Annals of Botany 107: 1213– 1222, 2011
doi:10.1093/aob/mcq229, available online at www.aob.oxfordjournals.org
REVIEW: PART OF A SPECIAL ISSUE ON THE PLANT CELL CYCLE
The organization of roots of dicotyledonous plants and the positions
of control points
Thomas L. Rost*
Department of Plant Biology, University of California, Davis, CA 95616, USA
* E-mail [email protected]
Received: 18 August 2010 Returned for revision: 8 September 2010 Accepted: 21 October 2010 Published electronically: 29 November 2010
Key words: Root apical meristem, RAM, cell cycle, differentiation, peripheral root cap, closed RAM organization,
open RAM organization, epidermis, module, determination, levels of organization, plasmodesmata, T-division, root
cap/protoderm initial, columella initial.
IN T RO DU C T IO N
Root tips of dicotyledonous plants have been studied for well
over a hundred years, with many papers published on root
apical meristem (RAM) organization and many other aspects
of root structure and development (Popham, 1966; Groot and
Rost, 2001a, b; Groot et al, 2003; Heimsch and Seago,
2008). Despite this attention, plant biologists tend to disagree
on many fundamentals of the structural organization of roots
and the terminology used to describe them. In this short
essay, I intend to step away from these controversies and
instead discuss observations on some aspects of root structure
made primarily in my laboratory over a 40 year period. I would
not be so bold as to state that every generalization I intend to
make will prove to be law in the long run, but the real intention
is to point out control points in the tissues of a root tip where
differentiation decisions are made and where molecular biologists need to focus their attention in determining the locations
of genetic action and how these decision points are affected by
such things as the root growth rate. Before continuing, I need
to acknowledge the work of the undergraduate and graduate
students, post-doctoral scholars and collaborators with whom
I have worked, and many of their published studies will be
referenced in what follows.
One reason for the apparent lack of agreement on root structure issues is that the organization and cellular patterns within
a root are constantly in flux during root development. As will
be pointed out, the organization of the RAM changes during
the root growth cycle, and also roots are remarkably responsive
to their environment and consequently keep changing (Rost
and Baum, 1988; De Tullio et al., 2010). Researchers who
fail to take this into consideration will sometimes jump to
different conclusions on structural issues when what they are
actually seeing is a change in structure due to the responsiveness of the root to its environment. Is it possible to describe a
normal condition for anything inside a root? I think it is, but it
is important to consider carefully and describe growth conditions and the root growth cycle stage and timing.
What should we know about differentiation control points in
the context of complex root tissues? One obvious place to start
# The Author 2010. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved.
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† Background The structure of roots has been studied for many years, but despite their importance to the growth
and well-being of plants, most researchers tend to ignore them. This is unfortunate, because their simple body
plan makes it possible to study complex developmental pathways without the complications sometimes found
in the shoot. In this illustrated essay, my objective is to describe the body plan of the root and the root apical
meristem (RAM) and point out the control points where differentiation and cell cycle decisions are made.
Hopefully this outline will assist plant biologists in identifying the structural context for their observations.
† Scope and Conclusions This short paper outlines the types of RAM, i.e. basic-open, intermediate-open and
closed, shows how they are similar and different, and makes the point that the structure and shape of the
RAM are not static, but changes in shape, size and organization occur depending on root growth rate and development stage. RAMs with a closed organization lose their outer root cap layers in sheets of dead cells, while
those with an open organization release living border cells from the outer surfaces of the root cap. This observation suggests a possible difference in the mechanisms whereby roots with different RAM types communicate
with soil-borne micro-organisms. The root body is organized in cylinders, sectors (xylem and phloem in the vascular cylinder), cell files, packets and modules, and individual cells. The differentiation in these root development
units is regulated at control points where genetic regulation is needed, and the location of these tissue-specific
control points can be modulated as a function of root growth rate. In Arabidopsis thaliana the epidermis and peripheral root cap develop through a highly regulated series of steps starting with a periclinal division of an initial
cell, the root cap/protoderm (RCP) initial. The derivative cells from the RCP initial divide into two cells, the
inner cell divides again to renew the RCP and the other cell divides through four cycles to form 16 epidermal
cells in a packet; the outer cell divides through four cycles to form the 16 cells making up the peripheral root
cap packet. Together, the epidermal packet and the peripheral root cap packet make up a module of cells
which are clonally related.
