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Cytoskeleton in plant development

Benedikt Kost*, Jaideep Mathur† and Nam-Hai Chua‡
The plant cytoskeleton has crucial functions in a number of ton in the development of multicellular plant organs. This
cellular processes that are essential for cell morphogenesis, problem has recently begun to be addressed by the char-
organogenesis and development. These functions have been acterization of Arabidopsis thaliana and maize (Zea mays)
intensively investigated using single cell model systems. With morphogenetic mutants that have primary defects in
the recent characterization of plant mutants that show aberrant cytoskeletal organization.
organogenesis resulting from primary defects in cytoskeletal
organization, an integrated understanding of the importance of The analysis of cytoskeletal functions, both in cellular
the cytoskeleton for plant development has begun to emerge. processes and in organogenesis, relies upon techniques
Newly established techniques that allow the non-destructive that allow observation of the cytoskeleton in cells and
visualization of microtubules or actin filaments in living plant organs. In the past year, the observation of the cytoskele-
cells and organs will further advance this understanding. ton has been significantly facilitated by the development
of green fluorescent protein (GFP)-based markers, which
Addresses allow the non-invasive visualization of microtubules and
*† Laboratory of Plant Cell Biology, Institute of Molecular Agrobiology, actin filaments in genetically transformed, living plant
National University of Singapore, Singapore 117604 cells and tissues.
*e-mail: benedikt@ima.org.sg
† e-mail: jaideep@ima.org.sg
‡ Laboratory of Plant Molecular Biology, The Rockefeller University, Here, we present an overview of the recent work on the
1230 York Avenue, New York, NY 10021 6399, USA; role of the cytoskeleton in the development of higher
e-mail: chua@rockvax.rockefeller.edu plants with respect to the aspects listed above. We empha-
Current Opinion in Plant Biology 1999, 2:462–470 size those cytoskeletal functions that are specifically
important for morphogenesis and that are not basic
1369-5266/99/$ — see front matter © 1999 Elsevier Science Ltd. requirements for cell reproduction and survival. A summa-
All rights reserved.
ry of the current knowledge on the hormonal control and
Abbreviations the intracellular regulation of the plant cytoskeleton,
GFP green fluorescent protein which is essential for a comprehensive understanding of
PPB preprophase band
TE tracheary element
how the cytoskeleton functions in plant development, is
beyond the scope of this review. For further information
interested readers are referred to [1–3].
Introduction
Cell division, cell expansion, cell differentiation and cell- Cell division
to-cell communication are fundamentally important Plant microtubules and actin filaments are essential com-
processes in the development of multicellular organisms. ponents of the basic machineries required for nuclear
During plant morphogenesis, the orientation of cell divi- division and cytokinesis [4–6]. Plants have evolved a
sion planes and the direction of cell expansion must be unique mode of cytokinesis, the process that results in the
strictly controlled because, unlike animal cells, plant cells separation of two daughter cells following completion of
are constrained within rigid cell walls and are unable to nuclear division. A new cell wall, the cell plate, is built by
rapidly change shape or to migrate. targeted secretion between the two daughter nuclei. Cell
plate formation is initiated in the cell center and proceeds
Microtubules and actin filaments, the two key compo- centrifugally until the new cell wall fuses with the
nents of the eukaryotic cytoskeleton, play important roles parental cell wall [7]. Positioning of the cell plate is of cru-
in all of these processes (Figure 1). Single cell model sys- cial importance for plant morphogenesis and is under
tems such as tobacco BY-2 suspension cells, Tradescantia cytoskeletal control.
virginiana stamen hair cells, guard cell initials, cultured
pollen tubes, root hairs, trichomes, Zinnia elegans tracheary Dividing cells in young plant organs generally form anticli-
elements developing in vitro, microinjected mesophyll nal — perpendicular to the organ surface — cell plates.
cells and algal cells have been widely exploited to investi- After exiting the cell cycle, newly formed cells elongate
gate cytoskeletal functions in the cellular processes that perpendicularly to the cell plate, resulting in the con-
affect development. trolled, directed expansion of developing organs. During
embryogenesis and meristem development, cell plates in
Although these model systems, which allow the easy single dividing cells are positioned periclinally (parallel to
manipulation and observation of cellular events, have the surface), leading to the establishment of new cell lay-
proven extremely valuable in elucidating the cellular func- ers. Together, these processes are responsible for the
tions of microtubules and actin filaments, they can provide organization of cells in files and layers that is typically
only indirect information about the role of the cytoskele- observed in plant tissues [8].
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Cytoskeleton in plant development Kost, Mathur and Chua 463

