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Acta Bot. Neerl. 39(1), March 1990, p. 1-
Review
The plant cytoskeleton: its significance in plant
development
J. Derksen F.H.A. Wilms and E.S. Pierson
Department of^ Experimental^ Botany, University^ of^ Nijmegen,^ Toernooiveld,^ NL-6525^ ED
Nijmegen, The^ Netherlands
CONTENTS
Introduction 1
Structure and composition of the cytoskeleton 1
General organization of the cytoskeleton 4
The cytoskeleton in cell morphogenesis and differentiation^7
The cytoskeleton in plant morphogenesis 12
Concluding remarks^12
Key-words: cytoskeleton, morphogenesis, cell^ differentiation.
INTRODUCTION
The informationconcerning the cytoskeleton in plant cells seems scanty when compared
to animal cells. However, as interest^ has^ been^ rapidly increasing, it^ will^ not be^ possible to
do justice to all^ studies^ concerning the^ plant cytoskeleton. Literature^ prior to 1980 has
been discussed extensively by Gunning & Hardham (1982), Hepler (1985) and Lloyd
(1982). For^ a recent review on the^ biochemistry and^ genetics of^ plant cytoskeletal proteins
see Fosket^ (1989).
As plant cells are immobile, a particular plant shape can only be^ obtained^ by directional
cell divisionand cell expansion, i.e. by polar growth, or specific cell death.^ Thus, cell^ division
and cell expansion within a tissue must occur in a co-ordinated way. Here we shall describe
first structure and composition of the cytoskeleton of eukaryotic cells in general and
following this^ we will^ discuss^ recent^ results^ on the^ organization of^ the^ cytoskeleton in^ higher,
i.e. embryonic, plants with special reference to morphogenesis and cell differentiation.
STRUCTURE AND^ COMPOSITION^ OF^ THE^ CYTOSKELETON
Microtubules Microtubules (^) are tubular structures with (^) an internal diameter of about 15nm and (^) an
average external^ diameter^ of^ about^25 nm.^ They^ consist^ of^ two^ evolutionally^ related
The cytoplasm of eukaryotic cells contains a three-dimensional network of filaments: the
cytoskeleton. It^ connects^ the^ various^ organelles and^ other^ cytoplasmic elements^ of^ the^ cell
with each other and with the plasma membrane and is involved in many dynamic pro-
cesses in^ the^ cell, including cell^ division, morphogenesis, redistribution^ of^ surface
components, endo-^ and^ exocytosis^ and^ the^ positioning^ of^ cytoplasmic^ elements^ (reviews:
Dustin 1984; Lackie 1986; Bershadsky & Vasiliev 1988; plants: Traas 1989).
The three major constituents^ of this^ system are microtubules, microfilaments and^ inter-
mediate filaments. They can be discriminated, based on their diameter and by means of
immunocytochemistry.
2 J. DERKSEN ET A L.
proteins, a-tubulin^ and^ p-tubulin, each^ with^ a molecular^ weight of^ approximately
50 000 kD. The microtubular wall is generally made up of 13 protofilaments, but different
numbers occur. Protofilaments consist of a chain of about 8 nm long, a-P tubulin dimers
that are asymmetrical and are all oriented in one direction, resulting in polar proto-
filaments. The a-tubulin side is called the -|-side, the P-tubulin side the —side. In a
microtubule all protofilaments have the same orientation and are shifted about 1 nm in
respect to^ each^ other.^ Microtubules^ thus^ show^ a^ distinct polarity^ with^ dimers^ in^ a^ 10°
left-handed helix^ (Dustin 1984).
