Genetic basis of the "sleeping leaves" revealed.
Genetic basis of the “sleeping leaves” revealed
Millán Cortizo and Patrick Laufs1
Institut National de la Recherche Agronomique, Unité Mixte de Recherche 1318, and AgroParisTech, Institut Jean-Pierre
Bourgin, F-78000 Versailles, France
morning glory, are open during the day or
part of the day and close at night. Based
on such observations, in 1751, the Swedish
botanist Carl von Linné suggested com-
bining several plant species in which the
flowers open or close at specific and
different times of the day to build a
“Horologium Florae” (flower clock) that
would accurately and colorfully predict
time. Such daily movements of plants are
not limited to flowers. In his book entitled
The Power of Movement in Plants, Darwin
(1880) described many examples of “sleep
movements of leaves” and provided a “List
of Genera, including species the leaves of
which sleep” (1). Among these, he noted
that the legume family “includes many
more genera with sleeping species than all
the other families put together.” He also
described a specialized organ, called a
joint, cushion, or pulvinus responsible for
such movement (1). In PNAS, Chen et al.
(2) identify the genetic determinant for the
formation of these pulvini in three legumes:
Pisum sativum (pea), Medicago truncatula
(barrel medic), and Lotus japonicus.
Contrary to intuition, plants are capable
of moving in response to environmental
stimuli. This movement is achieved
either through irreversible differential
growth or through reversible changes
in turgor. An example of differential
growth is the growth toward a light source
(phototropism) observed in the majority
of plant shoots. Tropisms are plant
movements induced by directional stimuli,
such as light or gravity. In addition, plants
can move in response to nondirectional
factors, such as humidity or contact. These
movements are called nastic responses.
Nyctinasty, the proper name for the “sleep
movements of leaves” is a well-known
example of a nastic response. In this case,
plants close up their leaves and petals in
response to the onset of darkness. Because
it is a rather fast response, it does not
involve differential growth but changes in
The pulvinus is the organ responsible
for the nyctinastic leaf movement. It is
a specialized structure located at the base
of the petiole of leaves or the petiolule of
leaflets in the case of compound leaves
(Fig. 1A). In the pulvinus (Fig. 1 B and C),
the central vascular bundles and the
supporting tissues (often, sclerenchyma)
gous to those of stomatal guard cells (4, 5).
ike most animals, plants also sleep
at night, or at least some of them.
For instance, the flowers of many
species, such as crocus, tulip, and
are surrounded by parenchyma. The outer
cells of the parenchyma, called the motor
cells, undergo water-driven volume
changes and are the ultimate effectors of
movement. Motor cells are distributed
into two positionally and functionally
opposed regions: extensor and flexor.
Extensors cells are located in the upper
side of the organ, whereas flexors are
located in the lower side. During leaf
opening, leaflets move downward by the
simultaneous increase of turgor pressure
in extensor and decrease in flexor cells.
During closing, the inverse occurs, exten-
sor cells shrink, and flexor cells swell,
moving leaflets upward (3). These turgor
changes in the motor cells are caused by
ion movements followed by massive water
flux across the plasma membrane. Swelling
is caused by proton pump-driven accu-
mulation in the cytoplasm of K+and Cl−.
This increase in solute concentration low-
ers water potential inside the cell, and thus
drives the entrance of water in the cell.
Shrinking is caused by a passive leaking of
solutes that is accompanied by water loss.
It is currently accepted that the osmotic
volume changes of motor cells are analo-
There is abundant literature describing
the anatomy of the pulvinus and the
physiology and biomechanics of nyctinastic
movements in legumes (e.g., reviewed in
3). However, nothing was known about
the development of this organ, probably
because Arabidopsis lacks an equivalent
structure. The report of Chen et al. (2)
starts to fill this void through the identifi-
cation of the genetic factor that determi-
nates pulvinus formation in legumes. This
collaborative work between teams of three
different continents working on three
different legume models nicely illustrates
what can be achieved when the use of
large, well-established collections of
mutants meets the most advanced plant
molecular genetic approaches.
The whole story started more than
50 y ago, when Stig Blixt identified a pea
mutant he called petiolulatus in which fo-
liar pulvini are replaced by petiolules (6).
depicts nyctinasty in M. truncatula WT and elp1 mutant.
Leaf of M. arabica (A) shows a detail of the pulvinus (B) and the extensor cells (C). (D) Schema
Author contributions: M.C. and P.L. wrote the paper.
The authors declare no conflict of interest.
See companion article 10.1073/pnas.1204566109.