Figure 1 guide the extending cell plate to the correct fusion site.
Local cell wall differentiations [16] and a cortical region
largely devoid of filamentous actin — zone of actin deple-
tion (ZAD) — that remains at the position of the PPB after
Organogenesis/
its disappearance [17], have been proposed to constitute
Cell-to-cell such landmarks. The observation of actin filaments
communication extending from the expanding cell plate to the cortical
fusion site [18,19], together with the demonstration of
actin assembly at the edge of growing cell plates [20•], has
Pollen tube led to the suggestion that the actin cytoskeleton may have
growth a function in cell plate guidance. This is consistent with
reports showing that treatment with actin depolymerizing
drugs results in abnormal cell plate positioning [11,21].
Cell
differentiation Cell plate positioning during asymmetric cell divisions
Trichome
morphogenesis
appears to be achieved by the same mechanisms as those
described above [5]. During such divisions, however, PPB
formation is preceded by a relocation of the nucleus
towards one cellular pole, which is mediated by filamen-
tous actin structures that connect the nucleus to the cell
Cell expansion Root hair cortex [22]. The PPB and the cell plate are established
(polarity elongation
later in a non-medial plane which is determined by the
establishment)
position of the nucleus.

Cell expansion
Cell division
Cell expansion in developing tissues
Current Opinion in Plant Biology After leaving the cell cycle, differentiating plant cells typ-
ically expand dramatically. Most cells in developing plant
The cytoskeleton has crucial functions in different cellular processes
that are essential for plant development. The experimental evidence tissues grow diffusely, with some cell extension dispersed
summarized in this review demonstrates that microtubules and/or actin over the entire surface, but still expand preferentially
filaments are required for correct cell plate positioning, for the control along one axis in a clearly polar manner [23]. The determi-
of directional cell expansion, for single cell morphogenesis (pollen tube nation of the main direction of this type of cell expansion
growth, root hair elongation, trichome morphogenesis), for cell
differentiation, for cell-to-cell communication through plasmodesmata plays a key role in organogenesis, as discussed above, and
and for organogenesis. is thought to be mediated by cytoskeletal elements.

Diffuse cell expansion is driven by turgor pressure. The

Most commonly, dividing plant cells form two identically cell wall of diffusely growing cells generally contains cross-
sized daughter cells. Some cells, however, including linked cellulose microfibrils that are arranged in parallel to
zygotes, microspores, certain cells in developing meris- one another and transversely to the direction of cell elon-
tems and guard cell initials, undergo asymmetric cell gation. Because this microfibrillar network resists radial
division, with the cell plate positioned in a non-medial expansion much more than longitudinal expansion, the
plane. This invariably results in the formation of two cells react to turgor pressure by elongating perpendicular-
daughter cells that differ in size, cytoplasmic content and ly to the microfibril orientation [24].
developmental fate [9].
Cellulose microfibrils are synthesized by a multi-subunit
Both microtubules and actin filaments appear to have enzymatic complex, the cellulose synthase, which is inte-
essential functions in cell plate positioning. At an early grated into the plasma membrane. A widely accepted
stage of cell division, cortical microtubules reorganize into model predicts that cellulose synthase moves along cortical
a ring of parallel bundles, the preprophase band (PPB) microtubules and uses them as a template for the oriented
[10]. Filamentous actin is also present in the microtubule deposition of microfibrils in the cell wall [25]. According to
PPB and may be required for the final restriction of this this hypothesis, the main axis of diffuse cell growth is ulti-
structure to a narrow ring [11,12]. The PPB predetermines mately determined by the orientation of cortical
the site where the maturing cell plate will fuse with the microtubules. This idea was initially based on microscopi-
parental cell wall [13], even though the mitotic spindle and cal observations of a co-alignment of microfibrils in the
the early cell plate are often obliquely oriented with vicinity of the plasma membrane with cortical microtubules
respect to the plane defined by this structure [14,15•]. The [26]. It was later supported by experiments showing that
PPB disappears completely when the spindle is formed the disruption of microtubules results in depolarized cell
[13], but may leave landmarks in the cell cortex that later growth [24,27,28] and by compelling evidence that plant
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464 Cell biology