Microtubules are no static elements but continuously assemble and disassemble. Two
mechanisms have been proposed to explain these processes: treadmilling (Margolis &
Wilson 1978) and dynamic instability (Mitchison & Kirschner 1984). The first model
assumes a steady state^ with^ a continuous^ assembly at^ the^ -(-side and^ disassembly at^ the
— side. Dynamic instability predicts the existence of a population of rapidly disassembling
microtubules together with a population of slowly, eventually at both ends, growing
microtubules. In mitotic spindles in vivo, microtubules appear to behave according to
the dynamic instability model (Salmon et al. 1984). However, in the interphase cells of
animals, part of^ the^ microtubules^ appeared to^ be^ much more stable. In such cases their
dynamics have been^ described^ as a tempered mode^ of^ dynamic instability (e.g. Sammak^ &
Borisy 1988). Tubulins^ of^ higher plants appear to^ be^ less^ conservative^ than^ in^ animal^ cells
(Cleveland et al.^ 1980; Fosket^ 1989), which^ has^ been^ attributed^ to^ a different^ evolutionary
pressure. This^ differencemay result^ from^ the^ disappearance^ of^ cilia^ and^ flagella^ (Cavalier-
Smith 1978), but may as well relate to the occurrence of a cell wall. This difference may also
explain the^ relative^ insensitivity of^ plant microtubules^ to colchicine^ and^ their^ sensitivity to
some herbicides^ (Bajer &^ Mole-Bajer 1986a). A^ variety of^ proteins has^ been^ found^ to
associate with microtubules: the microtubule-associated proteins (MAPs) (review:
Olmsted 1986, see also below). MAPs largely determine microtubule stability and its
association with^ other^ cytoskeletal elements and organelles. Their activity appears to be
regulated by a cascade^ of^ phosphorylating enzymes in which Ca 2+/calmodulin and cAMP
may play^ an^ important^ role^ (Theurkauf^ &^ Vallee^ 1982;^ Larsson^ et^ al.^ 1985,^ Schulman^ et^ al.
Microtubules have been related to various types of movements: besides their well
known role in movement of metaphase chromosomes and of cilia/flagella, they can be
involved in the transport of vesicles and organelles, like in neurons and other animal cells
(Dustin 1984; Bershadsky & Vasiliev^ 1988). Microtubules^ are probably also involved in
the organization and dynamics of the endoplasmic reticulum (Chen & Lee 1988; Vale &
Hotani 1988).
Microfilaments
The backbone protein of microfilaments is formed by actin which has a molecular weight
of about 42 kD. In the filaments actin molecules are piled up in such a way as to form a
double-stranded twisted rope. The major repeat distance is 38 nm, involving 13 molecules.
Individual microfilaments have a diameter of 5-7 nm and show a distinct polarity.
Microfilaments are also dynamic and both treadmilling and dynamic instability have been
considered to occur. Microfilaments often occur in large bundles with a regular organiz-
ation like in muscles and stress fibres, where they are very stable (reviews: Lackie 1986,
Bershadsky &^ Vasiliev^ 1988). They can also^ form^ loose^ bundles, such^ as in^ many plant
cells (review; Staiger & Schliwa^ 1986). The^ microfilaments^ that can be seen in electron
microscopic preparations represent actin^ filaments, which^ have^ recently been^ confirmed
(^4) J. DERKSEN ETAL.
The presence of ankyrin-like and spectrin-like proteins indicates that the associations of
actin with the membranes (^) may be similar to those in animal cells.
We expect that most cytoskeletal proteins of animal cells will be present somehow in
plant cells.
GENERAL ORGANIZATION OF THE CYTOSKELETON
The techniques used
The organization of cytoskeletal elements has been studied using a number of techniques.
Electron microscopy. Classical electron microscopical (EM) techniques enable visualiz-
ation of microtubules, microfilaments and intermediate filaments, but due to the con-
ditions used, cytoskeletal elements, especially actin filaments, may become depolymerized
during the^ procedure and^ fibrous elements^ may become^ only poorly contrasted^ as
compared to the^ embedding medium, or are obscured^ by soluble^ components of^ the
cytoplasm. Moreover, only small^ areas can be^ studied.
Fixation problems can be overcome by the use of cryo-techniques, (Hereward &
Northcote 1973; Howard & Aist 1979; Emons & Derksen 1986; Lancelle et al. 1986;
Craig &^ Staehelin^ 1988). The^ use of^ these^ techniques may be^ conditional for the use of
immunological probes at^ the^ ultrastructural^ level^ (e.g. Lancelle^ & Hepler 1989).
