Search

Actin filaments are visualised by means of indirect immunofluorescence in plant tissues that were fixed in formaldehyde, embedded in low-melting polyester wax and sectioned on a microtome. The technique described here avoids usage of detergents and organic solvents and is also compatible with immunolocalization of many other antigens.

Root apices represent an ideal model object for studies on plant cell growth and development. We have exploited this opportunity for detailed analysis of the actin-based cytoskeleton in cells of various root tissues throughout their cellular development. During mitosis, cells re-distribute their actin filaments (AFs) and myosin VIII molecules from the cytoplasm to the cell periphery where they accumulate at putative AF-organizing centres (AFOCs) facing the spindle poles. Postmitotic root cap columella cells differentiate first into gravity-sensing statocytes which are unique among postmitotic root cells due to the lack of any distinct cables of AFs. Later, statocytes, as well as peripheral cap cells, transform into secretory cells equipped with dense AF networks distributed throughout their cytoplasm. They retain abundant AFs after being shed from the root. Intriguingly, however, all root cap cells lack myosin VIII at their periphery. By contrast, all postmitotic cells of the root body, as they traverse the transition zone, show myosin VIII localized at their periphery. Myosin VIII localizes especially at the plasmodesmata in the non-growing cross walls. In cells of the transition zone, unique AF bundles develop which are proposed to participate in the onset of rapid cell elongation. These AF bundles are initiated at the nuclear peripheries and are organized via myosin VIII-enriched cross-walls, these two sites obviously act as the major AFOCs of postmitotic root-body cells. Treatment of roots with latrunculin B reveals that dynamic AFs are essential for both vacuome-driven cell elongation and root hair formation. In the transition zone and elongation region, cells of the inner cortex localize plant myosin VIII molecules abundantly at their pit-fields. These distinctive subcellular sites, like cross-walls and root hair apices, represent powerful AFOCs capable of organizing abundant AFs.

Root development is extremely sensitive to variations in nutrient supply, but the mechanisms are poorly understood. We have investigated the processes by which nitrate (NO 3 − ), depending on its availability and distribution, can have both positive and negative effects on the development and growth of lateral roots. When Arabidopsis roots were exposed to a locally concentrated supply of NO 3 − there was no increase in lateral root numbers within the NO 3 − -rich zone, but there was a localized 2-fold increase in the mean rate of lateral root elongation, which was attributable to a corresponding increase in the rate of cell production in the lateral root meristem. Localized applications of other N sources did not stimulate lateral root elongation, consistent with previous evidence that the NO 3 − ion is acting as a signal rather than a nutrient. The axr4 auxin-resistant mutant was insensitive to the stimulatory effect of NO 3 − , suggesting an overlap between the NO 3 − and auxin response pathways. High rates of NO 3 − supply to the roots had a systemic inhibitory effect on lateral root development that acted specifically at the stage when the laterals had just emerged from the primary root, apparently delaying final activation of the lateral root meristem. A nitrate reductase-deficient mutant showed increased sensitivity to this systemic inhibitory effect, suggesting that tissue NO 3 − levels may play a role in generating the inhibitory signal. We present a model in which root branching is modulated by opposing signals from the plant’s internal N status and the external supply of NO 3 − .

Direct contact of the radiating perinuclear microtubules (MTs) with the nuclear envelope was visualized with an immunogold technique using specific monoclonal tubulin antibody. The possibility that these perinuclear MT arrays are involved in establishing and maintaining nuclear organization during the interphase of cycling cells in maize root meristems was tested using taxol, a MT-stabilizing agent. Taxol not only stabilized all MTs against the action of the MT-disrupters colchicine and oryzalin but also prevented these agents from their usual induction of nuclear enlargement and decondensation of nuclear chromatin. On the contrary, nuclear size decreased and the chromatin became more compact in mitotically cycling cells of the taxol-treated root apices. Moreover, taxol prevented the stimulation, by colchicine and oryzalin, of the onset of the S phase in cells of the quiescent centre and proximal root meristem. Exposure of maize roots to taxol strongly decreased final cell volumes, suggesting that the more condensed nuclear chromatin is less efficient in genome expression and that this accounts for the restriction of cellular growth. All these findings support the hypothesis that MT arrays, radiating from the nuclear surface, are an essential part of an integrated plant 'cell body' consisting of nucleus and the MT cytoskeleton, and that they regulate, perhaps via their impact on chromatin condensation and activity, progress through the plant cell cycle.

