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33
Seed maturation: genetic programmes and control signals
Ulrich Wobus* and Hans Weber
Seed maturation is mainly governed by a few genes best
studied in maize and Arabidopsis. The isolation of the LEC1
and FUS3 genes, besides the previously known VP1/ABI3
genes, and their identification as transcriptional regulators
provides the first direct hints as to their molecular mode of
action. With the identification of new effector genes, the
investigation of the role of hormones with new methods such
as immunomodulation and the increasingly recognised role
of metabolites like sugars as important modulators of seed
development, we increasingly understand the complexity
and structure of the regulatory network underlying
seed maturation.
Addresses
Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK),
D-06466 Gatersleben, Germany
*e-mail: [email protected]
Current Opinion in Plant Biology 1999, 2:33–38
http://biomednet.com/elecref/1369526600200033
© Elsevier Science Ltd ISSN 1369-5266
Abbreviation
ABA
abscisic acid
Introduction
Higher plants are characterised by the formation of seeds
which contain the embryo, protected by the maternallyderived seed coat. The embryo first runs through a phase
of cell division and morphogenesis, followed by a maturation phase which includes the accumulation of storage
products, the suppression of precocious germination, the
acquisition of desiccation tolerance, water loss and often
the induction of dormancy (see [1•]). Maturation interrupts seedling development. The process progresses in a
wave-like manner forming temporally and spatially
determined patterns or gradients between and within
the different seed organs [2,3]. Several lines of evidence
strongly suggest that maturation evolved late in evolution and has been ‘inserted’ [1 •] at different
developmental time points in different higher plant taxa
[4•]. In depth molecular analyses, however, are mainly
carried out in Arabidopsis, grain legumes and cereals
(especially maize), in each case with a specific research
focus depending on the different advantages of the three
experimental systems. In this review, the following
exciting recent research achievements are briefly discussed; the isolation and initial functional analysis of
central transcriptional regulators of seed maturation and
some of their target genes, a new methodology to control
the process at the hormonal level and the role of metabolites as triggers of maturation processes. These data
necessarily cover only certain aspects of the complex
network of processes involved in seed maturation.
ABA3, FUS3 and LEC1: three major regulators
of seed maturation
Analyses of mutant phenotypes in Arabidopsis have provided
strong evidence that three genes — ABA-INSENSITIVE3
(ABI3), FUSCA3 (FUS3) and LEAFY COTYLEDON1
(LEC1) — are of utmost importance for seed maturation as
they affect a wide, broadly overlapping but not identical
spectrum of seed-specific characters (see Figure 9 in [5••]
for a good graphic representation). In essence, mutant
embryos enter the germination programme immediately
by skipping maturation. The description of the abi3
mutant as abscisic acid insensitive already indicates a link
to the most important hormonal regulator of seed maturation, abscisic acid (ABA). Until recently, only the molecular
structure of the ABI3 gene [6] and its maize homologue,
the transcriptional activator gene VP1 [7], were known. In
1998, however, the sequences of two other genes, FUS3
and LEC1, have been obtained [8••,9••] and led to the
important conclusions discussed below.
The FUS3 gene turned out to be related to the VP1/ABI3
gene family, but is of reduced length (ABI3 encodes 720
amino acid residues; FUS3 only 312 residues), and high
homology in the protein is restricted to a stretch of at least
100 amino acid residues [9••] defined as the B3 region in
the VP1 protein [10]. The B3 domain can mediate
sequence-specific DNA-binding in vitro [11•] and is critical
for gene activation at low or insignificant ABA concentrations [12•]. It was also used to define a B3-box family of
genes (or B3-domain family of proteins) [9••] of which
FUS3 clearly is a member of a — possibly more ancient —
subgroup. An auxin-response factor ARF1 [13] represents
another even more remote subfamily. In accordance with
the ABA independence of fus3 mutants, the FUS3 protein
lacks an amino-terminal A-domain which, in VP1 (and certainly in ABI3), mediates the ABA response [12•]. FUS3 is
often classified as an LEC1-type gene on the basis of
mutant phenotypes (see [1•,8••,14]), but the two genes are
structurally unrelated and should, therefore, stay separate.
LEC1 has recently been shown most probably to encode a
protein related to a transcription factor subunit of the HAP3
type [8••] — HAP3 in mammals is part of the heterotrimeric CCAAT box-binding factor CBF (see [15]).
On the basis of an extensive descriptive study of double
and the parental single mutants, Parcy et al. [5••] had
already concluded that the three genes act synergistically. The authors proposed a model for the concerted
action of the gene trio in controlling several major
aspects of seed maturation. They further speculated that
the three proteins could act as transcription factors and
form hetero-oligomeric combinations, which regulate
different developmental processes. These protein complexes could also feed-back regulate their own gene
34
Growth and development
promoters as shown, for instance, for the DEF A/GLO
heterodimer in Antirrhinum [16]. DEF A and GLO are
two MADS-box transcription factors involved in flower
development. Preliminary experiments at the molecular
level involving ABI3 and FUS3 [17•] provide first hints
that these suggestions may be valid. In a transient
expression assay both FUS3 and ABI3 strongly induce
the activity of maturation-specific promoters in a cooperative and synergistic way. This induction was strictly
dependent on an intact RY (CATGCAT) promoter-ciselement and suggests a rather direct mode of action,
perhaps direct binding. Also LEC1, due to its proposed
HAP3-like function [8••], could directly bind to the RY
repeat. In maize and other grasses, the same element
(here called the Sph motif) is functionally dependent on
the VP1 gene product, specifically on the B3 domain, in
promoters of genes like C1, a transcriptional regulator of
anthocyanin biosynthesis in seeds [11•]. VP1 not only
recognises the Sph cis-element [11•], but may also act by
its B2 domain as a protein association enhancer [10]. In
an Em-promoter G-box binding complex, VP1 is associated with a 14-3-3 protein (GF14), and VP1, the G-box
transcription factor EmBP1 and GF14 may all interact to
stabilise and/or activate the transcription complex
[18•,19•]. Em is an ABA-responsive late embryogenesis
abundant (LEA) gene expressed exclusively in maturing
embryos. Additional b-ZIP factors, modifying ABI3
homologue-dependent gene expression, have been characterised from pea [20,21] and underscore the complex
nature of the molecular machinery directing gene regulation during seed maturation.
Studies on genes affected by FUS3 [22•,23•] revealed
another interesting new aspect; among the genes controlled by FUS3 during maturation are upregulated [22•]
and downregulated [23 •] MYB transcription factors.
AtMYB13, for instance, is involved in some aspects of
meristematic vegetative growth. It is likely to be a representative of several regulators of vegetative growth
repressed during maturation by the concerted action of
at least FUS3 and LEC1 [23•]. The data also indicate that
FUS3 is, on the one hand, a central regulator controlling
other regulators such as MYB transcription factors but,
on the other hand, the protein probably directly regulates effector genes like maturation-specific storage
protein genes via the RY promoter cis-element.
The ABI3/VP1 transcriptional activator family is well
known from mutant phenotypes to be involved in establishing embryo dormancy. Recent work with Avena fatua
inbred lines suggests that VP1 plays a central role since
AfVP1 mRNA levels established during the final stages of
embryo maturation were positively correlated with the dormant phenotype in a way that qualifies the gene as a
molecular marker for dormancy potential/after-ripening
time [24•]. The most recent discovery of a role of an ABI3
homologue in quiescent apices of Arabidopsis [25•] signals
that the function of genes of the ABI3-type, and perhaps
also of the FUS3-type (low expression levels in non-seed
tissues [9••]), is not restricted to seeds. Interestingly,
another newly isolated transcriptional regulator, ABI4,
most likely to be involved in regulating seed responses to
ABA, is not expressed seed-specifically in spite of the fact
that a clear mutant phenotype is only seen in seeds [26•].
According to published data [8••], only LEC1 expression
seems to be strictly embryo-specific but studies with more
sensitive assays like RT-PCR have not yet been reported.
In summary, three major genes involved in seed maturation initiation and maintenance are structurally either not
(LEC1) or only distantly (ABI3/FUS3) related and their
expression patterns are temporally and spatially different,
but broadly overlapping especially during seed maturation.
At least ABI3 and FUS3 are expressed in tissues other than
seed, that is to say they are not strictly seed-specific, contrary to accepted beliefs. The door is now open to study
the molecular interactions of the gene products with each
other and other factors and their role in determining cell
fate and function in each specific process.