1214
Rost — Root organization and the position of control points
is the organization of the RAM. How is the RAM organized?
Is it static during root development? Is it the same in all plant
families? Does the RAM organization pattern have a function?
Relative to the body of the root, are there control points where
developmental decisions are made that regulate differentiation
events? Is the location of these control points static during root
development? What controls the spatial relationship between
differentiation events? Finally, relative to cell cycle events,
how is the location of cell cycle events regulated? How is
the plane of division controlled? Is there a mechanism to
‘count’ the number of divisions within a cell file and cell
module? Some of these questions will be addressed in the following from the perspective of where the control points are,
but not from the perspective of defining the genetic or hormonal control of those control points; that is left for the new
scientists to discover. I will start with the RAM.
Many papers on RAM organization have been published,
going back to the 1800s (Hanstein, 1870; Janczewski, 1874)
and forward to more recent papers by Clowes (1981), Groot
et al. (2003) and Heimsch and Seago (2008). Some analyses
have simplified patterns and others have plotted many complicated schemes with types and sub-types (Popham, 1966), and
in the exhaustive study by Heimsch and Seago (2008) 15
different types of RAM organization were identified. The
scheme I will describe is possibly overly simplified but was
used in the phylogenetic comparison study by Groot et al.
(2003). One thing to note, however, is that in some families
of plants more than one RAM organization type may exist
(Heimsch and Seago, 2008), so this analysis or any other
must be considered tentative and open for additional analysis.
In the scheme of Groot et al. (2003) there are three types of
RAM – closed (Fig. 1A), intermediate-open (Fig. 1B) and
basic-open (Fig. 1C). In their study they compared the RAM
organization of representatives from many different families
and plotted their RAM type based on the DNA sequence phylogeny data analysis of Soltis et al. (2000). This suggested that
(1) Closed RAM organization. In this type, there are three
specific tiers of initial cells (stem cells) from which all
other cells in the root body and cap are connected by
lineage. One tier for the vascular cylinder, one for the
cortex and the other for the epidermis and root cap (flax;
Fig. 1A). A fourth tier of initials called the ‘calyptrogen’
derives only the root cap, is found only in grasses and is
lacking in roots of dicotyledonous plants.
(2) Intermediate-open organization. In this type, examination
of the zone at the boundary of the root cap and body
shows some level of organization, particularly for the epidermis/root cap lineage, but the clear and specific identity
of the initial tiers is lacking (carrot; Fig. 1B).
(3) Basic-open organization. In this type, the identity of cell
files cannot be clonally predicted and instead a zone of seemingly disorganized initials replaces a zone of distinct initials ( pea, Fig. 1C).
The next question is, ‘Does the RAM organization pattern
remain constant throughout the growth cycle of a root?’ To
examine this question, we need first of all to look at the
growth cycle of a root; in this case we will use the primary
root of a seedling as the model. In studies of the primary
root growth cycle of pea (Rost and Baum, 1988; Gladish and
Rost, 1993), cotton (Reinhardt and Rost, 1995), Arabidopsis
(Zhu et al., 1998a, b) and five other species (Chapman
et al., 2003) a similar four-step pattern has been observed
F I G . 1. Root apical meristem (RAM) patterns in dicotyledonous angiosperm roots from Groot et al. (2003). (A) Linum grandiflorum (flax) has a closed organization with specific tiers of initials. (B) Daucus carota (carrot) has an intermediate-open organization and this is perhaps the ancestral condition. No specifically
apparent tiers of initials are present, but the cell files are seen in a clear lineage to the initial zone. (C) Pisum sativum ( pea) is the basic-open type found only in the
Fabaceae and the Cucurbitaceae. In this type no initials are obvious. Scale bars ¼ 50 mm.
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ROOT AP I CAL M E R IS T E M O R G AN IZ AT IO N
the intermediate-open type of RAM organization was ancestral, the basic-open type, which is found only in two families,
Fabaceae and Cucurbitaceae, is derived and the closed type is
advanced. Although these results should be considered open
for further analysis, the point being made here is that there
are specific types of RAM organization that appear throughout
dicotyledonous plant taxa and that generating these different
types will require taxon-specific cell cycle and differentiation
regulation. The fundamental differences between the types
are as follows.
