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Molecular Biology of Starch-Branching and
-Debranching Enzymes |
Starch composes about 65% of the
weight of a typical cereal grain. There are two main components of
starch: the linear molecule amylose and the highly branched molecule
amylopectin. Branches are formed when starch-branching enzymes (SBEs)
break the 
-(1,4) linkage of starch and reattach the chain with an
-(1,6) bond. There are two general types of SBEs (I and II). In
maize (Zea mays), rice (Oryza
sativa), and barley (Hordeum vulgare), SBEII is
further categorized into closely homologous types IIa (a leaf form
involved in transient starch production) and IIb (an endosperm form
involved in reserve starch formation). In this issue, Blauth et
al. (pp. 1396-1405) announce their discovery of the first monocot
mutant known to be defective for SBEIIa. The structure of the leaf
starch in the mutant resembles that of the endosperm starch extracted
from amylose extender mutants of maize (defective for SBEIIb). However,
no change is reported in the endosperm. This result suggests functional
redundancy between SBEIIa and SBEIIb. The evolution of two distinct
forms could be required to meet the specific needs of leaf versus
endosperm starch synthesis. Also in this issue, Rahman et al.
(pp. 1314-1324) report on their isolation of a gene for SBEII
from wheat (Triticum aestivum) endosperm. It is surprising
that this gene is more homologous to the genes that encode for
SBEIIa in maize than for those that encode for SBEIIb. The wheat
gene was traced to the long arm of chromosome 2 (Fig.
1), and reaches peak activity 15 to
18 d after anthesis. In a second starch-related paper in this
issue, Dinges et al. (pp. 1406-1418) study the effects of
three starch-debranching enzyme (SDBE) mutants on the biosynthesis
of starch in maize. Functional SDBEs [-(1,6) glucan hydrolases]
are necessary for the formation of crystalline starch granules.
Mutations of the maize sugary1 (su1) locus, which
encodes for a major SDBE, lead to a phenotype in which the maize
kernels have a glassy, translucent, and wrinkled appearance (Fig.
2). The most interesting of the three
mutations is su1-st, which results from the insertion of a
novel transposon-like sequence that causes alternative splicing of the
mRNA to occur. Three su1-st mutants are produced: one that is nonfunctional and two that encode for modified SU1
polypeptides. The authors propose that many anomalies in the
biochemical and genetic data can be explained if it is assumed that the
various su1 mutant polypeptides, by means of quaternary
protein interactions, influence one another in affecting the ultimate
activity of the SU1 holoenzyme.
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Figure 1.
Fluorescent in situ hybrization reveals that the
SBEIIa is located on the long arm of chromosome 2 in wheat.
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Figure 2.
Sugary maize mutants (upper) are defective in an
SDBE, and have kernels that are glassy, translucent, and shrunken
compared with wild type (lower).
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Gibberellin (GA) Breakdown and Inflorescence
Formation |
The catabolism of GAs is an important factor that
regulates the endogenous levels of GAs in plants. In many plant
species, GAs are 2
-hydroxylated to produce biologically inactive GAs
in a reaction catalyzed by GA 2-oxidase. In this issue, Sakamoto et al. (pp. 1508-1516) report about their cloning and
characterization of a GA 2-oxidase gene from rice. In situ
hybridization analysis reveals that this gene is expressed in a ring at
the basal region of leaf primordia and young leaves. The drastic
reduction in this expression pattern after the phase transition from
vegetative to reproductive growth suggests that the control of GA
2-oxidase expression may play a role in the early development of the
inflorescence meristem (Fig. 3).
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What Do You Get When You Cross Oat (Avena
sativa) with Maize? |
No, this is not a child's riddle, but a scientific question
that is yielding valuable new insights into the genome map of maize.
Oat and maize are among the most remotely related plant species that
can be sexually hybridized and produce stable fertile partial hybrids.