A fresh look at the role of abscisic acid:
probing abscisic acid function by expressing
ABA-antibodies
The important role of ABA in seed maturation is well documented but since the hormone triggers diverse processes
not only in the seed, analysis by established procedures
faces limitations. A method which blocks the ABA molecule itself very specifically in a tissue-, cell- and
compartment-specific manner [27•], therefore, promises
new insights. Phillips et al. [28••] directed an ABA-specific
single chain Fv (scFv) antibody into the lumen of the endoplasmic reticulum (ER) of developing tobacco seeds by
using the ER-target signal KDEL and a seed-specific promoter, active from the late phase of early development to
the first part of late embryogenesis. The anti-ABA scFv
caused a developmental switch to the germination programme characterised by the formation of chloroplasts
containing photosynthetic pigments and a dramatic reduction in the embryo of the most abundant storage proteins as
well as protein and oil bodies. The authors calculated that
the recombinant antibody bound nearly all available ABA
until day 20 of tobacco development. The overall concentration of antibody-bound ABA plus unbound ABA
increased considerably above wild-type level, possibly due
to a feed-back mechanism or protection from ABA metabolism. The seed phenotype was most similar to the
Arabidopsis aba/abi3 double mutant seeds [29]. The well
established fact that ABA biosynthetic mutants do not
markedly reduce storage protein mRNAs [29] led to the
question — how important is ABA in controlling storage
protein accumulation [30•]? The anti-ABA seeds now clearly underscore the dominating role of ABA in the process at
least in tobacco seeds [28••]. By using a broader range of
promoters and scFvs against other regulatory molecules,
immunomodulation promises to contribute considerably to
a deepened understanding of seed development.
Seed maturation: genetic programmes and control signals Wobus and Weber
Development and metabolism: renewed
interest in a long known connection
Metabolites as developmental signals in seeds
Metabolites as signal molecules in plant developmental
processes have been mainly neglected until recently when
sugars especially were shown to regulate diverse gene activities [31] and to act as important components in signal
transduction networks [32•]. In grain legume seeds, extensive, mainly correlative analyses (see [33•,34•] for reviews)
including transgenic plants which express a yeast-derived
invertase in developing seeds [35], suggested that soluble
sugars provide signals for cotyledon development but that
glucose and sucrose can play different roles; whereas a high
glucose/sucrose ratio was correlated with cell division, a high
sucrose/hexose ratio seems to be a major trigger of the storage product pathways. Of special interest is the discovery of
steep glucose gradients across the cotyledons of the grain
legume Vicia faba, which change during development and
correlate positively with mitotic activity but negatively with
storage starch accumulation [36•]. These gradients are likely
to be primarily established by the specific expression pattern
of a cell wall-bound invertase in the developing maternal
seed coat [37] and further modulated (enhanced?) by a hexose- and a sucrose transporter (mainly excluding each other
in the timing of expression) in the epidermal cells of the
embryo [38•]. Gradients were also visualised in developing
maize kernels at the level of the activity of a key enzyme —
sucrose synthase — of starch biosynthesis in sink organs
[39•]. The combination of methods which allow the (semi-)
quantitative determination not only of RNA and protein but
also of enzyme activity and metabolites at (or nearly at) cellular resolution will certainly reveal a refined net of
relationships. Whether sugar levels in such gradients function as morphogenic substances [36•], however, has not been
shown unequivocally. Whatever the exact role of sugars is,
the elicited signals have to interact with other signals to
evoke the complex processes of seed maturation.
One site of integration for different signals are promoter
sequences. Interestingly, it has recently been demonstrated
by analysing the phaseolin seed storage protein promoter in
transgenic tobacco seeds that the essential signal for the transcriptional activity of the phaseolin promoter provided by
ABA is modulated beside unknown developmental factors
by sucrose and calcium ions [40•]. In the barley embryo, glucose acting as a signal molecule interferes with the
gibberellic acid response of α-amylase gene expression, but
in a tissue-specific manner [41•]; the induction of α-amylase
by GA in the aleurone layer is unaffected by sugars, whereas
in the embryo, sugars repress the response. The site of action
of the sugar molecule has not been determined, but could be
at the transcriptional and also at the post-transcriptional
level, as rice α-amylase mRNA stability and abundance have
been shown to be controlled by sucrose [42•,43•].
Assimilate import and seed filling
If metabolites are able to act as signals in developmental
processes, the regulated import of photoassimilates into the
35
growing seed will influence seed development beside by
providing nutrients. Developing seeds import sucrose and
amino acids from the phloem. These assimilates are
unloaded from the seed coat and must be taken up by the
apoplastically isolated embryo or endosperm. Seed-located
apoplastic transport processes, therefore, can influence
assimilate partitioning and storage product synthesis. One
obvious regulatory control point could be provided by various transporters situated in the membranes of seed coat
and/or embryo epidermal cells. In Vicia faba a sucrose transporter gene is highly expressed in the epidermal transfer cell
layer covering the outer surface of developing cotyledons
[38•,44]. These transfer cells develop at the contact area of
the cotyledonary epidermis to the seed coat, starting first at
early cotyledon stage and subsequently spreading to the
abaxial region. The tissue underlying the epidermal transfer
cell layer differentiates into storage parenchyma cells, indicating an important role of the sucrose transporter in
providing sugars for storage product synthesis. Feeding high
concentrations of sugars in vitro suppressed both the sucrose
transporter gene expression and transfer cell differentiation,
suggesting a control mechanism by carbohydrate availability
[38•]. A more detailed study [45•] also demonstrated the
induction of transfer cell differentiation by sugars and the
modulation by the sugar state of sugar transport activity and
consequently storage product synthesis. Less detailed data
are available for amino acid transporters which have a broad
substrate specificity and are probably regulated by tissuespecific expression. Two members were found to be
expressed in developing seeds of Arabidopsis. Using promoter-GUS fusions it was shown that AAP1 is expressed in the
endosperm and the cotyledons whereas AAP2 is expressed in
the vascular strands of siliques and in funiculi [46•].
In addition to these seed-specific processes, there are
genes acting on the whole plant level in controlling
assimilate transport into the seed and seed growth. One
example is the SN gene in pea which regulates assimilate
partitioning between vegetative and reproductive
growth [47•]. One important factor could be gibberellic
acid (GA) since it appears that in the pea lh2 mutation
seeds are unable to use available assimilates. In developing lh2-seeds, GA levels are reduced and lead to reduced
growth and to a high frequency of seed abortion. GAs
may either promote assimilate uptake into developing
seeds or affect seed growth, that is sink strength [48•].
The general significance of seed-specific transport
processes for development, however, is still unknown.
Conclusions
Substantial progress has been made in the past two years
in elucidating the structure and the mode of action of
genes involved in embryo development and seed maturation. This progress will now gain much momentum,
especially by the Arabidopsis genome sequencing project.
Furthermore, some signal transduction chains with their
components and branch points are slowly emerging and
mostly correlative analyses suggest a number of mainly
36
Growth and development
hitherto neglected factors modulating or even triggering
developmental processes. The still scarce data suggest the
existence of an extended regulatory network rather than a
clear hierarchy of actions and allows first glimpses on its
structure. Large scale expression analysis by DNA chip
technology will uncover many more possible network components especially at the gene level. The real challenge,
however, will be to unravel their role and interconnection.
Beside the genetic approach, with its focus on gene structure and mRNA expression level, more extensive studies
at the protein and higher order structural level (subcellular,
cellular and also whole-plant level), together with more
integrated experimental approaches are necessary to further deepen and extend our knowledge of seed maturation
as part of seed development and plant life in general.
Acknowledgements
The authors would like to thank Helmut Bäumlein and Udo Conrad for
help with the manuscript and Simon Miséra and Antje Rhode for providing
manuscripts in press. Work in our lab is supported by the Deutsche
Forschungsgemeinschaft and the European Union.
References and recommended reading
Papers of particular interest, published within the annual period of review,
have been highlighted as:
• of special interest
•• of outstanding interest
Harada JJ: Seed maturation and control of germination. In Cellular
and Molecular Biology of Plant Seed Development. Edited by Larkins
BA, Vasil IK. Dordrecht: Kluwer Academic Publishers; 1997:545-592.
Extensive and recommended review covering all aspects of seed maturation.
1.
•
2.
Perez-Grau L, Goldberg RB: Soybean seed protein genes are
regulated spatially during embryogenesis. Plant Cell 1989,
1:1095-1109.
3.
Borisjuk L, Weber H, Panitz R, Manteuffel R, Wobus U:
Embryogenesis in Vicia faba L.: histodifferentiation in relation to
starch and storage protein synthesis. J Plant Physiol 1995,
147:203-218.
Kaplan DR, Cooke TJ: Fundamental concepts in the
embryogenesis of dicotyledons: a morphological interpretation of
embryo mutants. Plant Cell 1997, 9:1903-1919.
This paper is well worth reading. It represents, in essence, a minority view by
challenging the concept of pattern formation in plants, but also brings to
mind the comparative morphological approach to embryogenesis often
neglected by molecular biologists.