Rost — Root organization and the position of control points
Root elongation rate (mm d–1)
60
2
50
40
3
1
30
20
10
0
4
25
50
75
100 125 150
Root length (mm)
175
200
*
225
F I G . 2. Primary root elongation rate curve showing an acceleration phase (1)
just after germination, a period of constant growth rate (2), followed by a
deceleration phase (3) and finally reaching a determinate length (4).
Taller RAM = fast growth rate
Height of the RAM
changes in response
to root growth rate
Shorter RAM = slow growth rate
F I G . 3. Image of the tip of a Pisum sativum (pea) root tip showing the height
of the RAM. The height of the RAM changes such that in fast growing roots
the RAM is taller and in slow growing roots the RAM is shorter. Scale bar ¼
100 mm.
Hawes (1991) and Hawes et al. (1998, 2003, 2005) made the
observation that root caps of many plants release living cells
into the rhizosphere in a programmed way. These cells,
called border cells, live in the soil environment, may divide
and secrete materials into the soil and thereby interact with
soil micro-organisms. The numbers of border cells released
experimentally was from up to 10 000 border cells in 24 h
by a cotton root to no border cells in roots of Brassica
(Hawes et al., 2003). It was this observation that led
Hamamoto et al. (2005) to ask a broader question connecting
RAM organization to root function.
Hamamoto et al. (2005) studied 19 different plant species
from different families with all three types of RAM organization. They then examined the anatomy of the root caps,
looked for the presence of border cells and tested for
living cells using the fluorescein diacetate (FDA) method
(O’Brien and McCully, 1981). This examination revealed
that roots with an open organization (e.g. Cucuminus
sativus in Fig. 5; Daucus carota in Fig. 1B) released
border cells from the surface of the root cap. FDA staining
showed that sunflower and pea border cells showed a positive reaction indicating that the border cells were living
when released (Hamamoto et al., 2005). Roots with a
closed organization (Salvia; Fig. 6) tended to release their
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(Fig. 2). Immediately after emergence the primary root shows
an accelerated growth rate, followed by a period of relatively
constant growth, then a deceleration and finally the primary
root ceases elongation at a determinate length. The precise
determinate length is a function of growth conditions, e.g.
temperature (Gladish and Rost, 1993). This is the first point:
the primary root, of at least the eight species studied, shows
a three-part growth cycle followed by reaching a determinate
length.
The next point is that the size, shape and behaviour of the
RAM does not remain constant during the growth of the
root. I will give two examples. In pea, Rost and Baum
(1988) measured the RAM at different times during the
primary root growth cycle (Fig. 3). They showed that the
height of the meristem increased during the accelerative
phase of the growth cycle, it remained relative constant in
height during the constant phase of the cycle and it became
shorter during the deceleration phase of the root growth
cycle. The RAM also became narrower and had less volume
during the growth cycle. The conclusion here is nicely stated
by DeTullio et al. (2010), ‘. . . RAM organization evolved to
become highly plastic and dynamic’. Another example of
RAM plasticity is connected to the observation that the
primary roots of the examined plants tend to reach a determinate length. In Arabidopsis (Baum et al., 2002) and in five
other species (Chapman et al., 2003) with a closed RAM
organization, the pattern of the meristem was followed over
the growth cycle of the primary root. In each case
(Blumenbachia; Fig. 4), the closed pattern of the RAM
changed to an open pattern when the primary root reached
its determinate length. Furthermore, the cells of the RAM
cease cycling and become symplasmically isolated (Zhu and
Rost, 2000). Incidentally, determinacy in root growth is not
restricted to the primary root (Shishkova et al., 2007).
Clearly there are different patterns of cells in the RAM of
dicotyledonous plants, and these patterns, which are associated
with specific taxa of plants, obviously evolved under different
conditions over time. Why are there different conserved patterns, why not just one pattern? There is not any absolute
answer to this question, but an observation by Hawes (1991)
and a subsequent study by Hamamoto et al. (2005) makes
an interesting suggestion.