This chapter focuses on formal characterization of stem cells in the context of plant root (and shoot) development. Stem-cell populations in plants show many properties common to those of animal stem-cell systems, including proliferation, which is usually at a lower rate than in other dividing somatic cells; self-maintenance; occupation of a particular anatomical site from which recognizable cell lineages originate; asymmetric division, where one daughter cell may, given certain conditions, embark on a differentiation pathway; and capacity to respond, by an activation of more rapid proliferation, to the loss or destruction of neighboring differentiating cells. Just as the regenerative response of stem cells is a property of a more integrated organ level of organization, so the property of asymmetric division is a feature more satisfactorily interpreted at the organ level than at the cellular level. A stem cell is capable of proliferation and self-maintenance, the latter condition indicating an unlimited potential for mitotic division. This chapter discusses some of the features used empirically to define or to describe stem cells. It elaborates stem branching structures and founder zones. It explains open and closed types of root meristems and also describes structured and stochastic apices.

Indirect immunofluorescence, using monoclonal antibodies to actin and tubulin, applied to sections of root tips of Lepidium, Lycopersicon, Phleum, and Zea, revealed features of the cytoskeleton that were unique to the statocytes of their root caps. Although the cortical microtubules (CMTs) lay in dense arrays against the periphery of the statocytes, these same cells showed depleted complements of endoplasmic microtubules (EMTs) and of actin microfilament (AMF) bundles, both of which are characteristic of the cytoskeleton of other post-mitotic cells in the proximal portion of the root apex. The scarcity of the usual cytosketetal components within the statocytes is considered responsible for the exclusion of the larger organelles (e.g., nucleus, plastids, ER elements) from the interior of the cell and for the absence of cytoplasmic streaming. Furthermore, the depletion of dense EMT networks and AMF bundles in statocyte cytoplasm is suggested as being closely related to the elevated cytoplasmic calcium content of these cells which, in turn, may also favour the formation of the large sedimentable amyloplasts by not permitting plastid divisions. These latter organelles are proposed to act as statoliths due to their dynamic interactions with very fine and highly unstable AMFs which enmesh the statoliths and merge into peripheral AMFs-CMTs-ER-plasma membrane complexes. Rather indirect evidence for these interactions was provided by showing enhanced rates of statolith sedimentation after chemically-induced disintegration of CMTs. All these unique properties of the root cap statocytes are supposed to effectively enhance the gravity-perceptive function of these highly specialized cells.

A plant is a complex system of branched axes, any of which may be in either a vegetative or a reproductive phase of development. Indeed, given the amazing diversity of plant life, repetitive branching of morphologically distinctive axes is probably the only true generality that can be attributed to plants (see Schultz-Schultzenstein 1861). At a more detailed level, in those axes expressing the vegetative phase, it can be seen that their structure is comprised of a set of reiterated morphological units, a structure which, moreover, tends to be recapitulated in each new axis originating from a branching event. But what exactly is the reiterated structure that is so faithfully reproduced by branching? Where do the type of roots termed “adventitious” fit into this branching scheme, and what contribution do they make to the plant’s life cycle? We shall attempt to answer these questions in the following pages but, put briefly, our thesis is that so-called “adventitious roots” comprise a range of shoot-borne roots which are members of a set of reiterated morphological units that, in turn, are integral to plant architecture. Not unexpectedly, these roots are components of a strategem of plant development that is geared to vegetative propagation and nutrient acquisition. These developmental ploys are accomplished in diverse manners, revealing the astonishingly multifarious nature of root growth and development. First, however, it is necessary to clarify what is meant by the two terms alreadyused, “shoot borne-root” and “adventitious root”. In doing so, some explanation is required of the reiterative nature of plant construction which, although more evidently of relevance to the shoot system, does nevertheless emphasise the indissoluble link between root and shoot systems in the life of the plant.