Large surfaces^ allowing quantitative analyses can be^ obtained^ by using cleaving
techniques (Traas 1984; Traas^ et^ al.^ 1985). Also^ sections^ of^ polyethylene glycol (PEG)
embedded material (Wilms 1990) and whole mounts (Hawes 1985) may be useful. Larger
cell parts can also be studied using serial sectioning, though this procedure is laborious.
Light microscopy. Cytoskeletal elements appear to^ be evolutionally conservative and
antibodies raised against animal proteins, especially tubulin, could be used in plant cells.
Immunofluorescent (IF) probes allow study of the spatial organization in entire cells
(Lloyd et^ al.^ 1979; Wick^ et^ al.^ 1981) and^ in^ sections^ of^ PEG-embedded^ material^ (Hawes &
Table 1. Microtubule (MT) and actin filament (AF) associated proteins that have been identified in
the cytoplasm of^ cells of^ higher plants
Protein Group References Possible role
MAPs MT Cyr & Palevitz (1989) Microtubule bundling
Kinesin MT^ Moscatelli et al.^ (1988) Force-generating, organelle transport.
Calmodulin MT^ Wick^ et al. (1985) Ca 2+-binding protein, regulatory
functions
Troponin AF/MT Lim^ et al.^ (1986) Ca^ 2+-binding protein binds^ to
tropomyosin, regulatory functions
Ankyrin AF^ Wang& Van^ (1988) Connecting actin^ to^ membrane^ proteins
Myosin AF^ Vahey et^ al.^ (1982)
Parke et al. (1986)
Yan et al. (1986)
Grolig et al.^ (1988)
Tang et al.^ (1989)
Organelle movement, force^ generation,
intracellular (^) movements
Spectrin AF^ Wang &^ Yan^ (1988) Connecting actin^ to^ membrane^ proteins
CYTOSKELETON SIGNIFICANCE IN DEVELOPMENT 5
Horne 1985). The^ introduction of fiuorochrome conjugates of^ phalloidin (Wulf et al.
1979) allowed^ investigation of^ the^ three-dimensional^ organization of^ actin^ filaments,
though antibodies^ have^ been used^ as well^ (McCurdy et^ al.^ 1988)).
The IF methods are extremely useful for large scale-studies, but for a detailed analysis
EM techniques are required (e.g. Segaar 1990).
The use of advanced light microscopic techniques in plant cells such as confocal scan-
ning laser^ microscopy (CSLM; e.g. Quader &^ Schnepf 1986) and^ video/computer
enhanced image photography (e.g. Lichtscheidl & Weiss 1988) will contribute to a better
understanding of^ the^ dynamics of^ the^ cytoskeleton and^ its^ interactions^ with^ other^ cellular
structures.
General organization
Microtubules are present throughout the cytoplasm, but large arrays of parallel cortical
microtubules are always present, and especially conspicuous in vacuolated cells (reviews
in Gunning & Hardham 1982; Lloyd 1984; Traas 1989; see also Fig. 1). The cortical
microtubules (^) are interconnected and (^) thought to form (^) an almost (^) uninterrupted helix
throughout the^ cell^ (Lloyd 1984). Recently it^ could^ be^ shown^ that^ in^ protoplasts of
Nicotiana about 50% of^ the^ cortical^ microtubules^ is^ regularly interconnected^ (H. Kengen
& J. Derksen, unpublished data). If^ these^ interconnections were of a dynein-type such
interconnected microtubules could not only withstand strong forces, but could even exert
considerable forces (^) on their environment.
At the surface of the nucleus microtubule organizing centres (MTOCs) are present that
initiate microtubule assembly after cell division(e.g. Wick & Duniec 1983). They probably
do not determine the organization of the cortical cytoskeleton, as nucleating sites are
also present at the plasma membrane (Gunning et al. 1978) and in anucleate cytoplasts
the microtubular skeleton may self-assemble into highly organized patterns (Bajer &
Fig. 1.^ Microtubules^ in^ root cortex cells^ of^ Lepidium. Immunofluorescence^ preparation according to Traas^ el al. (1984). In^ the^ left^ two cells^ the^ microtubules^ are helical, in^ the^ two cells^ on the^ right, the^ microtubules^ are almost
parallel to the^ longitudinal axis^ of^ the^ cell.^ Magnification: x^ 1000.