1215
1216
Rost — Root organization and the position of control points
Blumenbachia
F I G . 5. Whole-mount image of a Cucuminus sativus root tip showing released
border cells. Scale bar ¼ 0.5 mm.
F I G . 6. Median longitudinal section of a Salvia splendens root with a closed
RAM and showing the outer root cap cells being released in sheets of cells.
Scale bar ¼ 0.1 mm.
border cells into the soil environment and roots with a
closed organization do not.
B
F I G . 4. Median longitudinal sections of the Blumenbachia hieronymi RAM.
(A) Just after emergence the root has a closed organization. (B) When the
root reaches its determinate length and stops elongation the organization
pattern changes to open. This figure is redone from Fig. 3G and H of
Chapman et al. (2003). Scale bar ¼ 70 mm.
surface root cap cells in sheets of cells. In Brassica these
cells did not show a positive FDA test (Hamamoto et al.,
2006). In Arabidopsis a TUNEL (terminal deoxynucleotidyl
transferase dUTP nick end labelling) reaction test showed
the surface root cap cells to be undergoing programmed
cell death prior to release of the surface root cap cells
(Fig. 7) (Zhu and Rost, 2000). The results of these studies
show that roots with an open organization release living
ROOT ORG A NI Z AT I ON AN D T H E PO S IT I ON OF
CON T ROL POI NT S
Roots have a deceptively simple body plan (Fig. 8) consisting
of cylinders of different tissues, sectors (e.g. xylem and
phloem), individual cell files, groups of cells within files and
individual cells. All of these levels of structural organization
must be considered when trying to sort out how specific
events are controlled. Let us briefly look at each level of organization (Rost, 1994; Rost and Bryant, 1996).
Tissues are aggregates of cells with a common origin (meristem) and a common collective function, and in roots they
tend to be organized into cylinders of cells and sectors
(Fig. 8). Cells within a specific cylinder or sector tend to
cycle and differentiate together as a unit. The signals
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A
Rost — Root organization and the position of control points
1217
Sectors
Cell files
Cylinders
Individual cells
Packets or modules
A
B
F I G . 7. An Arabidopsis thaliana root tip stained with propidium iodide (red
colour) and by the TUNEL reaction for programmed cell death (green
colour) photographed by confocal microscopy; the images to the left are propidium iodide only, those in the centre are TUNEL only, and the images to the
right show the two superimposed. (A) Longitudinal view; (B) transverse view,
showing a positive TUNEL reaction in dead cells on the periphery. From
Figure 4 of Zhu and Rost (2000) with permission. Scale bars ¼ 50 mm.
controlling this obviously affect all cells within the unit and
presumably move from cell to cell longitudinally, transversely
and sometimes tangentially, to the exclusion of tissues of
different identity nearby. This shows either that the signal is
unique to the cylinder, or that the nearby cells are insensitive
to its effect or are structurally ( plasmodesmically) isolated.
Juniper and Barlow (1969), Gunning (1978), Gunning and
Overall (1983) and Zhu et al. (1998a, b) examined the distribution of plasmodesmata (PD) in root tips and noted the tissue
specificity of the pattern, and that the most abundant frequency
of PD was in the transverse cell walls within files of cells. In
Arabidopsis (Zhu et al., 1998a) the frequency of PD was
lowest in the outer and inner tangential walls, possibly indicating less signalling between cylinders (Fig. 9). The identity of
the signal, possibly auxin, is still an open question (Benkova
et al., 2009; DeSmet et al., 2010).
The vascular cylinder is organized into xylem and phloem
sectors (Rost et al., 1988). Xylem and phloem tissues are
levels of organization in a root tip. Cylinders of
and the vascular cylinder; the vascular cylinder is
phloem; files of cells, files sometimes organized
finally individual cells. (Redrawn from Fig. 2.9 of
Rost, 1993).
composed of a complex of several cell types, vessel
members, tracheids, parenchyma cells, pericycle cells, fibres,
sclereids, sieve tube members, companion cells, and possibly
other cells organized into a pattern unique to each taxa. The
differentiation of the cells making up these tissues occurs in
a relatively synchronous pattern such that when there are multiple xylem sectors separated by phloem sectors the events in
the xylem of all sectors occur at the same time. As an
example, the pericycle cells in a pea root continue to divide
in a transverse plane some distance from the root tip just
outside of the three protoxylem points, while cells in the
cortex and the pericycle of adjacent phloem sectors are not
dividing (Rost et al., 1988). Evidence of this at the molecular
level can also be seen in cross-sections of pea roots where, at
about 10 mm from the root tip, only pericycle cells in the
xylem sector showed a positive signal for labelled histone
H2A mRNA (Tanimoto and Rost, 1993; Tanimoto et al., 1993).