Acacia Senegal is an economically valuable tree species growing in the arid and semi- arid Sahel grassland of northern Africa. Not only is it a protein-rich forage crop, but it is also the sole source of gum arabic, a commercially valuable water-soluble polysaccharide. Natural regeneration of A. Senegal is compromised by over-grazing, fires, drought, etc., and so it has been necessary to implement re-forestation programmes in order to ward off the continual threat of desertification. In this context, it is fortunate that A Senegal has a root system that (a) stabilizes soil structure and (b) enriches the soil with symbiotically-fixed nitrogen. However, during the replanting of young trees, the main root easily becomes damaged. Although a new root system does eventually become re-established, this takes time, leaving the plantlets vulnerable to unfavourable conditions and hence making recovery of the ecosystem less certain.

Although the microtubular cytoskeleton of plant cells is important in maintaining the direction of cell growth, its natural lability can be harnessed in such a way that new growth axes are permitted. In these circumstances, the system which fabricates the cytoskeleton is presumably responsive to morphogenetic information originating from outside the cell. Spatial patterns of hormonal and metabolic signals within the tissue or organ that house the responsive cells are one possible source of this information. However, a contrasting source takes the form of biophysical information, such as the supracellular patterns of stresses and strains.

The nucleus and the microtubular cytoskeleton of eukaryotic cells appear to be structurally and functionally interrelated. Together they constitute a "cell body". One of the most important components of this body is a primary microtubule-organizing center (MTOC-I) located on or near the nuclear surface and composed of material that, in addition to constitutive centrosomal material, also comprises some nuclear matrix components. The MTOC-I shares a continuity with the mitotic spindle and, in animal cells, with the centrosome also. Secondary microtubule-organizing centers (MTOC-IIs) are a special feature of walled plant cells and are found at the plasma membrane where they organize arrays of cortical MTs that are essential for ordered cell wall synthesis and hence for cellular morphogenesis. MTOC-IIs are held to be similar in origin to the MTOC-I, but their material has been translocated to the cell periphery, perhaps by MTs organized and radiating from the MTOC-I. Many intranuclear, matrix-related components have been identified to participate in MT organization during mitosis and cytokinesis; some of them also seem to be related to the condensation and decondensation of chromatin during the mitotic chromosome cycle.

The origin and subsequent evolution of life on Earth have taken place within an environment where a 1g gravitational field is omnipresent. Living organisms, at whatever stage in their evolution, have accommodated this variable in both their structure and their function. Systems have also evolved whereby gravitational accelerations are perceived by gravisensors and these, in turn, have led to responses that give particular spatial orientations to living processes. It is proposed that, the higher the evolutionary status of an organism, the more likely it is that it will possess multiple systems for gravisensing because evolution discards little that assists fitness and hence supplements with new gravisensing systems those which already existed within evolutionarily older, less complex organisms. Moreover, in comparison with a single gravisensing system, a multiplicity of systems permits gravity to participate in a wider range of developmental programmes, such as taxes, morphisms and tropisms, through the action of different sensory mechanisms coupled to distinct signalling and response pathways. Whatever the precise mechanism of graviperception in any given set of conditions, all may transduce the g-force by means of a membrane system. Transduction may involve the endoplasmic reticulum and thence the plasma membrane.