4.
•
Parcy F, Valon C, Kohara A, Miséra S, Giraudat J: The ABSCISIC
ACID-INSENSITIVE3, FUSCA3, and LEAFY COTYLEDON1 loci act
in control multiple aspects of Arabidopsis seed development.
Plant Cell 1997, 9:1265-1277.
In this important paper, the authors, by quantifying several developmental
processes in a range of mutants, clearly show that the three genes work synergistically and not in different pathways as earlier models suggested. They
also provide evidence that the abundance of ABI3 is regulated by FUS3 and
LEC1. The genetic model provided can now be tested at the molecular level
due to the availability of FUS3 and LEC1.
5.
••
Homology of LEC1 to HAP3 subunit sequences of the CCAAT-box binding factor CBF is restricted to the HAP3 B-domain.
Luerßen H, Kirik V, Herrmann P, Miséra S: FUSCA3 encodes a
protein with a conserved VP1/ABI3-like B3 domain which is of
functional importance for the regulation of seed maturation in
Arabidopsis thaliana. Plant J 1998, 15:755-764.
A third key regulatory gene of seed maturation beside ABI3 and LEC1,
FUS3, has been isolated by postional cloning and its product shown to
share the B3 domain with ABI3-like proteins, whereas significant features of
A- and B1-domains are missing, obviously due to the short amino-terminal
part of this protein. Whether some similarities to B2-domain sequences are
of functional importance has still to be shown. The analysed allelic variants
(most of them causing abnormal splicing) provide hints for the essential role
of the B3 domain in seed maturation (but see [12•]).
9.
••
10. Hill A, Nantel A, Rock CD, Quatrano RS: A conserved domain of the
VIVIPAROUS-1 gene product enhances the DNA binding activity
of the bZIP protein EmBP-1 and other transcription factors. J Biol
Chem 1996, 271:3366-3374.
11. Suzuki M, Kao CY, McCarty DR: The conserved B3 domain of
•
VIVIPAROUS1 has a cooperative DNA binding activity. Plant Cell
1997, 9:799-807.
Since its isolation, VP1 has been implicated in gene regulation, but the question remained open as to whether or not the protein binds directly to DNA.
Now the McCarty lab provides convincing evidence that the B3 domain of
VP1 has a highly co-operative, sequence-specific DNA-binding activity in an
in vitro assay. Specific binding to the Sph-motif TCCATGCAT provides a
good link to the expected functions and leads to the suggestion that the B3
domain in general binds to this element (also known as the RY-motif) present
in many seed protein gene promoters.
12. Carson CB, Hattori T, Rosenkrans L, Vasil V, Vasil IK, Peterson P,
•
McCarty DE: The quiescent/colourless alleles of viviparous-1 show
that the conserved B3 domain of VP1 is not essential for
ABA-regulated gene expression in the seed. Plant J 1997,
12:1231-1240.
By analysing several vp1 alleles in a further attempt to map functional
domains within the VP1 protein in more detail, the authors conclude that B3
is not absolutely essential for completion of seed maturation. Contrary to
predictions in [9••], a two amino acid deletion in the B3 domain of FUS3 still
led to the Fus3– phenotype in Arabidopsis. Whether this apparent discrepancy is of general importance has still to be worked out.
13. Ulmasov T, Hagen G, Guilfoyle TJ: ARF1, a transcription factor that
binds to auxin response elements. Science 1997, 276:1865-1868.
14. Vernon DM, Meinke DW: Late embryo-defective mutants of
Arabidopsis. Dev Genet 1995, 16:311-320.
15. Maity SN, de Crombrugghe B: Role of the CCAAT-binding protein
CBF/NF-Y in transcription. Trends Biochem Sci 1998, 23:174-178.
16. Schwarz-Sommer Z, Hue I, Huijser P, Flor PJ, Hansen R, Tetens F,
Lönnig W-E, Saedler H, Sommer H: Characterization of the
Antirrhinum floral homeotic MADS-box gene deficiens:
evidence for DNA binding and autoregulation of its persistent
expression throughout flower development. EMBO J 1992,
11:251-263.
17.
•
Wohlfarth T, Braun H, Kirik V, Kölle K, Czihal A, Tewes A, Luerssen H,
Miséra S, Shutov A, Bäumlein H: Regulation and evolution of seed
globulin genes. J Plant Physiol 1998, 152:600-606.
This symposium paper contains interesting data which have still to be published in detail. They suggest a co-operative interaction of FUS3 and ABI3
in the regulation of seed-specific promoters with the RY cis-motif as target.
6.
Giraudat J, Hauge BM, Valon C, Smalle J, Parcy F, Goodman HM:
Isolation of the Arabidopsis ABI3 gene by positional cloning. Plant
Cell 1992, 4:1251-1261.
18. Schultz TF, Medina J, Hill A, Quatrano RS: 14-3-3 proteins are part
•
of an abscisic acid-VIVIPAROUS1 (VP1) response complex in the
Em promoter and interact with VP1 and EmbP1. Plant Cell 1998,
10:837-847.
The paper provides a first and important example of a transcription complex
involving VP1 and identifies members of the complex including 14-3-3 proteins which are supposed to act as mediators in protein–protein interactions
involved in kinase-related steps of signal-transduction pathways.
7.
McCarty DR, Carson CB, Stinad PS, Robertson DS: Molecular
analysis of viviparous-1: an abscisic acid-insensitive mutant
maize. Plant Cell 1989, 1:523-532.
19. Quatrano RS, Bartels D, Ho T-HD, Pagés M: New insights into
•
ABA-mediated processes. Plant Cell 1997, 9:470-475.
A meeting-based review including the role of ABA in seed maturation.
Lotan T, Ohto M-A, Yee KM, West MAL, Lo R, Kwong RW, Yamagishi
K, Fischer RL, Goldberg RB, Harada JJ: Arabidopsis LEAFY
COTYLEDON1 is sufficient to induce embryo development in
vegetative cell. Cell 1998, 93:1195-1205.
The authors isolated the LEC1 gene and demonstrated its major role, not
only during late, but also in early embryogenesis including suspensor and
endosperm development. Ectopic LEC1 expression, in contrast to ABI3,
induced embryonic programmes in vegetative cells in rare cases.
20. Chern MS, Bob AJ. Bustos MM: The regulator of MAT2 (ROM2)
protein binds to early maturation promoters and represses
PvALF-activated transcription. Plant Cell 1996, 8:305-321.
8.
••
21. Chern MS, Eiben HG, Bustos MM: The developmentally regulated
bZIP factor ROM1 modulates transcription from lectin and
storage protein genes in bean embryos. Plant J 1996,
10:135-148.
Seed maturation: genetic programmes and control signals Wobus and Weber
22. Kirik V, Kölle K, Miséra S, Bäumlein H: Two novel MYB homologues
•
with changed expression in late embryogenesis-defective
Arabidopsis mutants. Plant Mol Biol 1998, 37:819-827.
See [23•] for annotation to this reference.
23. Kirik V, Kölle K, Wohlfarth T, Miséra S, Bäumlein H: Ectopic
•
expression of a novel MYB gene modifies the architecture of the
Arabidopsis inflorescence. Plant J 1998, 13:729-742.
Papers [22•] and [23•] contribute to the elucidation of the key role of the
ABI3/LEC1/FUS3 gene trio in late embryogenesis by the identification,
via differential display, of effector genes — in this case members of the
MYB group of transcriptional regulators. The genes described are either
positively [22•] or negatively [23•] regulated. AtMYB13 is activated in late
embryogenesis defective fus3 and lec1 mutants but not abi3 mutants,
that is to say, in wild-type embryos this MYB protein should be at least in
part responsible for repressing genes involved in vegetative meristem
activation. The report [23•] provides the first evidence at the molecular
level for different action spectra of ABI3 on the one hand and FUS3 plus
LEC1 on the other.
24. Jones HD, Peters NCB, Holdsworth MJ: Genotype and environment
•
interact to control dormancy and differential expression of the
VIVIPAROUS-1 homologue in embryos of Avena fatua. Plant J
1997, 12:911-920.
This is yet another good example that the well-chosen use of plants other
than Arabidopsis can considerably contribute to the better understanding of
specific developmental processes.
25. Rhode A, Van Montagu M, Boerjan W: The abscisic acid-insensitive
•
gene ABI3 is expressed during vegetative quiescence processes
in Arabidopsis. Plant Cell Environ 1998, in press.
Here, Rhode et al. show, in contrast to present conceptions, that ABI3 most
probably has additional functions to those in seed development. During
growth in the dark ABI3 is expressed in the seedling apex after the arrest of
cell division. Concomitantly, the 2S napin seed storage protein is induced.