Gunning (1982) conducted a series of studies on the organization of the Azolla fern root tip. This fern has a large apical
cell that divides acropetally to form root cap initials and
sequentially from its three basal surfaces to make a sequence
of groups of cells called merophytes. This developmental
unit, or merophyte, divides, differentiates and enlarges in a
set pattern, resulting in mature cells and tissues. During subsequent development, cell files differentiate in continuity
between merophytes, which would necessitate communication
between them. A merophyte system has been reported in other
lower vascular plants but not in the roots of higher vascular
plants. Barlow (1987) described a somewhat analogous structural unit in roots of the cortex of corn, and he called them cell
packets. Cell packets are also found in pea roots (Webster,
1980). In Arabidopsis, Wenzel and Rost (2001), and in
Trifolium repens Wenzel et al. (2001) reported on how the epidermis and peripheral root cap divide in units that they called a
module consisting of a packet of epidermal cells and a packet
of peripheral root cap cells. This will be described in detail
later.
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F I G . 8. Diagram of the
tissues, epidermis, cortex
sectored into xylem and
into packets of cells, and
1218
Rost — Root organization and the position of control points
TA B L E 1. Selected pea root (Pisum sativum) cell differentiation
level data from 21-d-old roots grown in liquid medium
Tissue
Plasmodesmal density (mm–2)
8·0–12·5
4·0–8·0
2·5–4·0
0·0–2·5
Endodermis cylinder
Pericycle cylinder
Protoxylem
Protophloem
Cortical parenchyma vacuolated
Endodermis casparian bands
Xylem lignified walls
Phloem fibres
82
82
176
229
650
3815
11 000
26 071
These data make the point that the distance from the root tip where
differentiation events occur is tissue specific. Note also that the location of
differentiation events is modulated by root growth rate: ‘With few exceptions,
levels of tissue differentiation were nearer the apex in non-aerated
(slow-growing) than in aerated (faster-growing) roots of comparable age and
genotype’ (Popham, 1955, p. 539; data extracted from table 2)
F I G . 9. Three-dimensional drawing of the plasmodesmal distribution in a
1-week-old Arabidopsis thaliana RAM. Plasmodesmal frequencies in different
tissues are grouped into four ranges and colour-coded in all cell walls. The
largest number of plasmodesmata tend to be located in the transverse cell
walls between cells in files. From Figure 5 of Zhu et al. (1998a) with
permission.
5·0
*
4·0
*
*
3·0
*
*
*
2·0
*
*
0·5
A
B
C
Distance from tip (mm)
Cell files are a basic unit of structure in the root where all
the cells in a cell file are clonally related to each other and
originate in a lineage from a tier or zone of initial cells depending on the RAM organization type. Popham (1955) in a rather
classic study examined the location of many different differentiation events in pea roots growing in hydroponic conditions
under aerated and non-aerated conditions. The roots grew
faster in aerated conditions and more slowly in non-aerated
conditions. Popham (1955) sampled roots and measured the
position of differentiation events. His noteworthy observation
is that the position of differentiation events is tissue specific.
Table 1 shows some selected data from his paper simply
making the important point that different cell files do different
things at different tissue-specific times and locations, and consequently whatever controls these processes needs to also be a
tissue-specific process.
Figure 10 illustrates this further, but for cell cycle events, in
a general way from pea roots, with diagrams of four different
cell files representing xylem pericycle, phloem pericycle, a
xylem tracheary element file and a cortical parenchyma file.
On the right side of the illustration are the approximate distances from the root body –cap boundary where each event
occurs. A note on the pericycle might be useful here. The pericycle is a unique tissue and in the xylem sector it is usually a
single cell layer located immediately outside the outermost
protoxylem tracheary element. Within the xylem sector the
pericycle is the site of the initiation of lateral root primordia.