The patterns of cell growth and division characteristic of the apex of tomato roots grown in vitro were simulated by computer using a growth tensor (GT). The GT was used to clarify the basis of the altered cell patterns found within apices of roots whose gibberellin levels had been depressed by mutation (at the GIB-1 locus) or through application of the gibberellin-biosynthesis inhibitor, 2S,3S paclobutrazol. At the pole of wild-type roots, where the cell files of the cortex converge, there are commonly only one or two tiers of cortical cells sandwiched between the pole of the stele and the cap initials. By contrast, root apices of the gib-1 mutant contain additional tiers in this region. The development of these additional tiers is suppressed when roots of the mutant are grown in the presence of gibberellic acid (GA3), but could be induced in wild-type roots when they are grown in 2S,3S paclobutrazol. The wild-type cell pattern can be simulated using the GT and by the application of appropriate rules that govern cell growth and division. The induced variations in cell pattern are interpreted as being due to displacements, within the apex, of the principal directions of growth (PDGs), which are represented, in part, by the set of periclines and anticlines seen in the cell wall network; these, in turn, are utilized in the specification of the GT. During normal (wild-type) root growth, the PDGs maintain a stable pattern and the corresponding cell pattern is also stable. However, in order to interpret the cellular behaviour found in wild-type roots grown in 2S, 3S paclobutrazol, simulation using the GT shows that, if the pattern of PDGs is destabilized and displaced distally along the root axis, the cell pattern reorganizes into that typical of gib-1 mutant roots. Conversely, the cell pattern of gib-1 roots, which reverts to wild-type upon exposure to GA3, can be simulated if the PDGs are displaced proximally to the inside of the apex whereupon the number of cortical tiers at the root pole decreases. These results suggest a link between endogenous gibberellin level and the specification of the PDGs in the growing tomato root apex. Furthermore, the evidence of cell patterns from gib-1 roots suggests that, in order to achieve stability of PDGs with concomitant stable cellular patterning, an optimal gibberellin level is necessary. In practice, this can be attained by culturing the mutant roots in medium containing 1 μM GA3.

Plant cells are assumed to embark on rapid elongation immediately after the cessation of their mitotic divisions at the proximal end of the meristem. However, a comprehensive appraisal of most of the data previously published on this matter, as well as several recently obtained findings, has convincingly revealed, at least for root cells that this belief is incorrect. For example, morphometric analysis of the maize root apex has clearly defined a distinct transitional region between the meristem and the zone of rapid cell elongation. This growth region is operationally defined as the post-mitotic isodiametric growth (PIG) region. In the middle of the cortex, this region may have a length similar to that of the meristem in steady-state growing roots. We believe that PIG is a specific phase in root cell ontogeny and that cells of the PIG region have more in common with the meristematic cells than with the rapidly elongating cells, although many of the metabolic properties of cells in the PIG region are rather unique. In this regard, certain properties of cells in the PIG region, such as the distributions of their microtubules and their sensitivity towards auxin and calcium, were found to be of crucial importance for the induction of differential cell growth patterns essential for the graviresponse of maize roots. Other data indicate that the PIG region is of considerable significance for the overall development of roots because of its characteristic plasticity under unfavourable external conditions.

The summit of roots of various plant species may be occupied by a single, rapidly proliferating tetrahedral apical cell (as in ferns), or by a multicellular and slowly proliferating quiescent centre (as in angiosperms), or by intermediate types of cellular organizations. The present paper attempts to deduce the phylogeny of these various types of cellular patterning at the root apex.

The mechanism by which the rate of cell proliferation is regulated in different regions of the root apical meristem is unknown. The cell populations comprising the root cap and meristem cycle at different rates, proliferation being particularly slow in the quiescent centre. In an attempt to detect the control points in the cell cycle of the root apical meristem of Zea mays L. (cv. LG 11), quiescent-centre cells were stimulated to synthesise DNA and to enter mitosis either by decapping or by immersing intact roots in an aqueous 3,3-dimethyl-glutaric acid buffer solution. From microdensitometric and flow-cytometric data, we conclude that, upon immersion, the G2 phase of the cell cycle of intact roots was shortened. However, when 50 μM abscisic acid (ABA) was added to the immersion buffer, parameters of the cell cycle were restored to those characteristic of intact roots held in a moist atmosphere. On the other hand, decapping of primary roots preferentially shortened the G1 phase of the cell cycle in the quiescent centre. When supplied to decapped roots, ABA reversed this effect. Therefore, in our model, applied ABA retarded the completion of the cell cycle and acted upon the exit from either the G1 or the G2 phase. Immersion of roots in buffer alone seems to trigger cells to more rapid cycling and may do so by depleting the root of some ABA-like factor.

