple mutant), the double and triple hydroxylase mutants still contain at least 50{\%} of the wild-type amount of hydroxylated β-rings. This finding suggests that it is the presence of minimal amounts of hydroxylated β-rings, rather than minimal amounts of specific β-carotene-derived xanthophylls, that are essential for light-harvesting complex II assembly and function in vivo. The carotenoid profiles in wild-type seeds and the effect of single and multiple hydroxylase mutations are distinct from those in photosynthetic tissues, indicating that the activities of each gene product differ in the two tissues. Overall, the hydroxylase mutants provide insight into the unexpected overlapping activity of carotenoid hydroxylases in vivo.",
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