hormones, such as ethylene and gibberellins, exert their these processes. The longitudinally oriented pollen tube
effects on the expansion of stem cells via the control of cor- actin bundles are believed to provide tracks for myosin-
tical microtubule orientation [2]. The complex microfibril dependent organelle movement [37,38], whereas the fine
organization observed in the wall of some cell types has actin structures in the tube apex may have a direct func-
been repeatedly used as an argument to challenge the tion in tip-directed targeted secretion. The polarity of the
model described above (e.g. [29]). However, fluorescently actin-dependent secretion machinery that mediates pollen
labeled microtubules in living cells have been shown to be tube elongation was shown to be controlled by a tip-
very dynamic structures that can potentially provide tem- focused Ca2+ gradient [41,42] and by Rac-related small
plates for even complex microfibril patterns [30]. GTPases, which induce the accumulation of phos-
phatidylinositol 4,5-bisphosphate specifically in the tip
Actin depolymerization also causes the abnormal deposi- membrane [43,44•].
tion of microfibrils [31] and affects polarized growth [32].
A function of filamentous actin in the control of micro- Microtubules are not essential for the elongation of
tubule organization has been determined in a range of pollen tubes, nor for the maintenance of their shape.
different types of plant cells [31,33–35]. Also, some sort of The disruption of cytoplasmic microtubules in pollen
polarized secretion, which may depend on actin filaments, tubes by various microtubule depolymerizing drugs
is likely to be required for polar cell expansion by diffuse alters cytoplasmic organization and moderately reduces
growth [32]. growth rate, but does not arrest cell elongation or affect
overall cell morphology [37,38,45].
Cell expansion during single cell morphogenesis
Certain types of plant cells with highly specialized func- Root hair elongation
tions undergo dramatic cellular morphogenesis during Root hairs are highly elongated, straight protrusions of spe-
differentiation. This group of cells includes pollen tubes, cialized epidermal cells that are oriented perpendicularly
root hairs and A. thaliana trichomes, which are extremely with respect to the root surface. Their main function is
elongated single cells that are either unbranched in the believed to be the absorption of water and nutrients from
cases of pollen tubes and root hairs, or branched, in the the soil. Root hair morphogenesis has been divided into two
case of A. thaliana trichomes. The cytoskeletal functions steps: first, the initial formation of a localized ‘bulge’ on the
that are required for the establishment of the shape of surface of a differentiating root epidermal cell, and second,
these three cell types, which are easily accessible for the subsequent elongation of this ‘bulge’ into a root hair
experimental manipulation and observation, are currently [46]. Bulge formation was recently shown to proceed nor-
under intensive investigation. mally in the presence of actin-depolymerizing drugs,
whereas pharmacological interference with microtubular
Pollen tube growth dynamics resulted in the outgrowth of multiple and mis-
Growing pollen tubes transport male generative cells localized bulges (J Mathur, N-H Chua, unpublished data).
through the style to the embryo sac where fertilization These results may indicate that root hair bulges are formed
can occur. Pollen tubes have a diameter of only 10–20 µm by microtubule and cell wall controlled growth that is driven
and can reach a final length of several centimeters [36]. by turgor pressure. By contrast, ample evidence suggests
They expand very rapidly by ‘tip growth’, during which that root hairs, once initiated, elongate by tip growth in a
new cell membrane and cell wall material is delivered similar manner to pollen tubes. Growing root hairs show
exclusively to the extreme apex by strictly polarized essentially the same cytoskeletal organization as pollen
secretion. Rapid movement of cell organelles through the tubes, exhibit cytoplasmic streaming and display a tip-
cytoplasm, known as cytoplasmic streaming, appears to focused Ca2+ gradient [47,48•,49•]. As observed with pollen
be required for sustained secretion at the tip and for tube tubes, the treatment of elongating root hairs with actin
elongation [37,38]. depolymerizing agents strictly inhibits cytoplasmic stream-
ing and cell elongation [48•]. The inhibition of cytoplasmic
Longitudinally oriented, thick actin bundles and micro- streaming by actin depolymeriszing drugs was found to be
tubules are present throughout the pollen tube cytoplasm reversible only in the presence of intact microtubules, indi-
with the exception of the apex. In most studies micro- cating that these cytoskeletal elements may be involved in
tubules have been found to be absent from the tip, actin organization in root hairs [50]. Interestingly, disruption
whereas the question of how actin is organized in this or stabilization of the microtubules in elongating root hairs
region is somewhat controversial [37,38]. Earlier studies was shown to cause a loss of growth directionality and to
describe a dense apical actin network, which more recent induce branching [49•], effects that are not observed in
work using improved methods has failed to detect [39,40•]. pollen tubes. Microtubules appear to play a key internal role
Instead, the presence of fine, elaborate actin structures at in ensuring that root hairs remain straight, and correctly ori-
the same location has been revealed. Treatment with actin ented with respect to the root surfaces as they grow out into
depolymerizing drugs results in the immediate inhibition the soil. In contrast, the direction of pollen tube growth
of cytoplasmic streaming and pollen tube growth, which within the style seems to be controlled externally, possibly
proves that intact actin filaments are required for both of by chemoattractants, and not internally by microtubules.
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Cytoskeleton in plant development Kost, Mathur and Chua 465