The pericycle cells in the phloem sector are also a single
layer, but they have a unique identity and behaviour, as will
be discussed later.
Xylem pericycle cells divide in the transverse plane and
pass through several rounds of the complete cell cycle. The
cells then arrest for a time, then start to cycle again, some
A. Xylem pericycle
B. Phloem pericycle
C. Xylem tracheary elements
D. Cortical parenchyma
* G1›S›G2›M
* G1›››S
ARREST G1/G2
D
F I G . 10. Drawing of four different cell files (A, xylem pericycle; B, phloem
pericycle; C, xylem tracheary element file; D, cortical parenchyma file)
showing the control points for cell cycle regulation. The pericycle is a
unique tissue; in the xylem sector it is usually a single cell layer located
immediately outside the outermost protoxylem tracheary element, and is the
site of the initiation of lateral root primordia. The pericycle cells in the
phloem sector are also a single layer, but they have a unique identity and
behaviour.
lateral root founder cells are then created, and they then continue to cycle and divide, but in a tangential plane. These
events would involve cell cycle on/off switches, a cell cycle
counter and finally a mechanism to change the orientation of
the cell plate at specific lateral root initiation sites.
Phloem pericycle cells are located in the adjoining vascular
cylinder sector, but have a much simpler cell cycle control
path. These cells divide several times through complete cell
cycles, then arrest and never divide again.
Xylem tracheary elements are a water-conducting cell type.
They are interesting because they tend to be large cells with a
sometimes high ploidy level, they tend to have sculptured secondary cell walls and they go through programmed cell death
on the way to reaching their mature function. The procambium
primary meristem cells are progenitors of xylem tracheary
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Root cap
Protoderm/Epidermis
Ground meristem/Cortex
Ground meristem/Endodermis
Procambium/Vascular tissue
Plerome
Periblem
Dermatogen/Calyptrogen
Distance from root cap columella (mm)
Rost — Root organization and the position of control points
elements and they divide in a transverse plane some number of
times through complete cell cycles, then the G2 M part of
their cycle ceases and the cells pass through several rounds
of G1 S to reach some level of ploidy. Coupled with this,
the cells lay down their secondary cell wall and initiate programmed cell death.
The last cell file in this illustration is a cortical parenchyma
cell type. These cells tend to be fairly large and specialized for
storage, often of starch. These cells divide through a few complete cell cycles, then switch to a G1 S cycle to become
polyploidal during the period when they accumulate starch.
So, here are four different cell files, all in close proximity,
and all having different cell cycle patterns and processes.
Since PD are predominantly in the transverse cell walls, this
is the most likely pathway for signal transduction, but the identity of the signal, although most likely to be auxin, has yet to
be fully understood.
In the ‘classical view’ a root tip is considered to be organized into the following regions: the root cap, the meristem,
the elongation region and the maturation region (Rost, 1994).
These boundaries do not really exist, as we have just discussed,
and each cell file, or group of files in a cylinder or sector, tends
to act independently. The boundaries between the region of the
meristem and the region of elongation, therefore, may be
different in cells of the cortex, for example compared with
cells of the epidermis or the vascular sectors. The manner in
which this happens involves the idea of transition points
(Rost, 1994). This concept, originally described by Ivanov
(1973), suggests that within each cell file there are developmental switches. One would be the point where cell division
is turned off in a phloem pericycle cell file, which would be
one file in the procambium primary meristem. Another transition point would exist at the boundary of the elongation
region, and another at the termination of cell maturation. In
this way each file or group of files may act independently of
each other. Exactly what happens at the transition point is
not known, but perhaps specific genes are expressed, which
turn events on or off in a cell file-specific manner.
The position of transition points, as we have seen, is not static.
As the growth rate of a root changes, the location of the transition
point also changes (Popham, 1955). An example in cotton is that
as the root growth rate speeds up, the position of the maturation of
xylem vessel members becomes farther away from the root tip,
and as the root growth rate slows down the position of maturation
becomes closer to the root tip (Reinhardt and Rost, 1995).
