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Adenosine Kinase-Deficient Mutants |
Adenosine kinase (ADK) catalyzes
the salvage synthesis of adenine monophosphate from adenosine and ATP.
ADK is encoded in Arabidopsis by two closely related cDNAs that are
constitutively, yet differentially, expressed in leaves, stems, roots,
and flowers. To investigate the role of ADK in plant metabolism,
Moffatt et al. (pp. 812-821) have created ADK-deficient
lines by sense and antisense expression of the ADK1 cDNA.
Transgenic plants with less than 10% ADK activity are small with
rounded, wavy leaves and a compact, bushy appearance. Due to the lack
of elongation of the primary shoot, the siliques extend in a cluster
from the rosette (Fig. 1). Fertility is
decreased because the stamen filaments do not elongate normally;
hypocotyl and root elongation are reduced also. The authors propose
that the numerous morphological changes observed in the ADK-deficient
lines may reflect the methylation requirements of different tissues.
Because previous biochemical studies have shown that the hydrolysis of
S-adenosyl-L-homo-Cys (SAH) is major
source of adenosine, the authors hypothesize that adenosine must be
steadily removed by ADK to prevent feedback inhibition of SAH
hydrolase. Consistent with this idea, the ADK-deficient lines were
shown to have increased levels of SAH. Since all methyltransferases that use S-adenosyl-Met (SAM) are inhibited by SAH, the lack
of adenosine salvage in the ADK-deficient lines and the consequent increase in SAH levels may, by negative feedback mechanisms, inhibit SAM-dependent transmethylation reactions needed for proper
development.
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Figure 1.
Arabidopsis mutants deficient in adenosine kinase
activity exhibit a squat phenotype, possibly because the buildup of
adenosine causes, by negative feedback mechanisms, a decrease in
transmethylation reactions.
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Multiple Gene Suppression by Chimeric Silencing
Constructs |
Much work describes the use of transgenic technologies to
manipulate genes of important biochemical pathways. In most cases, only
single genes have been manipulated, usually by down-regulating their
activity using antisense RNA or co-suppression. Although this work has
been extremely illuminating, full exploitation of the potential for
plant metabolic engineering will likely necessitate the manipulation of
multiple genes. A case in point concerns the lignin biosynthetic
pathway. Although the basic pathway was outlined many years ago, recent
data has led to multiple suggestions being made concerning alternative
routes by which the synthesis of certain intermediates may occur. In
this issue, Abbott et al. (pp. 844-853) employ a clever
method for achieving the coordinate suppression of multiple genes that
encode for proteins involved in lignin biosynthesis. Single chimeric
transgenes were produced by fusing partial sense sequences for all
possible combinations of two, or of all three genes encoding for the
lignin biosynthesis enzymes, caffeate/5-hydroxyferulate
O-methyltransferase (COMT), cinnamoyl-CoA reductase (CCR),
and cinnamoyl alcohol dehydrogenase (CAD). Compared with the results
obtained when sexual crossings were used as a means to combine distinct
antisense transgenes for the same target genes, plants transformed with
the chimeric partial sense constructs had consistently higher levels of
suppression of target enzymes. They also exhibited significant changes
in lignin content and plant structure (Fig.
2). Chimeric silencing constructs offer
great potential for the rapid and co-ordinate suppression of multiple
genes encoding for enzymes involved in other biochemical
pathways.
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Figure 2.
Tobacco (Nicotiana tabacum) mutants
deficient in three genes encoding for lignin biosynthesis enzymes
(right) exhibit a squat phenotype compared with wild type (left).
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Different Functional Roles for Ethylene
Receptors |
Current evidence suggests that tomato (Lycopersicon
esculentum) has at least five ethylene receptors
(LeETR1-LeETR5) that have distinct expression patterns and
somewhat overlapping functions. The binding of ethylene inhibits the
signaling of these receptors. Ethylene receptors generally have three
domains: a sensor, a His kinase, and a receiver domain (response
regulator), but in some cases certain of these domains are lacking. For
example, the mutant gene here referred to as LeETR3,
which causes the Never-Ripe phenotype in tomato, lacks the receiver
domain. Using the Arabidopsis ETR1 cDNA as a probe, two
orthologs of the Arabidopsis cDNA in tomato (recently renamed
LeETRI and LeETR2) have been identified.