CYTOSKELETON SIGNIFICANCE IN DEVELOPMENT 7
Single microfilaments were co-aligned with^ microtubules^ over distances^ up to 1 -46 pm in
dry cleaved^ preparations of^ Nicotiana protoplasts (H. Kengen & J. Derksen, unpublished
data); see also^ Fig. 3). However, doubt exists about the^ actual^ nature of^ these^ filaments
(see below).
A clear co-localization over large parts of the cell has been observed in immunofluores-
cent preparations of pollen tubes (Raudaskoski et al. 1987; Pierson et al. 1989) and the
distribution of the^ microtubules has been reported to depend, at least partly, on actin
distribution(Derksen & Traas 1984).
Intermediate filaments of plant cells co-localize with microtubules in a patchy way
(Goodbody et^ al.^ 1989). The^ present proteins may be^ at^ least^ partly identical^ to^ the
filaments that are visible in dry-cleaved cells and that accompany microtubules and con-
nect coated pits with microtubules (Emons&Traas 1986; Quadereta/. 1986; H. Kengen&
J. Derksen, unpublished data). The^ distribution^ of^ the^ larger bundles^ is^ independent from
an intact^ actin^ skeleton, but^ depends on microtubules.^ They become^ dispersed after
microtubule degradation (Goodbody et al. 1989).
THE CYTOSKELETON IN CELL MORPHOGENESIS AND
DIFFERENTIATION
Determination of the^ division^ plane
The position of the nucleus may be largely determined by microtubules, as anti-
microtubular drugs may cause disposition of the nucleus (Clayton & Lloyd 1984).
Prior to cell division, the nucleus will move towards the future division plane, which
may be^ determined^ by^ microtubules^ radiating^ from^ the^ nucleus^ to^ the^ cells^ periphery^ and
vice versa (Burgess 1970; Pickett-Heaps 1974). These microtubules may already be present
during phragmosome formationand^ coincide^ in^ time^ and^ place with the formationof the
pre-prophase bands^ (Venverloo &^ Libbenga 1987). However, a role^ of the^ preprophase
bands in nuclear positioning is not unquestioned (Clayton & Lloyd 1984; Mineyuki et al.
1988). The^ fundamental^ regulatory mechanisms^ are unknown.^ Also, microfilaments^ are
Fig. 3.^ Micrograph of^ a dry-cleaved preparation of^ a protoplast from^ a cell^ culture.^ Numerous^ putative
microfilaments are (^) present at the surface (^) (J.J.). Often these filaments seem (^) to be connected with coated (^) pits (^) (\J7), or to^ accompany microtubules^ (T). Magnification; x^59 000.^ (Photograph: H.^ Kengen.)
Nicotiana
8 J. DERKSEN ET AL
present in^ the^ preprophase^ bands^ (McCurdy^ et^ al.^ 1988;^ Lloyd^ &^ Traas^ 1988).^ During
metaphase the preprophase band microtubules disappear but the preprophase band actin
filaments remain, and^ may help to^ guide the^ cytokinetic apparatus out along the^ pre-
determined path (Traas et al. 1987; Lloyd 1988). Both microtubules and actin filaments
show distinct configurations during the meiotic divisions (see Lammeren et al. 1989; Traas
et al. 1989; Bednara et al. 1990) and are probably involved in the co-ordination of the
meiotic division process, but probably only microtubules are involved in the establish-
ment of^ cell^ polarity (Traas et al.^ 1989).
Calmodulin is associated with the spindle and the phragmoplast, and not with the pre-
prophase band^ (Gunning &^ Wick^ 1985), which^ indicates^ a regulatory role^ in^ cell^ division
but not in determination of the division plane. The actual mechanisms underlying the
changes in^ cytoskeletal organization prior to^ cell^ division^ are still^ basically unknown.
Hormonal control of the cytoskeletal organization
Little is known about the precise relationship between hormones and the cytoskeletal
elements. Ethylene causes a re-orientation of cortical microtubules in epidermis cells of
Pisum sativum and Vigna radiata (Steen & Chadwick 1981; Lang et al. 1982; Roberts et al.