A large number of A. thaliana mutants with altered root hair mediates trichome expansion at this initial stage. When
morphology have been described [47], and their analysis applied at a later stage, microtubule depolymerizing or sta-
may provide additional clues on the role of the cytoskele- bilizing drugs completely inhibited branching (J Mathur,
ton in root hair morphogenesis and in tip growth. Mutants N-H Chua, unpublished data). The Zwichel gene, which is
that show exclusive growth defects either in root hairs disrupted in one of the ‘branching’ mutants, has been
alone, such as Rhd1, Rhd2 and Rhd4 [46], or in root-hairs found to encode a kinesin-type microtubular motor protein
and pollen tubes, such as Tip1 [51], are particularly inter- [54], and the Fass mutant, which has general microtubular
esting in this respect (Table 1). Tip1 mutants display short, defects, displays reduced trichome branching [55]
branched root hairs and reduced fertility, which is caused (Table 1). These findings provide strong evidence of a key
by a moderate inhibition of the elongation of morphologi- function for microtubules in the initiation of trichome
cally normal pollen tubes. The Tip1 defects are strongly branching. Evidently, the microtubular cytoskeleton deter-
reminiscent of the effects observed after treating root hairs mines the basic features of trichome shape by controlling
and pollen tubes with drugs that affect microtubule orga- both stalk formation and branch initiation. Filamentous
nization. The Tip1 gene product may therefore be required actin seems to be required during later phases of trichome
for microtubular functions specifically during tip growth. development and appears to be important for branch elon-
gation. Exactly how the two cytoskeletal elements
Trichome morphogenesis function in concert to bring about normal trichome mor-
Trichomes (i.e. leaf hairs) cover all aerial parts of higher phogenesis is under investigation in our laboratory.
plants. They are postulated to maintain a beneficial micro-
climate close to the plant surface and/or to serve as Establishment of growth polarity in Fucus zygotes
pathogen repellants. A. thaliana trichomes are single cells The zygotes of Fucus, a marine brown algae, serve as a
consisting of a stalk, which extends perpendicularly to the model system for the analysis of the early events involved in
epidermis, and typically three branches that are arranged the establishment of growth polarity in plant cells. These
in a particular orientation with respect to the axis of the zygotes are initially apolar but then polarize in response to
underlying organ. They are formed by specialized epider- external cues such as light. The polarization is reversible
mal cells, which undergo a regular, well defined expansion during an early phase, but becomes fixed after several hours.
program [52]. At present, very little is known about the cel- Both the establishment of a reversible polar axis and the
lular mechanisms involved in this process. Genetic screens subsequent stabilization of this axis are sensitive to actin
have, however, identified a considerable number of depolymerizing drugs. Actin-dependent relocalization of
mutants that are defective in trichome morphogenesis. unknown plasma membrane factors to one cellular pole is
These mutants were classified into three main groups: thought to represent the basis of the initial reversible phase
first, mutants completely impaired in the local outgrowth of the polarization process. These factors are believed to
of stalks from differentiating trichome cells, second, form a complex that organizes a site of actin-dependent tar-
mutants showing a ‘distorted’ phenotype with trichomes geted secretion, which locally delivers another unknown
that are twisted and have branches which do not elongate marker to the cell wall. It has been hypothesized that the
properly, and third, ‘branching’ mutants with trichomes irreversible accumulation of this marker in the cell wall sta-
that appear relatively normal but display increased or bilizes the pre-established polarity and triggers the
decreased branching [53]. subsequent outgrowth of a protrusion, the rhizoid tip, which
is the first visible manifestation of polarity during Fucus
We have recently started to examine the role of the development [56]. Similar actin-dependent processes may
cytoskeleton in trichome morphogenesis. Pharmacological be involved during the early stages of growth polarity deter-
disruption of the actin filaments was found to have no mination in the cells of higher plants.
effect on the initial outgrowth of stalks, but to severely
affect the later stages of trichome morphogenesis and to Cell differentiation
cause the formation of aberrant trichomes that very closely The development of tracheary elements (TEs), the build-
resemble those formed by mutants of the ‘distorted’ class. ing blocks of the water-conducting tubes in the xylem, from
In addition, we have observed that a number of mutants parenchyma cells is an impressive example of plant cell dif-
that belong to this class show striking defects in trichome ferentiation. Cells of Zinnia elegans that are derived from
actin organization (J Mathur, N-H Chua, unpublished mesophyll protoplasts can be induced to undergo differenti-
data). These findings indicate that a functional actin ation to TEs in vitro [57]. This system has been used to
cytoskeleton is essential for the later stages of trichome examine cytoskeletal functions in the formation of the trans-
morphogenesis and for branch elongation. versely oriented bands of secondary cell wall thickening that
are characteristic of mature TEs. At the onset of secondary
The treatment of developing trichomes during stalk out- cell wall synthesis, loosely and longitudinally arranged
growth with drugs that interfere with microtubule microtubular bundles become reoriented into dense trans-
dynamics led to a loss of growth polarity and to the forma- verse bands, which underlay exactly the pattern of the
tion of balloon-like structures. This evidence suggests that forming cell wall thickening. Actin patches, which are ini-
turgor-driven, microtubule and cell wall controlled growth tially dispersed at random between the longitudinally
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Table 1