ABI3-promoter/GUS fusion constructs are expressed in still other vegetative organs. These results — together with the observed expression of an
ABI3 homologue in dormant poplar buds (A Rhode, Molecular strategies
to study bud dormancy in Populus [PhD thesis]. Universiteit Gent
1998) — led the authors to conclude that ABI3 is one of several components
of a molecular network underlying all types of quiescence.
26. Finkelstein RR, Wang ML, Lynch TJ, Rao S, Goodman H: The
•
Arabidopsis abscisic acid response locus ABI4 encodes an
APETALA2 domain protein. Plant Cell 1998, 10:1043-1054.
ABA-sensitive abi4 mutants are defective in seed development but exhibit a
less severe phenotype than known abi3 mutants. The gene, isolated by positional cloning, might represent a new element in the signal transduction
pathway involving ABI3.
Conrad U, Fiedler U: Compartment-specific accumulation of
recombinant immunoglobulins in plant cells: an essential tool for
antibody production and immunomodulation of physiological
functions and pathogen activity. Plant Mol Biol 1998, 38:101-109.
A review focusing on the importance of cell compartment-specific expression
of antibodies with respect to both functional studies or biotechnological goals.
27.
•
28. Phillips J, Artsaenko O, Fiedler U, Horstmann C, Mock H-P, Müntz K,
•• Conrad U: Seed-specific immunomodulation of abscisic acid
activity induces a developmental switch. EMBO J 1997,
16:4489-4496.
The paper demonstrates for the first time the successful use of transgene
encoded antibodies directed against low molecular weight regulatory compounds for the study of plant developmental processes. This new experimental tool will be certainly used in the future in a variety of situations
including other hormones and non-protein regulators.
29. Koornneef M, Hanhart CJ, Hilhorst HWM, Karssen CM: In vivo
inhibition of seed development and reserve protein accumulation
in recombinants of abscisic acid biosynthesis and
responsiveness mutants in Arabidopsis thaliana. Plant Physiol
1989, 90:463-469.
30. Merlot S, Giraudat J: Genetic analysis of abscisic acid signal
•
transduction. Plant Physiol 1997, 114:751-757.
One of several recommendable recent reviews (see also [19•]) on ABA signal transduction.
31. Koch KE: Carbohydrate-modulated gene expression in plants.
Annu Rev Plant Physiol Plant Mol Biol 1996, 47:509-540.
32. Jang J-C, Sheen J: Sugar sensing in higher plants. Trends Plant Sci
•
1997, 2:208-214.
Recommended review on a rapidly expanding research field.
33. Weber H, Borisjuk L, Wobus U: Sugar import and metabolism
•
during seed development. Trends Plant Sci 1997, 2:169-174.
See [34•] for annotation for this reference.
37
34. Weber H, Heim U, Golombek S, Borisjuk L, Wobus U: Assimilate
•
uptake and the regulation of seed development. Seed Sci Res
1998, 8:331-345.
Reviews (condensed [33•] and more extended [34•]) focusing on the possible role of sugars in regulating cotyledon development.
35. Weber H, Heim U, Golombek S, Borisjuk L, Manteuffel R, Wobus U:
Expression of a yeast-derived invertase in developing cotyledons
of Vicia narbonensis alters the carbohydrate state and affects
storage functions. Plant J 1998, 16:163-172.
36. Borisjuk L, Walenta S, Weber H, Müller-Klieser W, Wobus U: High
•
resolution histographical mapping of glucose concentrations in
developing cotyledons of Vicia faba in relation to mitotic activity
and storage processes: glucose as a possible developmental
trigger. Plant J 1998, 15:583-591.
A luciferase-coupled enzyme reaction together with quantitative bioluminescence and single photon imaging is used to visualise glucose concentration
gradients which are related to specific cell types. In addition, they correlate
positively with mitotic activity and negatively with starch accumulation. The
authors speculate about glucose concentration ‘landscapes’ as possible
morphogenetic gradients.
37.
Weber H, Borisjuk L, Heim U, Buchner P, Wobus U:
Seed coat-associated invertases of fava bean control both
unloading and storage functions: cloning of cDNAs and cell-type
specific expression. Plant Cell 1995, 7:1835-1846.
38. Weber H, Borisjuk L, Heim U, Sauer N, Wobus U: A role for sugar
•
transporters during seed development: molecular
characterization of a hexose and a sucrose carrier in Fava bean
seeds. Plant Cell 1997, 9:895-908.
The authors back their hypothesis on the important role of sugars in cotyledon development by describing correlations between the expression of
sugar transporters, sugar levels and cell type differentiation. Additional
experimental evidence is drawn from work with explanted cotyledons.
39. Wittich PE, Vreugdenhil D: Localization of sucrose synthase activity
•
in developing maize kernels by in situ enzyme histochemistry.
J Exp Bot 1998, 49:1163-1171.
A much welcomed piece of work bringing enzyme activity studies in seeds
down to the cellular level. Such studies, together with high-resolution
metabolite analysis (see [36•]) and gene expression studies, will increasingly contribute to a better understanding of developmental processes.
40. Bustos MM, Iyer M, Gagliardi SJ: Induction of a ß-phaseolin
•
promoter by exogenous abscisic acid in tobacco: developmental
regulation and modulation by external sucrose and Ca2+ ions.
Plant Mol Biol 1998, 37:265-274.
Phaseolin (seed storage globulin) promoter-GUS mRNA and GUS activity in
transgenic tobacco seeds can be induced by ABA, a process superimposed
on development-dependent regulation. The ABA response itself is repressed
by exogenously added sucrose alone, but stimulated by the presence of both
sucrose and CaCl2. These data add to earlier published work [49] on the
involvement of Ca2+ in the sugar-inducible expression of plant genes.
41. Perata P, Matsukura C, Vernieri P, Yamaguchi J: Sugar repression of
•
a gibberellin-dependent signaling pathway in barley embryos.
Plant Cell 1997, 9:2197-2708
This study deals mainly with α-amylase gene expression in germinating barley grain, but is also certainly of relevance for processes in seed maturation,
as it clearly demonstrates the complex and tissue-dependent interaction of
sugar signals with hormone regulation.
42. Chan M-T, Yu S-M: The 3¢ untranslated region of a rice a-amylase
•
gene functions as a sugar-dependent mRNA stability
determinant. Proc Natl Acad Sci USA 1998, 95:6543-6547.
See [43•] for an annotation to this reference.
43. Chan M-T, Yu S-M: The 3¢untranslated region of a rice a-amylase
•
gene mediates sugar-dependent abundance of mRNA. Plant J
1998, 15:685-695.
Evidence is provided [42•,43•] that the studied α-Amy3 3′UTR is probably
the major determinant for controlling transcript abundance in response to
glucose deprivation. This is the first example of a structural element in a
developmentally regulated plant mRNA gene involved in sugar regulation.
44. Harrington GN, Franceschi VR, Offler CE, Patrick JW, Tegeder M,
Frommer WB, Harper JF, Hitz WD: Cell specific expression of three
genes involved in plasma membrane sucrose transport in
developing Vicia faba seed. Protoplasma 1997, 197:160-173.
45. Offler CE, Liet E, Sutton EG: Transfer cell induction in cotyledons
•
of V. faba. Protoplasma 1997, 200:51-64.
The authors convincingly show that sugar levels trigger the differentiation of
epithelial cells into transfer cells. This paper together with [38•] probably
provides the first evidence for the initiation of differentiation within a developing organ in response solely to a sugar signal.
38
Growth and development
46. Hirner B, Fischer WN, Rentsch D, Kwart M, Frommer WB:
•
Developmental control of H+/amino acid permease gene
expression during seed development of Arabidopsis. Plant J
1998, 14:535-544.
The two characterised genes, AAP1 and AAP2, show non-overlapping expression patterns and might play a role in supplying the seeds with organic nitrogen.
47. Kelly MO, Spanswick RM: Maternal, single-gene regulation of
•
assimilate partitioning in pea. Plant Physiol 1997, 114:1055-1059.
The paper stresses the point that important seed characteristics like rate and
duration of seed growth are strongly influenced by physiological processes
determined maternally.
48. Swain SM, Reid JB, Kamiya Y: Gibberellins are required for embryo
•
growth and seed development in pea. Plant J 1997, 12:1329-1338.
The pea mutants lh-1 and lh-2 provide the most unequivocal evidence for an
important role of gibberellins (GAs) in early seed development. They exclude
assimilate availability and lacking maternal GAs as causes for reduced final
seed weight or seed abortion, but suggest that GAs either promote assimilate uptake by the embryo or exert a direct influence on seed growth.
49. Ohto M, Hayashi K, Isobe M, Nakumura K: Involvement of Ca2+
signalling in the sugar-inducible expression of genes coding for
sporamin and b-amylase of sweet potato. Plant J 1995,
7:297-307.