During the early embryonic development of these hybrids, maize
chromosomes are preferentially eliminated. This enables one to isolate
allohaploid oat plants that retain a single maize chromosome. In this
issue, Kynast et al. (pp. 1216-1227) report that each of
maize's 10 chromosomes has been isolated as a separate oat-maize
addition. Fertile plants from eight of the 10 allohaploids have been
used to establish lines; plants carrying the remaining two chromosomes
are maintained clonally. This oat-maize addition is a valuable new tool
that enables any maize-specific sequence (relative to oat) to be
easily mapped to the correct chromosome. In a companion paper,
Okagaki et al. (pp. 1228-1235) demonstrate the power of
this new technique by physically mapping more than 400 sequences to the
appropriate maize chromosome by means of the PCR. By generating lines
that have only additions of partial maize chromosomes, it may be
possible to refine this technique even more so that a given
sequence can be mapped to specific regions of a chromosome.
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The Genomics of Sorghum (Sorghum bicolor): A
Stepping Stone to Maize? |
Sorghum ranks fifth in importance among the world's grain
crops. Because of its small genome (approximately 760 Mb), it will likely be the second grass species to be completely sequenced following
rice (a genomic bantam weight at only approximately 440 Mb). Sorghum,
maize, and sugarcane (Saccharum officinarum), all
members of the tribe Andropogoneae, are believed to have shared a
common ancestor as recently as 24 million yearsago, a relationship also
apparent in their similar chromosomal organizations. Draye et al.
(pp. 1325-1341) propose that sorghum, with its physically small
genome, may serve as a valuable "template" for deciphering the
larger and more complex genomes of its close relatives, especially maize. This idea gains strength from the general finding that there are
considerable similarities in gene order among the grasses. The authors
describe their progress in constructing a physical map of the sorghum
genome. The genomic map under construction is based on large-insert DNA
clones and is anchored to the recombination-based genetic
map by locus-specific sequence-tagged sites and bacterial artificial
chromosomes contigs. The authors hope ultimately to relate variations
at the molecular level to phenotypic diversity. Such "diversity
maps" will provide insights into the locations of economically
important quantitative trait loci and other genomic regions that have
been selected for during domestication. The authors also discuss the
use of cytomolecular markers as a tool for studying the cytogenetics of
sorghum, a species whose small and morphologically uniform chromosomes
have rendered it an extremely difficult organism to study by
conventional cytogenetic techniques.
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Stabilization of Transgene Expression in Cereal
Crops |
The commercial and agricultural success of transgenic crops
depends upon the stable and predictable transmission and expression of
the transgene in successive generations. It is unfortunate that the
inactivation (silencing) of transgene expression is especially common
in cereal crops, particularly in those harboring multiple copies of the
transgene. With presently available cereal transformation methods, the
number of single-copy transgenic plants generated is usually low
relative to the number of plants containing multiple copies. In this
issue, Koprek et al. (pp. 1354-1362) report on their
development of a gene delivery technique in barley that generates large
numbers of transgenic plants, each carrying a single transgene copy at
different locations. The technique is based on the maize transposable
elements Activator and Dissociation (Ac/Ds). In this system, the transgene is
inserted between the inverted repeats of the nonautonomous
Ds element and is translocated to different loci in the
genome as a result of the action of the Ac transposase. Some
of the Ds transgene cassettes transpose to genetically
unlinked sites where the chromatin may be uncondensed and where
transcription can occur. In these unlinked sites, the transgene
cassettes can segregate in the next generation from the remaining
transgene copies, as well as from other vector sequences and the
Ac transposase gene. Koprek et al. demonstrate this
technique by crossing barley plants expressing
Ac transposase with plants containing one or more copies of
a marker gene for herbicide (Basta) resistance (bar) located
between inverted repeat Ds ends (Ds-bar). Transgene expression in F2 plants with transposed
Ds-bar was 100% stable, compared with only 23% of the
F2 plants carrying Ds-bar at the
original site. This system also sheds light on the mechanism of
transgene silencing. Transposed Ds-bar was generally
inserted into low-copy regions of the genome whereas silenced
Ds-bar was generally inserted into highly repetitive
regions. Methylation of the transgene and its promoter, as well
as the higher condensation of chromatin around the integration site,
was associated with transgene silencing.
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Molecular Biology of Starch-...
Gibberellin (GA) Breakdown and...