A. thaliana and Z. mays developmental mutants with confirmed or proposed primary defects in cytoskeletal functions.

Mutant Species Developmental defect Cytoskeletal defect Gene product References

Pilz A. thaliana Early arrest of embryo development General defects in microtubule Unknown [63•]
Embryos formed are mis-shaped and consist organization
of only a few cells

Fass/Ton A. thaliana Abnormal seedlings with a highly Organization of cortical Unknown [55,69]
irregular internal cellular structure but microtubules affected: no
a clearly discernible apical-basal pattern preprophase bands irregular
Trichomes show reduced branching microtubule arrays during
interphase

Rhd1 A. thaliana Abnormal enlargement of root hairs at the base Root hair specific Unknown [46]
cytoskeletal defects?*

Rhd2 A. thaliana Root hairs are initiated but do not elongate Root hair specific Unknown [46]
cytoskeletal defects?*

Rhd4 A. thaliana Root hairs remain short and show Root hair specific Unknown [46]
highly abnormal morphology cytoskeletal defects?*

Tan1 Z. mays Morphologically normal leaves with highly Defective preprophase Unknown [15•]
irregular internal cellular structure band positioning and
cell plate guidance

Tip1 A. thaliana Short, branched root hairs and reduced Tip growth-specific Unknown [51]
pollen tube growth rate microtubular function disrupted?*

Titan A. thaliana Highly polyploid and enlarged endosperm nuclei Defects in cytoskeletal elements Unknown [73•]
Some Titan mutations cause embryo required for nuclear division?*
death after only a few cell divisions

Tso1 A. thaliana Aberrant floral organs and ovules with Defects in cytoskeletal elements Unknown [72•]
highly irregular internal cellular structure required for cell division and/or
cell expansion specifically during
flower and ovule development?*

Zwichel A. thaliana Short trichomes with reduced branching Defect in microtubule based Kinesin-like protein [54]
intracellular motility required
for trichome morphogenesis?*

*Cytoskeletal defects have been proposed based on the mutant phenotype and on the nature of the product of the mutant gene, but were not
confirmed by direct experimental evidence.