Seed maturation: genetic programmes and control signals
Ulrich Wobus* and Hans Weber
Seed maturation is mainly governed by a few genes best
studied in maize and Arabidopsis. The isolation of the LEC1
and FUS3 genes, besides the previously known VP1/ABI3
genes, and their identification as transcriptional regulators
provides the first direct hints as to their molecular mode of
action. With the identification of new effector genes, the
investigation of the role of hormones with new methods such
as immunomodulation and the increasingly recognised role
of metabolites like sugars as important modulators of seed
development, we increasingly understand the complexity
and structure of the regulatory network underlying
seed maturation.
Addresses
Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK),
D-06466 Gatersleben, Germany
*e-mail: [email protected]
Current Opinion in Plant Biology 1999, 2:33–38
http://biomednet.com/elecref/1369526600200033
© Elsevier Science Ltd ISSN 1369-5266
Abbreviation
ABA
abscisic acid
Introduction
Higher plants are characterised by the formation of seeds
which contain the embryo, protected by the maternallyderived seed coat. The embryo first runs through a phase
of cell division and morphogenesis, followed by a maturation phase which includes the accumulation of storage
products, the suppression of precocious germination, the
acquisition of desiccation tolerance, water loss and often
the induction of dormancy (see [1•]). Maturation interrupts seedling development. The process progresses in a
wave-like manner forming temporally and spatially
determined patterns or gradients between and within
the different seed organs [2,3]. Several lines of evidence
strongly suggest that maturation evolved late in evolution and has been ‘inserted’ [1 •] at different
developmental time points in different higher plant taxa
[4•]. In depth molecular analyses, however, are mainly
carried out in Arabidopsis, grain legumes and cereals
(especially maize), in each case with a specific research
focus depending on the different advantages of the three
experimental systems. In this review, the following
exciting recent research achievements are briefly discussed; the isolation and initial functional analysis of
central transcriptional regulators of seed maturation and
some of their target genes, a new methodology to control
the process at the hormonal level and the role of metabolites as triggers of maturation processes. These data
necessarily cover only certain aspects of the complex
network of processes involved in seed maturation.
ABA3, FUS3 and LEC1: three major regulators
of seed maturation
Analyses of mutant phenotypes in Arabidopsis have provided
strong evidence that three genes — ABA-INSENSITIVE3
(ABI3), FUSCA3 (FUS3) and LEAFY COTYLEDON1
(LEC1) — are of utmost importance for seed maturation as
they affect a wide, broadly overlapping but not identical
spectrum of seed-specific characters (see Figure 9 in [5••]
for a good graphic representation). In essence, mutant
embryos enter the germination programme immediately
by skipping maturation. The description of the abi3
mutant as abscisic acid insensitive already indicates a link
to the most important hormonal regulator of seed maturation, abscisic acid (ABA). Until recently, only the molecular
structure of the ABI3 gene [6] and its maize homologue,
the transcriptional activator gene VP1 [7], were known. In
1998, however, the sequences of two other genes, FUS3
and LEC1, have been obtained [8••,9••] and led to the
important conclusions discussed below.
The FUS3 gene turned out to be related to the VP1/ABI3
gene family, but is of reduced length (ABI3 encodes 720
amino acid residues; FUS3 only 312 residues), and high
homology in the protein is restricted to a stretch of at least
100 amino acid residues [9••] defined as the B3 region in
the VP1 protein [10]. The B3 domain can mediate
sequence-specific DNA-binding in vitro [11•] and is critical
for gene activation at low or insignificant ABA concentrations [12•]. It was also used to define a B3-box family of
genes (or B3-domain family of proteins) [9••] of which
FUS3 clearly is a member of a — possibly more ancient —
subgroup. An auxin-response factor ARF1 [13] represents
another even more remote subfamily. In accordance with
the ABA independence of fus3 mutants, the FUS3 protein
lacks an amino-terminal A-domain which, in VP1 (and certainly in ABI3), mediates the ABA response [12•]. FUS3 is
often classified as an LEC1-type gene on the basis of
mutant phenotypes (see [1•,8••,14]), but the two genes are
structurally unrelated and should, therefore, stay separate.
LEC1 has recently been shown most probably to encode a
protein related to a transcription factor subunit of the HAP3
type [8••] — HAP3 in mammals is part of the heterotrimeric CCAAT box-binding factor CBF (see [15]).
On the basis of an extensive descriptive study of double
and the parental single mutants, Parcy et al. [5••] had
already concluded that the three genes act synergistically. The authors proposed a model for the concerted
action of the gene trio in controlling several major
aspects of seed maturation. They further speculated that
the three proteins could act as transcription factors and
form hetero-oligomeric combinations, which regulate
different developmental processes. These protein complexes could also feed-back regulate their own gene
34
Growth and development
promoters as shown, for instance, for the DEF A/GLO
heterodimer in Antirrhinum [16]. DEF A and GLO are
two MADS-box transcription factors involved in flower
development. Preliminary experiments at the molecular
level involving ABI3 and FUS3 [17•] provide first hints
that these suggestions may be valid. In a transient
expression assay both FUS3 and ABI3 strongly induce
the activity of maturation-specific promoters in a cooperative and synergistic way. This induction was strictly
dependent on an intact RY (CATGCAT) promoter-ciselement and suggests a rather direct mode of action,
perhaps direct binding. Also LEC1, due to its proposed
HAP3-like function [8••], could directly bind to the RY
repeat. In maize and other grasses, the same element
(here called the Sph motif) is functionally dependent on
the VP1 gene product, specifically on the B3 domain, in
promoters of genes like C1, a transcriptional regulator of
anthocyanin biosynthesis in seeds [11•]. VP1 not only
recognises the Sph cis-element [11•], but may also act by
its B2 domain as a protein association enhancer [10]. In
an Em-promoter G-box binding complex, VP1 is associated with a 14-3-3 protein (GF14), and VP1, the G-box
transcription factor EmBP1 and GF14 may all interact to
stabilise and/or activate the transcription complex
[18•,19•]. Em is an ABA-responsive late embryogenesis
abundant (LEA) gene expressed exclusively in maturing
embryos. Additional b-ZIP factors, modifying ABI3
homologue-dependent gene expression, have been characterised from pea [20,21] and underscore the complex
nature of the molecular machinery directing gene regulation during seed maturation.
Studies on genes affected by FUS3 [22•,23•] revealed
another interesting new aspect; among the genes controlled by FUS3 during maturation are upregulated [22•]
and downregulated [23 •] MYB transcription factors.
AtMYB13, for instance, is involved in some aspects of
meristematic vegetative growth. It is likely to be a representative of several regulators of vegetative growth
repressed during maturation by the concerted action of
at least FUS3 and LEC1 [23•]. The data also indicate that
FUS3 is, on the one hand, a central regulator controlling
other regulators such as MYB transcription factors but,
on the other hand, the protein probably directly regulates effector genes like maturation-specific storage
protein genes via the RY promoter cis-element.
The ABI3/VP1 transcriptional activator family is well
known from mutant phenotypes to be involved in establishing embryo dormancy. Recent work with Avena fatua
inbred lines suggests that VP1 plays a central role since
AfVP1 mRNA levels established during the final stages of
embryo maturation were positively correlated with the dormant phenotype in a way that qualifies the gene as a
molecular marker for dormancy potential/after-ripening
time [24•]. The most recent discovery of a role of an ABI3
homologue in quiescent apices of Arabidopsis [25•] signals
that the function of genes of the ABI3-type, and perhaps
also of the FUS3-type (low expression levels in non-seed
tissues [9••]), is not restricted to seeds. Interestingly,
another newly isolated transcriptional regulator, ABI4,
most likely to be involved in regulating seed responses to
ABA, is not expressed seed-specifically in spite of the fact
that a clear mutant phenotype is only seen in seeds [26•].
According to published data [8••], only LEC1 expression
seems to be strictly embryo-specific but studies with more
sensitive assays like RT-PCR have not yet been reported.
In summary, three major genes involved in seed maturation initiation and maintenance are structurally either not
(LEC1) or only distantly (ABI3/FUS3) related and their
expression patterns are temporally and spatially different,
but broadly overlapping especially during seed maturation.
At least ABI3 and FUS3 are expressed in tissues other than
seed, that is to say they are not strictly seed-specific, contrary to accepted beliefs. The door is now open to study
the molecular interactions of the gene products with each
other and other factors and their role in determining cell
fate and function in each specific process.
