Smith and Wang, unpublished), loss of which
results in a slow-growing plant with a low starch
content. No protein of this class has previously
been reported to be necessary for starch metab-
olism. A knockout of the equivalent gene in
Arabidopsis results in slow growth, but not
reduced starch content, indicating that the influ-
ence on starch content inL. japonicusmay reflect
an indirect rather than a direct requirement for
the gene product.
Mutant analysis in pea showed that one iso-
form of sucrose synthase that is highly expressed
in the nodule is the main enzyme responsible for
delivering carbon for nitrogenfixation. Mutants
lacking this isoform derived their nitrogen lar-
gely from the soil rather than from theRhizobium
symbiosis (Craig et al. 1999 ). These mutants
retained a small amount of sucrose synthase
activity in the nodule, indicating a potential
contribution from a second isoform. Using L.
japonicus, Horst et al. ( 2007 ) showed that two
isoforms are present in the nodule,LjSUS1and
LjSUS3, and that both contribute to sucrose
catabolism. There was no nitrogenfixation in
nodules of a double mutant lacking both iso-
forms, leading to the conclusion that sucrose
synthase is vital for nodule function and that its
actions cannot be replaced by those of invertases
also present in the nodule.
Although neutral/alkaline invertase does not
seem to contribute to nodule function, mutant
analysis shows that a cytosolic isoform of this
class is essential for the cellular development and
growth of theL. japonicusplant as a whole. Loss
of expression ofLjinv1resulted in dramatically
reduced growth of root and shoot, and changes to
tissue organisation, with enlarged root meristems
and thickened roots with more cells. The change
in cell organisation was also observed in leaves,
stems and nodules, and there was a lack of pollen
(Welham et al. 2009 ). A knockout of the equiv-
alent gene inArabidopsishad only a very mild
phenotype, but a double mutant lacking two
cytosolic invertases closely related to Ljinv1
resulted in a very strong effect onArabidopsis
growth and development (Barratt et al. 2009 ).
The reasons for the importance of this class of
invertases are not yet understood, but they may
be related to the control of sugar signalling in
meristems (Pignocchi, Edwards, Welham, Wang
and Smith, unpublished).10.6 The Relationship Between
Starch Content and Growth
inL. japonicusThere is a general expectation that fast-growing
plant species maximise resource acquisition,
whereas slow-growing species or accessions
rather maximise the conservation of resources.
With regard to the metabolism of carbohydrates,
this means that faster-growing plants are expec-
ted to hold less carbohydrate in reserve as starch
that would allow them to cope with unexpected
changes in their environmental conditions. This
general expectation holds true forArabidopsis:a
study of 94Arabidopsis accessions showed a
negative correlation between starch level at the
end of the night and biomass production (Sulpice
et al. 2009 ). However, the picture forL. japoni-
cusis more complex. For thefirst 6 weeks of
growth, two ecotypes with different growth rates
accumulated similar amounts of starch during the
day and degraded about 95 % of it at night.
Starch levels then increased dramatically in both
ecotypes. At 8 weeks, only about 70 % of starch
accumulated during the day was degraded at
night, and this value dropped to 30 % or less by
10 weeks when the plants started to flower.
Importantly, the faster-growing ecotype con-
tained 80 % more starch at the end of the day
than the slower-growing ecotype at this point
(Vriet et al. 2010 ). We suggest that although
there may be a general relationship between
starch storage and growth rate during vegetative
growth, this relationship no longer holds in
perennial plants approaching flowering. Pre-
sumably, storage of carbon at this point is con-
trolled by signals that reflect a future requirement
for carbon to fuel growth of new vegetative
meristems once flowering (and perhaps sub-
sequent winter dormancy) hasfinished.
The genetic resources available inL. japoni-
cushave provided further insights into the role
and importance of starch storage for regrowth112 C. Vriet et al.