oriented microtubules, reorganize at the same time and endoplasmatic reticulum and are filled with unidentified,
eventually form structures that fill the gaps between the spike-like structures. Small molecules can freely diffuse
transverse microtubular bands. Actin depolymerizing drugs through the plasmodesmal cytoplasm, whereas the trans-
inhibit microtubule rearrangement and induce the forma- fer of macromolecules, such as proteins, is subject to
tion of a longitudinally oriented pattern of secondary cell control mechanisms and requires specific signals. Devel-
wall thickening [33]. These observations indicate that actin opmental regulators including the maize KNOTTED1
filaments are required for the reorientation and correct posi- protein are known to be transported through plasmodes-
tioning of microtubules during TE differentiation. mata. Cell-to-cell communication by plasmodesmal
Microtubules in differentiating TEs appear to serve as tem- transport could potentially play an important role in plant
plates for the ordered deposition of cell wall material, as development ([58]; see below).
they do in elongating cells in developing plant tissues.
Actin and myosin have both been immunolocalized to
Cell-to-cell communication plasmodesmata in plant and algal cells [59,60]. Treatment
The cytoplasm of large groups of plant cells is connected with actin depolymerizing drugs was found to enlarge
through plasmodesmata and forms a continuous symplas- plasmodesmata [60] and to allow the uncontrolled passage
tic domain. Plasmodesmata are plasma membrane-lined of macromolecules [61]. On the basis of these observa-
cytoplasmic channels in the cell wall that contain tubular tions, a key function for actin in the control of
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Cytoskeleton in plant development Kost, Mathur and Chua 467

plasmodesmal transport has been hypothesized. It was leaves, which appears to be caused by defective PPB posi-
suggested that the unidentified ‘spikes’ observed in the tioning and cell plate guidance. Amazingly, mature Tan1
cytoplasm of plasmodesmata are actin structures that leaves look morphologically normal, although their internal
block the free diffusion of macromolecules and control the cellular organization is completely irregular [15•] (Table 1).
channel opening [58].
Fass/Ton and Tan1 mutants clearly demonstrate that plant
Viral movement proteins, which enable plant viruses to morphogenesis is not simply governed by a cell-
spread from cell-to-cell by plasmodesmal transport, autonomous, internal program that controls the
were found to co-localize with actin filaments within orientation of cell division and the direction of cell expan-
cells and to bind actin in vitro [62]. This may indicate sion via the regulation of cytoskeletal organization.
that actin, possibly acting in conjunction with myosin, Instead, cells in developing tissues are apparently con-
has a function in actively transporting certain proteins stantly communicating with each other, reevaluating their
through plasmodesmata. position with respect to the growing organ to which they
belong and adjusting their behavior according to a global
Organogenesis morphogenetic plan.
The analysis of mutants has begun to complement studies
on single cell systems, which have provided most of our Visualization of cytoskeletal elements
knowledge about the function of the cytoskeleton in plant Electron microscopy, immunofluorescence techniques or
developmental processes. Mutations in the A. thaliana Pilz staining with fluorescently labeled phalloidin, a fungal
group of genes appear to disrupt microtubule functions metabolite that specifically binds to actin filaments, have
throughout the cell cycle and to arrest embryo develop- traditionally been used to visualize cytoskeletal elements
ment in its very early stages [63•] (Table 1). These in fixed plant cells or tissues. These powerful methods
observations may illustrate the fundamental importance of have allowed many important observations, although they
the cytoskeleton for plant development. are not without caveats. They work well for isolated single
cells, but are more difficult to apply to intact tissues. The
Interestingly, genetic evidence suggests that Pilz gene fixation procedures, which are involved in their use, may
products are not required for gametophyte development alter cytoskeletal organization; actin filaments are known
[63•], which involves mitotic and meiotic cell divisions as to be especially sensitive to such treatments. With the
well as dramatic cellular morphogenesis. Pilz genes there- introduction of techniques that allow microinjection of flu-
fore have highly tissue-specific functions. The copy orescent tubulin or phalloidin conjugates a decade ago, it
number of cytoskeletal genes in plants is generally surpris- became possible to visualize cytoskeletal elements in liv-
ingly high; for example, 10 actin genes [64], 15 tubulin ing plants cells [17,70]. This led to new and important
genes [65,66], 9 genes encoding actin depolymerizing fac- insights into the dynamics and the organization of the
tor (ADF; C-H Dong, N-H Chua, unpublished data) and at plant cytoskeleton. However, microinjection is technically
least 8 profilin genes [67,68] have been identified in demanding and its application is restricted to certain types
A. thaliana. Members of these large gene families show of cells. Recently published work demonstrated that the
high sequence homology and are likely to encode proteins expression of sequences encoding GFP fused to F-actin, or
with largely conserved functions. Tissue-specific and tem- to microtubule binding domains of animal proteins, can be
porally controlled expression of the different members of used to observe cytoskeletal elements in living plants cells
cytoskeletal gene families appears to be employed by in a non-invasive manner [40•,71•]. The stable expression
plants in order to control the various functions of the of GFP fused to the F-actin binding domain of mouse talin
cytoskeleton during organogenesis and development. in transgenic plants was shown to allow visualization of
actin filaments in a range of different plant tissues. The
A. thaliana Fass/Ton mutants have more specific defects in transgenic plants expressing this GFP fusion protein did
cytoskeletal organization than Pilz mutants: Fass/Ton cells not show any apparent abnormalities in actin organization
fail to establish both PPBs and normal cortical microtubule or development. The new GFP-based markers now allow
arrays during interphase, whereas other microtubular struc- researchers to perform studies that have previously been
tures, such as the mitotic spindle and the phragmoplast, impossible and will facilitate the future analysis of
are unaffected. Consequently, such cells undergo cell divi- cytoskeletal functions during plant development.
sion but can not control cell plate positioning or the
direction of cell expansion [69] (Table 1). Interestingly, Conclusions
mutant Fass/Ton seedlings, although they remain stunted Actin filaments and microtubules form a dynamic cyto-
with highly abnormal morphology and internal organiza- plasmic network that not only stabilizes spatial features
tion, are able to form an apical-basal axis, cotyledons and and controls the polarity of cells in developing plant tis-
even floral organs [55,69]. sues, but also provides the basis for the intracellular
motility and force generation that such cells require to
Disrupting the Tan1 gene in maize leads to a deregulation position forming cell plates, to expand in a controlled
of the orientation of cell division planes in developing manner and to communicate with each other. Generally,
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468 Cell biology