A fresh look at the role of abscisic acid:
probing abscisic acid function by expressing
ABA-antibodies
The important role of ABA in seed maturation is well documented but since the hormone triggers diverse processes
not only in the seed, analysis by established procedures
faces limitations. A method which blocks the ABA molecule itself very specifically in a tissue-, cell- and
compartment-specific manner [27•], therefore, promises
new insights. Phillips et al. [28••] directed an ABA-specific
single chain Fv (scFv) antibody into the lumen of the endoplasmic reticulum (ER) of developing tobacco seeds by
using the ER-target signal KDEL and a seed-specific promoter, active from the late phase of early development to
the first part of late embryogenesis. The anti-ABA scFv
caused a developmental switch to the germination programme characterised by the formation of chloroplasts
containing photosynthetic pigments and a dramatic reduction in the embryo of the most abundant storage proteins as
well as protein and oil bodies. The authors calculated that
the recombinant antibody bound nearly all available ABA
until day 20 of tobacco development. The overall concentration of antibody-bound ABA plus unbound ABA
increased considerably above wild-type level, possibly due
to a feed-back mechanism or protection from ABA metabolism. The seed phenotype was most similar to the
Arabidopsis aba/abi3 double mutant seeds [29]. The well
established fact that ABA biosynthetic mutants do not
markedly reduce storage protein mRNAs [29] led to the
question — how important is ABA in controlling storage
protein accumulation [30•]? The anti-ABA seeds now clearly underscore the dominating role of ABA in the process at
least in tobacco seeds [28••]. By using a broader range of
promoters and scFvs against other regulatory molecules,
immunomodulation promises to contribute considerably to
a deepened understanding of seed development.
Seed maturation: genetic programmes and control signals Wobus and Weber
Development and metabolism: renewed
interest in a long known connection
Metabolites as developmental signals in seeds
Metabolites as signal molecules in plant developmental
processes have been mainly neglected until recently when
sugars especially were shown to regulate diverse gene activities [31] and to act as important components in signal
transduction networks [32•]. In grain legume seeds, extensive, mainly correlative analyses (see [33•,34•] for reviews)
including transgenic plants which express a yeast-derived
invertase in developing seeds [35], suggested that soluble
sugars provide signals for cotyledon development but that
glucose and sucrose can play different roles; whereas a high
glucose/sucrose ratio was correlated with cell division, a high
sucrose/hexose ratio seems to be a major trigger of the storage product pathways. Of special interest is the discovery of
steep glucose gradients across the cotyledons of the grain
legume Vicia faba, which change during development and
correlate positively with mitotic activity but negatively with
storage starch accumulation [36•]. These gradients are likely
to be primarily established by the specific expression pattern
of a cell wall-bound invertase in the developing maternal
seed coat [37] and further modulated (enhanced?) by a hexose- and a sucrose transporter (mainly excluding each other
in the timing of expression) in the epidermal cells of the
embryo [38•]. Gradients were also visualised in developing
maize kernels at the level of the activity of a key enzyme —
sucrose synthase — of starch biosynthesis in sink organs
[39•]. The combination of methods which allow the (semi-)
quantitative determination not only of RNA and protein but
also of enzyme activity and metabolites at (or nearly at) cellular resolution will certainly reveal a refined net of
relationships. Whether sugar levels in such gradients function as morphogenic substances [36•], however, has not been
shown unequivocally. Whatever the exact role of sugars is,
the elicited signals have to interact with other signals to
evoke the complex processes of seed maturation.
One site of integration for different signals are promoter
sequences. Interestingly, it has recently been demonstrated
by analysing the phaseolin seed storage protein promoter in
transgenic tobacco seeds that the essential signal for the transcriptional activity of the phaseolin promoter provided by
ABA is modulated beside unknown developmental factors
by sucrose and calcium ions [40•]. In the barley embryo, glucose acting as a signal molecule interferes with the
gibberellic acid response of α-amylase gene expression, but
in a tissue-specific manner [41•]; the induction of α-amylase
by GA in the aleurone layer is unaffected by sugars, whereas
in the embryo, sugars repress the response. The site of action
of the sugar molecule has not been determined, but could be
at the transcriptional and also at the post-transcriptional
level, as rice α-amylase mRNA stability and abundance have
been shown to be controlled by sucrose [42•,43•].
Assimilate import and seed filling
If metabolites are able to act as signals in developmental
processes, the regulated import of photoassimilates into the
35
growing seed will influence seed development beside by
providing nutrients. Developing seeds import sucrose and
amino acids from the phloem. These assimilates are
unloaded from the seed coat and must be taken up by the
apoplastically isolated embryo or endosperm. Seed-located
apoplastic transport processes, therefore, can influence
assimilate partitioning and storage product synthesis. One
obvious regulatory control point could be provided by various transporters situated in the membranes of seed coat
and/or embryo epidermal cells. In Vicia faba a sucrose transporter gene is highly expressed in the epidermal transfer cell
layer covering the outer surface of developing cotyledons
[38•,44]. These transfer cells develop at the contact area of
the cotyledonary epidermis to the seed coat, starting first at
early cotyledon stage and subsequently spreading to the
abaxial region. The tissue underlying the epidermal transfer
cell layer differentiates into storage parenchyma cells, indicating an important role of the sucrose transporter in
providing sugars for storage product synthesis. Feeding high
concentrations of sugars in vitro suppressed both the sucrose
transporter gene expression and transfer cell differentiation,
suggesting a control mechanism by carbohydrate availability
[38•]. A more detailed study [45•] also demonstrated the
induction of transfer cell differentiation by sugars and the
modulation by the sugar state of sugar transport activity and
consequently storage product synthesis. Less detailed data
are available for amino acid transporters which have a broad
substrate specificity and are probably regulated by tissuespecific expression. Two members were found to be
expressed in developing seeds of Arabidopsis. Using promoter-GUS fusions it was shown that AAP1 is expressed in the
endosperm and the cotyledons whereas AAP2 is expressed in
the vascular strands of siliques and in funiculi [46•].
In addition to these seed-specific processes, there are
genes acting on the whole plant level in controlling
assimilate transport into the seed and seed growth. One
example is the SN gene in pea which regulates assimilate
partitioning between vegetative and reproductive
growth [47•]. One important factor could be gibberellic
acid (GA) since it appears that in the pea lh2 mutation
seeds are unable to use available assimilates. In developing lh2-seeds, GA levels are reduced and lead to reduced
growth and to a high frequency of seed abortion. GAs
may either promote assimilate uptake into developing
seeds or affect seed growth, that is sink strength [48•].
The general significance of seed-specific transport
processes for development, however, is still unknown.
Conclusions
Substantial progress has been made in the past two years
in elucidating the structure and the mode of action of
genes involved in embryo development and seed maturation. This progress will now gain much momentum,
especially by the Arabidopsis genome sequencing project.
Furthermore, some signal transduction chains with their
components and branch points are slowly emerging and
mostly correlative analyses suggest a number of mainly
36
Growth and development
hitherto neglected factors modulating or even triggering
developmental processes. The still scarce data suggest the
existence of an extended regulatory network rather than a
clear hierarchy of actions and allows first glimpses on its
structure. Large scale expression analysis by DNA chip
technology will uncover many more possible network components especially at the gene level. The real challenge,
however, will be to unravel their role and interconnection.
Beside the genetic approach, with its focus on gene structure and mRNA expression level, more extensive studies
at the protein and higher order structural level (subcellular,
cellular and also whole-plant level), together with more
integrated experimental approaches are necessary to further deepen and extend our knowledge of seed maturation
as part of seed development and plant life in general.
Acknowledgements
The authors would like to thank Helmut Bäumlein and Udo Conrad for
help with the manuscript and Simon Miséra and Antje Rhode for providing
manuscripts in press. Work in our lab is supported by the Deutsche
Forschungsgemeinschaft and the European Union.
References and recommended reading
Papers of particular interest, published within the annual period of review,
have been highlighted as:
• of special interest
•• of outstanding interest
Harada JJ: Seed maturation and control of germination. In Cellular
and Molecular Biology of Plant Seed Development. Edited by Larkins
BA, Vasil IK. Dordrecht: Kluwer Academic Publishers; 1997:545-592.
Extensive and recommended review covering all aspects of seed maturation.
1.
•
2.
Perez-Grau L, Goldberg RB: Soybean seed protein genes are
regulated spatially during embryogenesis. Plant Cell 1989,
1:1095-1109.
3.
Borisjuk L, Weber H, Panitz R, Manteuffel R, Wobus U:
Embryogenesis in Vicia faba L.: histodifferentiation in relation to
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147:203-218.
Kaplan DR, Cooke TJ: Fundamental concepts in the
embryogenesis of dicotyledons: a morphological interpretation of
embryo mutants. Plant Cell 1997, 9:1903-1919.
This paper is well worth reading. It represents, in essence, a minority view by
challenging the concept of pattern formation in plants, but also brings to
mind the comparative morphological approach to embryogenesis often
neglected by molecular biologists.
4.
