Isoprenoid Biosynthesis in Fruits and Vegetables - Postharvest Biology and Technology of Fruits, Vegetables, and Flowers - page 288

pyrophosphate (FPP) ionization by providing a proton from the hydroxyl group to assist
the separation of the pyrophosphate moiety and to stabilize the incipient carbocation with
its

π

system (Tansey and Shechter, 2000). There are several potential phosphorylation sites
directly adjacent to and within a few positions of these motifs. Finally, there are several
potential phosphorylation sites within and surrounding the SQS 50-TSRSF-54 (PSY 131-
YAKTF-135) motif, which forms a flap involved in creating a hydrophobic pocket to protect
the highly reactive carbocation intermediates from being exposed to the solvent (Pandit
et al., 2000). Thus, within the PSY Trans IPPS HH conserved domain, there a several
potential phosphorylation sites, each located within or near potentially important functional
domains.

13.7 PSY1 transgenes

Because of its role in carotenoid biosynthesis of ripening fruit, PSY1 has received much
attention. Transgenic tomato plants expressing antisense PSY1 cDNA (originally called
pTOM5 ) have pale-colored flowers and yellow fruit with a 97% reduction in carotenoid
levels (Bird et al., 1991). Understandably, this inhibition of PSY1 mRNA accumulation did
not affect leaf carotenoid levels because of the specificity of the knockout to PSY1 and
not PSY2 (Bramley et al., 1992). Constitutive PSY1 overexpression in transgenic yellow
fruit r,r-mutants (possessing a loss-of-function mutation in the PSY1 gene) restored ly-
copene synthesis to ripening fruit (Fray et al., 1995). However, these same plants exhibited
unscheduled pigment production in other cell types, and separate insertion events of the
same sense PSY1 construct caused cosuppression in some plants. In cosuppressed plants,
immature green fruit, leaves, and flowers displayed inhibited carotenoid production due to
reduced mRNA levels of both the introduced transgene and the endogenous gene (presum-
ably PSY2 ). The profound effects of these irregularities in PSY1 and PSY2 transcription
demonstrate the importance of these genes in the regulation of carotenoid biosynthesis.

13.8 1-Deoxy- D -xylulose-5-phosphate synthase

1-Deoxy- D -xylulose-5-phosphate synthase catalyzes the condensation of hydroxyethyl thi-
amine (derived by the decarboxylation of pyruvate), with the aldehyde group of GAP, to
form DXP (Lange et al., 1998; Rohmer, 1999). Via subsequent enzymatic steps, DXP is
converted to the C 5 building blocks of the isoprenoid pathway (IPP and DMAPP) (Charon
et al., 2000; Rodrıguez-Concepcion et al., 2000).
Complementary DNAs encoding DXS have been cloned from several plant sources
that include Arabidopsis (Mandel et al., 1996; Estevez et al., 2000), peppermint (Lange
and Croteau, 1999), pepper (Bouvier et al., 1998), and tomato fruit (Lois et al., 2000). So
far, only one DXS ortholog has been found in tomato. Its expression is regulated during
development and in an organ-specific manner, showing a strong correlation to carotenoid
accumulation in tomato. The tomato DXS cDNA has an open reading frame of 2,160 bp
flanked by a 156-bp 5 -UTR and a 252-bp 3 -UTR. The cDNA encodes a 719-amino acid
polypeptide, with a predicted molecular mass of 77.6 kDa. The polypeptide shows a high
degree of sequence similarity with other known DXS polypeptides: 96% identity with that
from pepper, 83% with that from Arabidopsis , 64% with that from peppermint, and 60%
with that from Escherichia coli . Plant DXS possesses an N-terminal domain containing