1985) within^ a few^ hours.^ In^ cortex^ cells^ of^ tobacco^ explants, re-orientation^ of^ micro-
tubules is accelerated after ethylene treatment (Wilms & Wolters-Arts 1989). Doonan
et al.^ (1985) showed^ that^ high concentrations^ of^ benzylaminopurine (100 gM) can cause
depolymerization of^ microtubules^ in^ tip-growing cells^ of^ Physcomitrella. Gibberellic^ acid
causes rearrangement of^ cortical^ microtubules^ in^ epidermal cells^ of^ pea internodes
(Akashi &^ Shibaoka^ 1987). It^ increases^ the^ number^ of^ transverse^ microtubules^ and^ it
prevents depolymerization^ of^ microtubules^ by^ colchicine,^ cremart^ and^ low^ temperature
(Mita &^ Shibaoka^ 1984). However, the^ effects^ of^ gibberellic acid^ are diverse:^ it^ may protect
growth against cochicine^ inhibitionbut^ conversely, growth stimulation^ by gibberellic acid
may be^ inhibited^ by^ colchicine.^ The^ differences^ however,^ may depend^ on^ the^ colchicine
concentration used (Fragata 1974; for discussion see also Mita & Shibaoka 1984).
Hormones also may act indirectly on the organization of the cytoskeleton. Auxin
treatment effects^ the^ direction^ of^ cell^ expansion and^ the^ orientation^ of^ the^ cortical micro-
tubules in epidermal cells of maize coleoptiles (Bergfield et al. 1988). Since auxin induces
ethylene production (Imaseki 1985), it^ may be^ concluded^ that^ the^ action^ of^ auxin^ on the
organization of^ the^ cytoskeleton is^ indirect.^ However, the^ presence of^ auxin^ seems to^ be
conditional for the development of a cortical microtubular skeleton in protoplasts of
Medicago mesophyll cells^ (Meijer &^ Simmonds^ 1988). Hormones^ may act^ on cell^ polarity
by changing the^ distribution^ of^ cation^ pumps and^ channels^ on the^ plasma-membrane
(Saunders &^ Hepler 1981; Saunders^ 1986; review:^ Schnepf 1986). As^ Ca^ 2+may affect
both microtubules and microfilaments (see above), such changes may also affect the
organization of cytoskeletal elements.
Control of cell wall deposition
It is^ generally believed^ that^ the^ orientation^ of^ nascent^ cellulose^ microfibrils^ is^ controlled^ by
cortical microtubules. Several models have been proposed to describe microtubular con-
trol of microfibril orientation (reviews; Robinson & Quader 1982; Heath & Seagull 1982).
All models imply that microtubules connected to the plasma membrane, i.e. the cortical
microtubules, would^ prevent free^ diffusionof^ cellulose^ microfibril^ synthesizing complexes
in the^ membrane, leading to parallelism of^ microtubules^ and^ nascent microfibrils^ (see also:
Herth 1985). Also the insertion of Golgi-vesicles with non-cellulosic wall material has
10 J.^ DERKSEN^ ET^ AL
In stretching cells, microtubules generally appear to be oriented transverse to^ the direc-
tion of^ cell expansion (Gunning 1981; Gunning & Hardham^ 1982). In many cells, the
microtubular skeleton shows helix-like configurations with different pitches (Lloyd 1983,
1984; Traas^ et al.^ 1985; Traas^ 1989; see also^ Fig. 1).
Traas (^) et al. (^) (1984) supposed that (^) microtubules in (^) expanding root cortex (^) cells of
Raphanus would^ be^ oriented^ transverse^ to^ the^ vector-sum^ of^ both^ axial^ and^ circumfer-
ential cell expansion, more or less as in^ stretching cells.^ The^ orientation^ of^ microtubules^ in
the tip of tip-growing cells^ would be determined in^ a similar^ way (Traas et al. 1985; Emons
1989). The^ presence of^ randomly organized microtubules^ at^ non-expanding surfaces, of
meristematic and cortex cells in these roots, may support this assumption (Derksen et al.
1986a).
It remains open whether these helical organizations result from^ cell expansion, or
whether cell^ expansion is^ determined^ by these^ microtubules.^ The^ latter^ could^ occur
indirectly, by the^ control^ of^ cellulose^ deposition, or directly by resisting turgor pressure.