•
Parcy F, Valon C, Kohara A, Miséra S, Giraudat J: The ABSCISIC
ACID-INSENSITIVE3, FUSCA3, and LEAFY COTYLEDON1 loci act
in control multiple aspects of Arabidopsis seed development.
Plant Cell 1997, 9:1265-1277.
In this important paper, the authors, by quantifying several developmental
processes in a range of mutants, clearly show that the three genes work synergistically and not in different pathways as earlier models suggested. They
also provide evidence that the abundance of ABI3 is regulated by FUS3 and
LEC1. The genetic model provided can now be tested at the molecular level
due to the availability of FUS3 and LEC1.
5.
••
Homology of LEC1 to HAP3 subunit sequences of the CCAAT-box binding factor CBF is restricted to the HAP3 B-domain.
Luerßen H, Kirik V, Herrmann P, Miséra S: FUSCA3 encodes a
protein with a conserved VP1/ABI3-like B3 domain which is of
functional importance for the regulation of seed maturation in
Arabidopsis thaliana. Plant J 1998, 15:755-764.
A third key regulatory gene of seed maturation beside ABI3 and LEC1,
FUS3, has been isolated by postional cloning and its product shown to
share the B3 domain with ABI3-like proteins, whereas significant features of
A- and B1-domains are missing, obviously due to the short amino-terminal
part of this protein. Whether some similarities to B2-domain sequences are
of functional importance has still to be shown. The analysed allelic variants
(most of them causing abnormal splicing) provide hints for the essential role
of the B3 domain in seed maturation (but see [12•]).
9.
••
10. Hill A, Nantel A, Rock CD, Quatrano RS: A conserved domain of the
VIVIPAROUS-1 gene product enhances the DNA binding activity
of the bZIP protein EmBP-1 and other transcription factors. J Biol
Chem 1996, 271:3366-3374.
11. Suzuki M, Kao CY, McCarty DR: The conserved B3 domain of
•
VIVIPAROUS1 has a cooperative DNA binding activity. Plant Cell
1997, 9:799-807.
Since its isolation, VP1 has been implicated in gene regulation, but the question remained open as to whether or not the protein binds directly to DNA.
Now the McCarty lab provides convincing evidence that the B3 domain of
VP1 has a highly co-operative, sequence-specific DNA-binding activity in an
in vitro assay. Specific binding to the Sph-motif TCCATGCAT provides a
good link to the expected functions and leads to the suggestion that the B3
domain in general binds to this element (also known as the RY-motif) present
in many seed protein gene promoters.
12. Carson CB, Hattori T, Rosenkrans L, Vasil V, Vasil IK, Peterson P,
•
McCarty DE: The quiescent/colourless alleles of viviparous-1 show
that the conserved B3 domain of VP1 is not essential for
ABA-regulated gene expression in the seed. Plant J 1997,
12:1231-1240.
By analysing several vp1 alleles in a further attempt to map functional
domains within the VP1 protein in more detail, the authors conclude that B3
is not absolutely essential for completion of seed maturation. Contrary to
predictions in [9••], a two amino acid deletion in the B3 domain of FUS3 still
led to the Fus3– phenotype in Arabidopsis. Whether this apparent discrepancy is of general importance has still to be worked out.
13. Ulmasov T, Hagen G, Guilfoyle TJ: ARF1, a transcription factor that
binds to auxin response elements. Science 1997, 276:1865-1868.
14. Vernon DM, Meinke DW: Late embryo-defective mutants of
Arabidopsis. Dev Genet 1995, 16:311-320.
15. Maity SN, de Crombrugghe B: Role of the CCAAT-binding protein
CBF/NF-Y in transcription. Trends Biochem Sci 1998, 23:174-178.
16. Schwarz-Sommer Z, Hue I, Huijser P, Flor PJ, Hansen R, Tetens F,
Lönnig W-E, Saedler H, Sommer H: Characterization of the
Antirrhinum floral homeotic MADS-box gene deficiens:
evidence for DNA binding and autoregulation of its persistent
expression throughout flower development. EMBO J 1992,
11:251-263.
17.
•
Wohlfarth T, Braun H, Kirik V, Kölle K, Czihal A, Tewes A, Luerssen H,
Miséra S, Shutov A, Bäumlein H: Regulation and evolution of seed
globulin genes. J Plant Physiol 1998, 152:600-606.
This symposium paper contains interesting data which have still to be published in detail. They suggest a co-operative interaction of FUS3 and ABI3
in the regulation of seed-specific promoters with the RY cis-motif as target.
6.
Giraudat J, Hauge BM, Valon C, Smalle J, Parcy F, Goodman HM:
Isolation of the Arabidopsis ABI3 gene by positional cloning. Plant
Cell 1992, 4:1251-1261.
18. Schultz TF, Medina J, Hill A, Quatrano RS: 14-3-3 proteins are part
•
of an abscisic acid-VIVIPAROUS1 (VP1) response complex in the
Em promoter and interact with VP1 and EmbP1. Plant Cell 1998,
10:837-847.
The paper provides a first and important example of a transcription complex
involving VP1 and identifies members of the complex including 14-3-3 proteins which are supposed to act as mediators in protein–protein interactions
involved in kinase-related steps of signal-transduction pathways.
7.
McCarty DR, Carson CB, Stinad PS, Robertson DS: Molecular
analysis of viviparous-1: an abscisic acid-insensitive mutant
maize. Plant Cell 1989, 1:523-532.
19. Quatrano RS, Bartels D, Ho T-HD, Pagés M: New insights into
•
ABA-mediated processes. Plant Cell 1997, 9:470-475.
A meeting-based review including the role of ABA in seed maturation.
Lotan T, Ohto M-A, Yee KM, West MAL, Lo R, Kwong RW, Yamagishi
K, Fischer RL, Goldberg RB, Harada JJ: Arabidopsis LEAFY
COTYLEDON1 is sufficient to induce embryo development in
vegetative cell. Cell 1998, 93:1195-1205.
The authors isolated the LEC1 gene and demonstrated its major role, not
only during late, but also in early embryogenesis including suspensor and
endosperm development. Ectopic LEC1 expression, in contrast to ABI3,
induced embryonic programmes in vegetative cells in rare cases.
20. Chern MS, Bob AJ. Bustos MM: The regulator of MAT2 (ROM2)
protein binds to early maturation promoters and represses
PvALF-activated transcription. Plant Cell 1996, 8:305-321.
8.
••
21. Chern MS, Eiben HG, Bustos MM: The developmentally regulated
bZIP factor ROM1 modulates transcription from lectin and
storage protein genes in bean embryos. Plant J 1996,
10:135-148.
Seed maturation: genetic programmes and control signals Wobus and Weber
22. Kirik V, Kölle K, Miséra S, Bäumlein H: Two novel MYB homologues
•
with changed expression in late embryogenesis-defective
Arabidopsis mutants. Plant Mol Biol 1998, 37:819-827.
See [23•] for annotation to this reference.
23. Kirik V, Kölle K, Wohlfarth T, Miséra S, Bäumlein H: Ectopic
•
expression of a novel MYB gene modifies the architecture of the
Arabidopsis inflorescence. Plant J 1998, 13:729-742.
Papers [22•] and [23•] contribute to the elucidation of the key role of the
ABI3/LEC1/FUS3 gene trio in late embryogenesis by the identification,
via differential display, of effector genes — in this case members of the
MYB group of transcriptional regulators. The genes described are either
positively [22•] or negatively [23•] regulated. AtMYB13 is activated in late
embryogenesis defective fus3 and lec1 mutants but not abi3 mutants,
that is to say, in wild-type embryos this MYB protein should be at least in
part responsible for repressing genes involved in vegetative meristem
activation. The report [23•] provides the first evidence at the molecular
level for different action spectra of ABI3 on the one hand and FUS3 plus
LEC1 on the other.
24. Jones HD, Peters NCB, Holdsworth MJ: Genotype and environment
•
interact to control dormancy and differential expression of the
VIVIPAROUS-1 homologue in embryos of Avena fatua. Plant J
1997, 12:911-920.
This is yet another good example that the well-chosen use of plants other
than Arabidopsis can considerably contribute to the better understanding of
specific developmental processes.
25. Rhode A, Van Montagu M, Boerjan W: The abscisic acid-insensitive
•
gene ABI3 is expressed during vegetative quiescence processes
in Arabidopsis. Plant Cell Environ 1998, in press.
Here, Rhode et al. show, in contrast to present conceptions, that ABI3 most
probably has additional functions to those in seed development. During
growth in the dark ABI3 is expressed in the seedling apex after the arrest of
cell division. Concomitantly, the 2S napin seed storage protein is induced.
