Plant Molecular Biology 50: 129–142, 2002.
© 2002 Kluwer Academic Publishers. Printed in the Netherlands.
Expression of a bacterial carotene hydroxylase gene (crtZ) enhances UV
tolerance in tobacco
, Gerhard Sandmann
and Susanne Römer
Lehrstuhl für Physiologie und Biochemie der Pﬂanzen, Universität Konstanz, P.O. Box 5560, 78434 Konstanz,
author for correspondence; e-mail Susanne.Roemer@uni-konstanz.de);
Botanisches Institut, J.W.
Goethe Universität, P.O. Box 111932, 60054 Frankfurt, Germany
Received 8 May 2001; accepted in revised form 9 January 2002
Key words: β-carotene hydroxylase, carotenoids, lipid peroxidation, Nicotiana tabacum L., transgenic plants, UV
tolerance, xanthophyll cycle
Carotenoids are essential components of the photosynthetic apparatus involved in plant photoprotection. To in-
vestigate the protective role of zeaxanthin under high light and UV stress we have increased the capacity for
its biosynthesis in tobacco plants (Nicotiana tabacum L. cv. Samsun) by transformation with a heterologous
carotenoid gene encoding β-carotene hydroxylase (crtZ) from Erwinia uredovora under constitutive promoter
control. This enzyme is responsible for the conversion of β-carotene into zeaxanthin. Although the total pigment
content of the transgenics was similar to control plants, the transformants synthesized zeaxanthin more rapidly
and in larger quantities than controls upon transfer to high-intensity white light. Low-light-adapted tobacco plants
were shown to be susceptible to UV exposure and therefore chosen for comparative analysis of wild-type and
transgenics. Overall effects of UV irradiation were studied by measuring bioproductivity and pigment content. The
UV exposed transformed plants maintained a higher biomass and a greater amount of photosynthetic pigments than
controls. For revelation of direct effects, photosynthesis, pigment composition and chlorophyll ﬂuorescence were
examined immediately after UV treatment. Low-light-adapted plants of the crtZ transgenics showed less reduction
in photosynthetic oxygen evolution and had higher chlorophyll ﬂuorescence levels in comparison to control plants.
After 1 h of high-light pre-illumination and subsequent UV exposure a greater amount of xanthophyll cycle pig-
ments was retained in the transformants. In addition, the transgenic plants suffered less lipid peroxidation than the
wild-type after treatment with the singlet-oxygen generator rose bengal. Our results indicate that an enhancement
of zeaxanthin formation in the presence of a functional xanthophyll cycle contributes to UV stress protection and
prevention of UV damage.
Abbreviations: bp, base pair; DTT, dithiothreitol; DW, dry weight; FW, fresh weight; PCR, polymerase chain
reaction; PMSF, phenylmethylsulﬂuoride; RNA, ribonucleic acid; SD, standard deviation; SDS, sodium dodecyl
sulfate; UV, ultraviolet
Carotenoids are naturally occurring pigments with
antioxidative properties (Krinsky, 1989) which are in-
volved in photoprotection, light harvesting and mem-
brane stabilization (Havaux, 1998; Niyogi, 1999).
Photoprotection by carotenoids includes dissipation
of excess light energy (especially the de-excitation
Chl) and quenching of free radicals.
Chl leads to the formation of highly destructive
singlet-oxygen species which can also be de-excited
by carotenoids. In fact, carotenoids have been found
to be some of the most competent scavengers of
surpassing the capacity of other antioxidants such as
α-tocopherol, ascorbate and glutathione (Di Mascio
et al., 1989; Devasagayam et al., 1992). The im-