This built-in mechanism allows the root to accommodate its
growth rate events (cell division and elongation) spatially to its
cell differentiation events. In cotton roots a linear relationship
exists between root growth rate and the position of protoxylem
maturation (Reinhardt and Rost, 1995). Fast growing roots
show protoxylem maturation farther from the root tip than do
slow growing roots.
reported on the modular structure involved in the development
of the epidermis and peripheral root cap which together are
clonally related and initiated by a specialized periclinal division of an initial cell they termed the root cap/protoderm
(RCP) initial. The idea for a modular structure for the epidermis and peripheral root cap was first put forward by Kuras
(1978) in a study of embryo development in Brassica napus,
and the basic format and developmental sequence is similar
in A. thaliana, which is in the same family, the
Brassicaceae. What follows is a description of that sequence
as another example of a regulated series of cell cycle behaviour (Wenzel and Rost, 2001).
In median longitudinal view (Figure 11) an Arabidopsis
RAM is closed with three tiers of initial cells; the upper tier
forms the procambium which differentiates into the vascular
cylinder and becomes sectored into primary xylem and
primary phloem tissues. The middle tier forms the ground meristem which differentiates into the cortical cylinder with its
inner layer the endodermis. The third tier of initials has two
components, the columella initials (CIs) which form the
layers of the columella root cap, and the RCP initials that
form the protoderm/epidermis and the peripheral root cap.
The definition of an initial is that it is a ‘meristematic’ cell
that divides to renew itself and to form another derivative
cell that differentiates into one of many different cell types.
The behaviour of these initials and the sequence of divisions
progressing from their derivative cells provides an example of
very specific cell cycle control. CIs typically consist of four
internal cells surrounded by 8 – 11 outer CI cells (Wenzel
and Rost, 2001). The divisions of the CIs is relatively synchronized to produce discrete layers of columella root cap; in the
image shown there are four layers (Figure 11). In three dimensions the RCP initials form a circle around the CIs. The RCP
initials divide in sequence around the CIs (Baum and Rost,
1996; Wenzel and Rost, 2001). The division of the RCP
initial is periclinal, forming a T-division where the ‘shaft’ of
the T is the newly formed cell wall. In Figure 11, there are
Procambium/Vascular
cylinder
Protoderm/Epidermis
Ground meristem/Cortex
T-divisions
Peripheral root cap
Root cap/Protoderm initial (RCP)
Columella initial (CI)
Columella root cap
Modular development of the epidermis and peripheral root cap
Baum and Rost (1996) reported on the structure of the
closed RAM in Arabidopsis thaliana. In that study they
F I G . 11. Median longitudinal section view of a Arabidopsis thaliana RAM.
The three T-divisions from prior divisions of the root cap/protoderm (RCP)
initials are circled. Scale bar ¼ 0.04 mm.
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Transition points
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Rost — Root organization and the position of control points
ci
A
ci
ci
a
1
ci
a
ci
a
ci
1
1
1
2
1
B
ci
a
ci
2
1
ci
a
ci
a
ci
b
a
ci
b
a
ci
b
a
2
1
2
16
2
1
C
1 2
4
3
4
8
ci
c
b
a
4
2
1
rcp
Protoderm
3
8
2
4
4
prcA
prcB
Inner ci
8
Inner cd
Outer ci
Outer cd
F I G . 12. Division steps for the development of the columella root cap from the columella initials (CIs), and the epidermis/peripheral root cap modules from the
root cap/protoderm (RCP) initials. (A) The inner CI divides prior to the outer CI. (B) Divisions of the outer CI and adjacent RCP initial which generate the three
modules are coordinated, 1 is the oldest, 3 is the youngest. (C) Axial divisions in the protoderm, prcA and prcB in the three modules. New divisions are shown
with red lines. From Figure 3 of Wenzel and Rost (2001) with permission.
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The division sequence of the modules formed by the
T-divisions of the RCP initial follows a fairly consistent
pattern. The protoderm/epidermal cell divides four times to
form 1 2 4 8 16 cells that make up the packet of
epidermal cells. This number is quite consistent. After the
full increment of 16 cells forms, the cells proceed to elongate.