ABI3-promoter/GUS fusion constructs are expressed in still other vegetative organs. These results — together with the observed expression of an
ABI3 homologue in dormant poplar buds (A Rhode, Molecular strategies
to study bud dormancy in Populus [PhD thesis]. Universiteit Gent
1998) — led the authors to conclude that ABI3 is one of several components
of a molecular network underlying all types of quiescence.
26. Finkelstein RR, Wang ML, Lynch TJ, Rao S, Goodman H: The
•
Arabidopsis abscisic acid response locus ABI4 encodes an
APETALA2 domain protein. Plant Cell 1998, 10:1043-1054.
ABA-sensitive abi4 mutants are defective in seed development but exhibit a
less severe phenotype than known abi3 mutants. The gene, isolated by positional cloning, might represent a new element in the signal transduction
pathway involving ABI3.
Conrad U, Fiedler U: Compartment-specific accumulation of
recombinant immunoglobulins in plant cells: an essential tool for
antibody production and immunomodulation of physiological
functions and pathogen activity. Plant Mol Biol 1998, 38:101-109.
A review focusing on the importance of cell compartment-specific expression
of antibodies with respect to both functional studies or biotechnological goals.
27.
•
28. Phillips J, Artsaenko O, Fiedler U, Horstmann C, Mock H-P, Müntz K,
•• Conrad U: Seed-specific immunomodulation of abscisic acid
activity induces a developmental switch. EMBO J 1997,
16:4489-4496.
The paper demonstrates for the first time the successful use of transgene
encoded antibodies directed against low molecular weight regulatory compounds for the study of plant developmental processes. This new experimental tool will be certainly used in the future in a variety of situations
including other hormones and non-protein regulators.
29. Koornneef M, Hanhart CJ, Hilhorst HWM, Karssen CM: In vivo
inhibition of seed development and reserve protein accumulation
in recombinants of abscisic acid biosynthesis and
responsiveness mutants in Arabidopsis thaliana. Plant Physiol
1989, 90:463-469.
30. Merlot S, Giraudat J: Genetic analysis of abscisic acid signal
•
transduction. Plant Physiol 1997, 114:751-757.
One of several recommendable recent reviews (see also [19•]) on ABA signal transduction.
31. Koch KE: Carbohydrate-modulated gene expression in plants.
Annu Rev Plant Physiol Plant Mol Biol 1996, 47:509-540.
32. Jang J-C, Sheen J: Sugar sensing in higher plants. Trends Plant Sci
•
1997, 2:208-214.
Recommended review on a rapidly expanding research field.
33. Weber H, Borisjuk L, Wobus U: Sugar import and metabolism
•
during seed development. Trends Plant Sci 1997, 2:169-174.
See [34•] for annotation for this reference.
37
34. Weber H, Heim U, Golombek S, Borisjuk L, Wobus U: Assimilate
•
uptake and the regulation of seed development. Seed Sci Res
1998, 8:331-345.
Reviews (condensed [33•] and more extended [34•]) focusing on the possible role of sugars in regulating cotyledon development.
35. Weber H, Heim U, Golombek S, Borisjuk L, Manteuffel R, Wobus U:
Expression of a yeast-derived invertase in developing cotyledons
of Vicia narbonensis alters the carbohydrate state and affects
storage functions. Plant J 1998, 16:163-172.
36. Borisjuk L, Walenta S, Weber H, Müller-Klieser W, Wobus U: High
•
resolution histographical mapping of glucose concentrations in
developing cotyledons of Vicia faba in relation to mitotic activity
and storage processes: glucose as a possible developmental
trigger. Plant J 1998, 15:583-591.
A luciferase-coupled enzyme reaction together with quantitative bioluminescence and single photon imaging is used to visualise glucose concentration
gradients which are related to specific cell types. In addition, they correlate
positively with mitotic activity and negatively with starch accumulation. The
authors speculate about glucose concentration ‘landscapes’ as possible
morphogenetic gradients.
37.
Weber H, Borisjuk L, Heim U, Buchner P, Wobus U:
Seed coat-associated invertases of fava bean control both
unloading and storage functions: cloning of cDNAs and cell-type
specific expression. Plant Cell 1995, 7:1835-1846.
38. Weber H, Borisjuk L, Heim U, Sauer N, Wobus U: A role for sugar
•
transporters during seed development: molecular
characterization of a hexose and a sucrose carrier in Fava bean
seeds. Plant Cell 1997, 9:895-908.
The authors back their hypothesis on the important role of sugars in cotyledon development by describing correlations between the expression of
sugar transporters, sugar levels and cell type differentiation. Additional
experimental evidence is drawn from work with explanted cotyledons.
39. Wittich PE, Vreugdenhil D: Localization of sucrose synthase activity
•
in developing maize kernels by in situ enzyme histochemistry.
J Exp Bot 1998, 49:1163-1171.
A much welcomed piece of work bringing enzyme activity studies in seeds
down to the cellular level. Such studies, together with high-resolution
metabolite analysis (see [36•]) and gene expression studies, will increasingly contribute to a better understanding of developmental processes.
40. Bustos MM, Iyer M, Gagliardi SJ: Induction of a ß-phaseolin
•
promoter by exogenous abscisic acid in tobacco: developmental
regulation and modulation by external sucrose and Ca2+ ions.
Plant Mol Biol 1998, 37:265-274.
Phaseolin (seed storage globulin) promoter-GUS mRNA and GUS activity in
transgenic tobacco seeds can be induced by ABA, a process superimposed
on development-dependent regulation. The ABA response itself is repressed
by exogenously added sucrose alone, but stimulated by the presence of both
sucrose and CaCl2. These data add to earlier published work [49] on the
involvement of Ca2+ in the sugar-inducible expression of plant genes.
41. Perata P, Matsukura C, Vernieri P, Yamaguchi J: Sugar repression of
•
a gibberellin-dependent signaling pathway in barley embryos.
Plant Cell 1997, 9:2197-2708
This study deals mainly with α-amylase gene expression in germinating barley grain, but is also certainly of relevance for processes in seed maturation,
as it clearly demonstrates the complex and tissue-dependent interaction of
sugar signals with hormone regulation.
42. Chan M-T, Yu S-M: The 3¢ untranslated region of a rice a-amylase
•
gene functions as a sugar-dependent mRNA stability
determinant. Proc Natl Acad Sci USA 1998, 95:6543-6547.
See [43•] for an annotation to this reference.
43. Chan M-T, Yu S-M: The 3¢untranslated region of a rice a-amylase
•
gene mediates sugar-dependent abundance of mRNA. Plant J
1998, 15:685-695.
Evidence is provided [42•,43•] that the studied α-Amy3 3′UTR is probably
the major determinant for controlling transcript abundance in response to
glucose deprivation. This is the first example of a structural element in a
developmentally regulated plant mRNA gene involved in sugar regulation.
44. Harrington GN, Franceschi VR, Offler CE, Patrick JW, Tegeder M,
Frommer WB, Harper JF, Hitz WD: Cell specific expression of three
genes involved in plasma membrane sucrose transport in
developing Vicia faba seed. Protoplasma 1997, 197:160-173.
45. Offler CE, Liet E, Sutton EG: Transfer cell induction in cotyledons
•
of V. faba. Protoplasma 1997, 200:51-64.
The authors convincingly show that sugar levels trigger the differentiation of
epithelial cells into transfer cells. This paper together with [38•] probably
provides the first evidence for the initiation of differentiation within a developing organ in response solely to a sugar signal.
38
Growth and development
46. Hirner B, Fischer WN, Rentsch D, Kwart M, Frommer WB:
•
Developmental control of H+/amino acid permease gene
expression during seed development of Arabidopsis. Plant J
1998, 14:535-544.
The two characterised genes, AAP1 and AAP2, show non-overlapping expression patterns and might play a role in supplying the seeds with organic nitrogen.
47. Kelly MO, Spanswick RM: Maternal, single-gene regulation of
•
assimilate partitioning in pea. Plant Physiol 1997, 114:1055-1059.
The paper stresses the point that important seed characteristics like rate and
duration of seed growth are strongly influenced by physiological processes
determined maternally.
48. Swain SM, Reid JB, Kamiya Y: Gibberellins are required for embryo
•
growth and seed development in pea. Plant J 1997, 12:1329-1338.
The pea mutants lh-1 and lh-2 provide the most unequivocal evidence for an
important role of gibberellins (GAs) in early seed development. They exclude
assimilate availability and lacking maternal GAs as causes for reduced final
seed weight or seed abortion, but suggest that GAs either promote assimilate uptake by the embryo or exert a direct influence on seed growth.
49. Ohto M, Hayashi K, Isobe M, Nakumura K: Involvement of Ca2+
signalling in the sugar-inducible expression of genes coding for
sporamin and b-amylase of sweet potato. Plant J 1995,
7:297-307.