This might occur either by the rigidity of the microtubular skeleton or by active force
generation involving microtubule-based^ motors.
Lloyd and^ coworkers^ assumed^ that^ the^ helical^ configurations would^ behave^ more or
less like a spring, which becomes^ stretched out during cell expansion (Lloyd 1984; Roberts
et al.^ 1985). Such^ a behaviour^ cannot^ be^ reconciled^ with^ the^ occurrence of^ helical^ patterns
as observed^ in^ Raphanus and^ Pisum^ root^ cells^ (Traas et al.^ 1984; Hogetsu &^ Oshima^ 1986)
and in Avena coleoptiles and mesocotyls, and Pisum epicotyls (Iwata & Hogetsu 1988). In
these studies the changes from transverse to oblique or longitudinal occur mainly after
elongation has^ ceased.
Both assumptions relate the microtubule orientation to cell expansion but fail to point
out the exact mechanisms.
The first assumption requires a rather dynamic behaviour of microtubules, whereas the
second one demands a more static, passive one, yet needs considerable reorganization of
the microtubules to compensate for the loss in diameterof the helix during stretching.
In tissue explants of^ Nicotiana,the orientation^ of^ microtubules^ in^ the cells changes
from transverse to longitudinal to the long cell axis immediately after explantation. This
change occurs gradually without^ appreciable cell^ extension^ and^ has^ been^ related^ to^ de-
differentiationand a change in cell polarity. However, ethylene production by wounding
might also^ play a role^ (Wilms &^ Derksen^ 1988). To^ explain the^ mechanism^ involved,
Wilms & Derksen took into account the dynamic properties of the microtubules. They
supposed that^ the^ change in^ orientation^ resulted^ from^ polarity determining factors^ in^ the
cytoplasm, i.e.^ at^ the^ plasma membrane, during a dynamic phase of^ the^ microtubulesafter
explantation.
The behaviour of the cortical actin filaments during cell elongation has not yet been
described, but^ the organization of the endoplasmic bundles remain^ essentially the same
(Derksen et^ al.^ 1985).
Differentiation
Cell differentiation is the event leading to cells with quantitatively or qualitatively
different functions. In plants, differentiation is often accompanied by a local wall
deposition. Here^ we will^ discuss^ a few^ examples, namely vascular^ elements, stomatal^ cells
and statocytes.
The densities of microtubules increase gradually just before the initiation of wall
thickening along the^ lateral^ walls^ of^ young sieve^ elements^ in^ root^ protophloem of^ wheat
CYTOSKELETON SIGNIFICANCE IN DEVELOPMENT 11
(Eleftheriou 1987). Xylogenesis is^ one of^ the^ best^ studied^ cases with^ respect to^ the^ cyto-
skeletal organization during cell differentiation.In xylem cells, wall material is deposited
locally in^ close^ relation^ with^ cytoskeletal elements^ (Goosen-de Roo 1973, Herth 1985).
The formation of bands of microtubules before local wall deposition starts has also been
reported for^ Zinnia elegans cells^ in^ culture, which differentiate^ into^ xylem elements
(Falconer &^ Seagull 1988; Kobayashi et al.^ 1988). These^ cells^ have been^ extensively
studied by Falconer & Seagull and Fukuda & Kobayashi, who recently reviewed this
particular system (Fukuda &^ Kobayashi 1989).
The specific bands^ of microtubules at the sites of local wall deposition are reached via a
characteristic sequence of actin and microtubule patterns. The initially more or less axial
patterns of^ actin^ filaments^ disappear^ and^ large^ dots^ of^ actin^ are^ seen^ regularly^ distributed
over the^ surface.^ Meanwhile^ the^ axial^ microtubules^ change their^ orientation^ to^ oblique,
forming a network^ with^ the^ actin^ dots^ in^ the^ darns.^ This^ pattern changes gradually until
transverse and oblique orientations are predominant. At the same time the microtubules
form bundles. The actin dots are present exclusively between the microtubule bands and
are finally^ located^ under^ the^ sites^ of^ cell^ wall^ deposition.^ Colchicine^ destroys the^ regular
microtubule pattern and cell wall deposition (see also Herth 1985). Cytochalasin disrupts
the regular pattern of actin filaments and also affects the microtubule pattern, like in
pollen tubes^ (Derksen &^ Traas^ 1984). Microtubules^ and^ microfilamentsappear to^ act^ in^ a
co-ordinated way. In Zinnia, calmodulin is found between the regions of microtubule
bundles and wall deposition (Dauwalder et al. 1986), which may indicate a role in the
changes of^ the^ specific microtubule^ and^ actin^ filament^ patterns.