The peripheral root cap cells also progress through a division
series 1 2 4 8 16 cells to form a root cap packet
of cells. The outer layer will eventually pass through programmed cell death (Zhu and Rost, 2000) and be sloughed
off as a more or less intact layer. Together, the protoderm/epidermis packet and the peripheral root cap packet make up a
module showing a very prescribed and consistent cell division
timing sequence and a cell counting mechanism. In a second
study, Wenzel et al. (2001) examined the development of the
epidermis and peripheral root cap of white clover (T. repens)
which has an open RAM organization, and it is quite noteworthy that this root also formed modules in the same
pattern even though the RAM type was different. The only
difference between Arabidopsis and Trifolium was that the
packets tended to be composed of 32 cell of epidermis and
32 cells in the peripheral root cap in the latter, showing that
three T-divisions apparent along one lineage series, shown
inside circles on the right side.
Figure 12 illustrates the sequence of divisions that occur to
form the root cap and epidermis in A. thaliana. The CIs
(Fig. 12A) divide in a transverse plane to form the next increment of the columella root cap shown by the letter ‘a’. The
adjoining RCP initial will divide in concert with the CI or
immediately after it by a periclinal T-division (Fig. 12B).
The resulting two cells will divide again, the inner one will
renew the RCP initial and form the first increment of protoderm/epidermis and the outer one forms the first two cells of
the peripheral root cap. This cluster of a protoderm/epidermal
cell and two peripheral root cap cells is the first step in
module formation, noting that both components are clonally
related and derivatives of the RCP initial. The next step in
the process is that the CI will divide again, followed by the
next division of the RCP initial so that the columella root cap
and the peripheral root cap have components added and grow
in concert with each other as shown in Fig. 12B. Figure 12B
shows three T-divisions in a lineage sequence and the three
modules which form, with number 1 as the oldest.
Rost — Root organization and the position of control points
in these roots the module had one more cell division, but it was
still a multiple of eight cells.
CON C L U S IO NS
How are these very specific genetic events controlled in a complex
tissue?
Rost and Bryant (1996) wrote a review on root organization
and gene expression where the authors tried to identify the
genes known at that time which were involved in various
root differentiation and cell cycle events. The number of
involved genes was quite large, but the details of how events
are regulated were not known. Many questions remain unanswered. How many genes are involved in differentiation regulation? Where are these genes activated and how? How do
cells ‘communicate’ in cylinders, sectors and files? What is
the nature of the communication signals? What is the role of
auxin, cytokinin and other regulators? How does positional
information regulation work in roots?
It is now left to the current and next generation of scientists
to sort out how events are controlled in roots.
ACK N OW L E DG E M E N T S
I acknowledge the partnership of many undergraduate and
graduate students, post-doctoral scholars and collaborations
over almost 40 years of research on root structure and
development. I would like to note in particular the work of
Jack Van’t Hof, Stuart Baum, Dan Gladish, Gene Tanimoto,
Maud Hinchee, Carol Wenzel, Edwin Groot, Sue Nichol,
Tong Zhu, Harold Reinhardt, Joseph Dubrovsky, Alexander
Lux, Arnold Bloom and Jim Seago, and I dedicate this paper
to everyone who has passed through my laboratory over the
years.
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1. There are three general RAM types in dicotyledonous
angiosperms: basic-open, intermediate-open and closed;
the intermediate-open is ancestral.
2. RAM size and shape change with the root growth cycle.
3. The height of a RAM correlates with the root growth rate.
4. Roots with a closed RAM organization have specific initials
to which all cells making up the root cap and body are connected by lineage. Roots with an open RAM organization
have no specifically identifiable initials cells and consequently the lineage connections for cell files are less clear.
5. Primary roots show a three-part growth pattern and become
determinate.
6. The RAM pattern changes when the root reaches its determinate length and the cells of the RAM cease cycling.
7. RAM type is related to root cap function. Roots with a
closed apical organization have a smooth outside surface;
they tend to slough off their root cap in layers as dead
cells and not as individual living border cells. Roots with
an open organization release many border cells that continue
to divide and secrete substances.
8. Cell cycle and differentiation control points are tissue
specific and their position also correlates with root growth
rate.
9. The epidermis and peripheral root cap in A. thaliana form
through a precise series of cell divisions from a RCP
initial cell forming modules of 16 epidermal cells and 16
peripheral root cap cells. A similar pattern occurs in
T. repens, a root with an open RAM organization, the difference being that 32 cells form in each packet.
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