1To whom correspondence should be addressed. E-mail:
www.pnas.org/cgi/doi/10.1073/pnas.1209532109PNAS Early Edition
| 1 of 2
This mutant was later renamed apulvinic
(apu), according to another mutant
independently found in 1979 by D. M.
Harvey (7). Although pea is a very nice
model for performing genetic analyses, it
is somehow more difficult to clone genes
from this species, and this probably ex-
plains why no further progress toward this
has been made. In 2003, Kawaguchi (8)
identified from a population of chemically
mutagenized L. japonicus a mutant that
could not close its leaflets at night because
of an absence of differentiated pulvini,
which was therefore called sleepless (slp).
Again, cloning the SLP gene through fine
mapping was not an easy task. Finally,
a similar mutant called elongated petiolule1
(elp1) with pulvini replaced by longer
petiolule-like structures was described in
M. truncatula. In this mutant, leaves
remain open at night (2) (Fig. 1D). The
combination of fine genetic mapping of
the elp1 mutation with the identification of
the flanking sequences of newly generated
elp1 alleles through the insertion of a
retrotransposon allowed the identifica-
tion of the ELP1 gene. From this, the
APU and SLP genes could be identified as
ELP1 orthologs bearing mutations in the
respective pea and L. japonicus mutants.
Therefore, the work of Chen et al. (2)
reveals that the formation of the pulvinus
in legumes is likely to be regulated by
a conserved genetic network orchestrated
by the ELP1/APU/SLP1 genes. Identi-
fication of these genes also provides
a straightforward way to test whether
pulvinus formation in more distantly
related species is controlled by the same
What do the mutant phenotypes tell us
about ELP1/APU/SLP functions? In the
absence of these genes, the small, iso-
diametric, epidermal pulvinus cells with
a highly convoluted surface are replaced
by much larger and elongated petiole-like
epidermal cells (2). This change in the cell
type and expansion pattern may explain
the elongated petiolule phenotype of the
elp1/apu/slp mutants. Conversely, over-
expression of ELP1 in M. truncatula leads
to dwarf plants, with shorter petioles and
leaf rachises, which correlates with a
reduction in the size of the epidermal cells
Formation of the
pulvinus in legumes is
likely to be regulated by
a conserved genetic
by the ELP1/APU/SLP1
in the transgenic lines. Interestingly, in
these plants, the small epidermal cells of
the petiole and rachis show some convo-
lution at their surface reminiscent of
pulvinus cells, indicating that ELP1 may
be sufficient to some degree for the
acquisition of the pulvinus identity. Con-
firmation of this hypothesis awaits further
identification of molecular markers of
ELP1/APU/SLP1 codes for nuclear-
localized proteins belonging to the plant-
specific LATERAL ORGAN BOUND-
ARIES domain (LBD) transcription factor
family (2). In the past years, LBD mem-
bers have been shown to have essential
regulatory functions for the development
of plant lateral organs (9, 10). LBD pro-
teins have a characteristic N-terminal lat-
eral organ boundaries (LOB) domain that
contains a putative DNA-binding domain
consisting of four conserved cysteines in
a CX2CX6CX3C motif and a 30-aa-long
domain predicted to form a coiled coil
reminiscent of a leucine zipper that may
be involved in interaction with LBD or
other proteins. The LBD family contains
43 members in either Arabidopsis or
maize, and the LOB domain accounts for
the specificity of the members of this
family (11). Close homologs of the ELP1/
APU/SLP1 genes can been found in other
species in which no similar pulvinus are
observed, such as maize and Arabidopsis.
Some of them, like LOB in Arabidopsis,
have been implicated in the establishment
of the frontiers between meristem and
lateral organs (10). Similarly, ELP1 is
expressed very early on in the basal region
of the leaflet, the region that will later
differentiate into the pulvinus, in a fashion
reminiscent of a frontier gene (2). Over-
expression of ELP1, like overexpression of
LOB, leads to dwarf plants (10), suggest-
ing that one common function of these
genes would be to control cell expansion.
Therefore, ELP1 may share some func-
tions with related LBD genes from other
species, although retaining species- and
even organ-specific roles, as shown by the
differential response of M. truncatula
rachis and petiolule to ELP1 over-
expression (2). Further comparative anal-
ysis of the role of these genes between
different species, including the elucidation
of the downstream genetic network, will
be necessary to understand how ELP1
triggers pulvinus differentiation. By iden-
tifying a key determinant of the formation
of a specialized plant structure, the work
of Chen et al. (2) provides a unique op-
portunity to understand better the genetic
basis and evolution of the diversity observed
ACKNOWLEDGMENTS. M.C. is supported by
a postdoctoral fellowship from the Fundación
Alfonso Martín Escudero (Spain).
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