microtubules tend to be responsible for stabilizing, frame- Note added in proof

work-providing functions, whereas actin filaments are A manuscript describing our data on the functions of actin
often involved in dynamic processes and structural filaments and microtubules in trichome morphogenesis
rearrangements. Analysis of single cell model systems has has been accepted for publication [85].
provided substantial insights into the cellular functions of
the plant cytoskeleton. Although many of these functions
are expected to prove important for plant organogenesis,
Acknowledgements
We would like to thank Ulrich Klahre for critical reading of the manuscript.
the global role of the cytoskeleton in plant development is This work was supported by the Swiss National Science Foundation (grant
not yet understood well. Clearly, the analysis of plant #823A-046686 awarded to B Kost) and by the US Department of Energy
(grant #DOE94ER20143 awarded to N-H Chua).
developmental mutants is needed for a better definition
of this role. A large number of such mutants have been
described and many of them may have primary defects in References and recommended reading
Papers of particular interest, published within the annual period of review,
cytoskeletal functions (e.g. Tso1 [72•]; Titan [73•]; see have been highlighted as:
Table 1). The disruption of genes that encode cytoskele- • of special interest
ton-associated proteins (e.g. microtubule associated •• of outstanding interest
proteins or actin binding proteins), which constitute a 1. Nick P: Signaling to the microtubular cytoskeleton in plants.
large group of the key factors involved in either the regu- Int Rev Cyt 1998, 184:33-80.
lation or the function of the cytoskeleton, results in 2. Shibaoka H: Plant hormone-induced changes in the orientation of
aberrant morphogenesis in different organisms [74–76] cortical microtubules. Annu Rev Plant Physiol Plant Mol Biol 1994,
45:527-544.
and is also likely to cause developmental defects in plants
(e.g. the Zwichel trichome morphogenetic mutant [54]; see 3. Staiger CJ, Gibbon BC, Kovar DR, Zonia LE: Profilin and actin-
depolymerizing factor: modulators of actin organization in plants.
above and Table 1). Trends Plant Sci 1997, 2:275-281.
4. Hepler PK, Hush JM: Behavior of microtubules in living plant cells.
Plant Physiol 1996, 112:455-461.
As exemplified by the Pilz group of mutants, disruption of
5. Cleary AL: F-actin redistributions at the division site in living
cytoskeletal genes may often arrest plant development at a Tradescantia stomatal complexes as revealed by
very early stage. In such cases, the use of conditional microinjection of rhodamine-phalloidin. Protoplasma 1995,
mutants could allow the analysis of gene functions at later 185:152-165.
developmental stages. Alternatively, reverse genetics can 6. Staehelin LA, Hepler PK: Cytokinesis in higher plants. Cell 1996,
84:821-824.
be employed to study the role of cloned cytoskeletal genes
during particular developmental events. In this process 7. Samuels AL, Giddings TH, Staehelin LA: Cytokinesis in tobacco
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