Both LeETR1 and LeETR2 possess the same
three domains as the Arabidopsis ethylene receptor
AtETR1 (sensor, His kinase, and receiver domain), whereas LeETR3, like its counterpart in Arabidopsis,
lacks the receiver domain. Whitelaw et al. (pp. 978-987)
examine the possibility that these structural variations may serve as the basis for functional differences between the receptors. They transformed tomato plants with a construct containing the antisense sequence for the receiver domain and the 3' untranslated portion of
LeETR1. Because of the high nucleotide sequence identity
between the receiver domains of LeETR1 and
LeETR2, they presume that both of these receptors were
inhibited by their construct. The two most consistently observed
phenotypes in the transgenic lines were delayed abscission and reduced
plant size. Fruit coloration and softening and the triple response of
seedlings from the next generation were all unaffected. Delayed
abscission and shorter internode length segregated with the transgene
and were correlated with the degree of reduced LeETR1
transcript accumulation. The authors propose that ethylene signal
transduction occurs through parallel pathways that partially intersect
to regulate shared ethylene responses.
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Structural Analysis of a Soybean Seed Desiccation
Protein |
Late embryogenesis abundant (LEA) proteins accumulate
in developing plant embryos prior to the desiccation that marks the last stage in seed development. LEA proteins also accumulate in vegetative tissues subjected to drought, osmotic or low-temperature stress, or treated with exogenous ABA. LEA proteins, the majority of
which are highly hydrophilic, are part of a larger, evolutionarily conserved group of hydrophilic proteins termed "hydrophilins" that
are involved in various adaptive responses to dehydrating conditions.
Group 1 LEA proteins are distinguished from other groups of LEA
proteins by being very conserved along the entire length of the
protein. Although much evidence indicates a role for group 1 LEA
proteins in desiccation tolerance, the mechanism by which they work
remains controversial. In this issue, Soulages et al. (pp.
822-832) employ several physical techniques to analyze the
structure of a group 1 LEA protein from soybean (Glycine max). Circular dichroism spectral measurements indicated that a
minimum of 14% of the amino acid residues of this LEA protein exist in
a solvent-exposed, left-handed extended helical or poly (L-Pro)-type (PII) conformation, with the
remainder of the protein being unstructured. The authors hypothesize
that by favoring the adoption of a PII structure, instead of the
formation of 
-helical or 
-sheet structures, group 1 LEA proteins
retain a high content of surface area available for interaction with
water. This feature may constitute the basis by which LEA proteins
function in preventing freezing, desiccation, or osmotic stress damage.
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Extensin Cell Wall Protein Involved in Root Hair Tip
Growth |
Extensins are abundant cell wall proteins. In this
issue, Bucher et al. (pp. 911-923) report on their cloning
of a cDNA encoding an extensin-like protein (LeExt1) from a
tomato (Lycopersicon esculentum) root hair cDNA library.
Patterns of mRNA distribution indicated that the expression of the
LeExt1 gene was initiated in the root hair differentiation
zone of the tomato epidermis. A number of findings suggest a direct
correlation between LeExt1 expression and cellular tip
growth. For example, LeExt1/-glucuronidase
(Lext1/GUS) expression was detectable only in the root
hair-forming cells of the root epidermis. Moreover, both hair formation
and LeExt1 expression were inducible by the plant hormone
ethylene. Comparative analyses of the LeExt1/GUS expression
patterns were performed in other Solanaceous plants and in Arabidopsis.
In the apical/basal dimension, GUS staining was absent from the root
cap and undifferentiated cells at the root tip in all species
investigated. It was induced at the distal end of the differentiation
zone and remained high proximally to the root/hypocotyl boundary. In
the radial dimension, GUS expression was root hair-specific in all of
the Solanaceous species. These data support a role for
LeExt1 in root hair tip growth.
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