During stomalal^ development in^ grasses, asymmetric divisions^ of^ the^ guard mother
cells take place, forming the guard cells and the subsidiary cells. Their function depends at
least partly on the local deposition of wall material which finally results in the kidney-like
shape of^ the^ guard cells.^ The^ organization of^ microtubules^ reflects^ the^ orientation^ of
cellulose microfibrils in periclinal walls. In these cells a clear correlation between micro-
tubules and (^) cellulose microfibril orientation (^) appears to (^) be (^) firmly established. Like (^) in
vascular elements, microtubule density increases prior to the deposition of the secondary
cell wall (reviews; Palevitz 1982; Kristen 1986).
The initial, asymmetric, radial^ division^ of^ the mother cell is preceded by the asymmetric,
radial organization of^ cortical^ microtubules and^ preprophase bands^ in^ the^ plane of the
new cell^ wall.^ The^ radial^ arrays in^ both^ the^ pairs of^ guard cells^ and^ the^ pairs of^ subsidiary
cells from mirror^ images (Cho & Wick 1989; Cleary & Hardham 1989; Mullinax &
Palevitz 1989; Palevitz & Mullinax 1989, and references in these papers). During further
development, the^ organization of^ the^ microtubules^ changes from^ radial^ to transverse^ in
the subsidiary cells and from radial, over transverse and oblique, to axial in guard cells. In
Avena, in^ the^ last^ stage of^ differentiation, both^ guard cells^ and^ subsidiary cells^ show^ axial
microtubule patterns (Palevitz & Mullinax 1989). Thus, here too, a clear sequence in
microtubule patterns occurs in a co-ordinated way in the different cells.
Statocytes of^ Lepidium show^ a distinct^ cytoplasmic organization. The^ nucleus^ is
situated in the top of the cell, whereas in the distal part of the cell, amyloplasts and rough
endoplasmic reticulum^ are present that^ are involved^ in^ graviperception (Volkmann &
Sievers 1979). This typical organization is entirely actin dependent, as has been shown by
Hensel (1985, 1987). Microtubules appear to be less involved but may yet contribute in
stabilizing the^ distal^ endoplasmic^ reticulum^ (Hensel^ 1984). Thus,^ endoplasmic^ reticulum
organization seems to^ depend on actin^ filaments^ and^ not^ on microtubules.^ Also the
position of^ the^ nucleus^ seems to^ depend on actin^ filaments^ here, whereas^ in^ other^ systems it
CYTOSKELETON SIGNIFICANCE IN DEVELOPMENT 13
determine from static images alone, the use of newly developed light microscopic
techniques also^ appears to^ be^ very promising in^ plant cells.
However, the^ basis^ for^ a further^ understanding of^ the^ organization and^ function^ of^ the
cytoskeleton is^ set, as a start^ has^ been^ made^ in^ the^ identification, detection^ and^ localiz-
ation of intermediate filaments and the cytoskeleton-associated proteins. The latter may
be of particular interest also for the still pending discussion on microtubular control of
cellulose microfibril deposition, a control that must occur via an interaction with the
plasmamembrane and^ that^ limits^ lateral^ diffusion^ of^ the^ cellulose^ synthesizing complexes.
As several groups are studying differentialgene expression of cytdskeletal proteins and
genetic studies^ to^ the function of the^ cytoskeleton are also^ en route, the near future
promises to^ be^ exciting for^ those^ botanists^ interested^ in^ the^ cues of^ plant morphogenesis
and plant cell differentiation.
ACKNOWLEDGEMENT
The authors are indebted to Professor M.M.A. Sassen for his critical support during the
preparation of^ the^ manuscript.
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