The cells at the summit of an actively growing primary root, derived from the anti- apical pole of the embryo, are the source of all of the other cells of that organ. Some of these latter cells then generate new growth centres which initiate new roots, again with a new set of ‘source’ cells at their summit. The source cells thus comprise a minimal set of structural initials for the root. In some ferns, the structural initial is reduced to a single ‘apical’ cell which divides frequently, whereas in gymnosperms and angiosperms there are usually a number of initials which have a low division frequency. Experimentally, it has been shown that, in the latter case, cells immediately neighbouring the structural initials share their proliferative properties and, together with them, constitute a quiescent centre (QC). The slow rate of proliferation in the QC is associated with a prolonged G1 phase of the mitotic cycle which, in turn, is linked to a slow rate of cell growth. These properties, together with its special position within the root, characterize the QC as a stem-cell population, a view reinforced by the ability of QC cells to grow and proliferate more rapidly in circumstances that interfere with the proliferative behaviour of cell in the bulk of the meristem (Barlow 1978). The interrelationship of the various types of cells and their different behaviours within the root tip are summarized in Figure 1.

The term “adventitious root” is widely used to designate a root that arises either on an already lateralized root axis or at a site on the plant that is not itself a root (e.g., on a shoot or leaf) (Esau, 1953). In the latter case, the root, strictly speaking, need not be adventitious since, etymologically, this refers to a root located at an unusual site on the plant whereas such a root might be developing at a site in a way that is entirely consistent with the normal ontogenic pattern of the plant. It would be more exact to designate such a root as “shoot-borne.” This, in turn, leads to the idea that there are two types of roots: one is the shoot-borne type whose origin is self-defining, the other is the pole-borne root whose origin is from one of the poles of the embryo. According to Guedes, (Guedes et al, 1979) there is only one type of root: all roots are shoot-borne because even the embryonic root derives from the shoot-pole of the embryo. The same argument has been made for grasses (Tillich, 1977). Although this view may seem a reasonable argument from an evolutionary perspective, it is not one accepted by all morphologists. Whatever view is taken, it does seem that the term “adventitious root” can be restricted to its “true” meaning as referring to a root which develops out of the normal temporal sequence and/or at an unusual location. In most if not all cases, this would apply to a root that develops as a result of wounding and is thus evidence of a regenerative response. Adventitious roots, therefore, are simply a class of shoot-borne (or root-borne) roots developed under rather special circumstances (see the chapter by Haissig and Davis in this volume).

What is a meristem? This question is thrown into re lief by the very title of this book. Etymology should lead us to the original conception of the meristem as a zone where division takes place. But what type of division? The first reference, in 1844, to a ‘meristem’ is due to F. Unger. He not only described plant cell division, but also, using Tradescantia shoots, determined the location from which new cells were produced that directly contributed to the growth and development of the shoot. This was the era of Schleiden’s Cell Theory, and ‘meristem’ conveniently fitted into the cellular concept of plant growth. Indeed, it is the view taken in many of what are now classic plant anatomy texts. However, as is evident from many of the chapters of this book, meristem is given a wider meaning than this. It is also a region of the plant where a certain degree of tissue differentiation is taking place, or has already done so. One speaks of ‘shoot meristem’ and ‘root meristem’, for example, meaning thereby not only a zone of cell division but that the cells in this zone also express the characteristics of tissues that define shoot and root. It was from this histogenetic rather than cellular conception of the meristem that Hanstein’s Histogen Theory was developed. Thus, meristems are sites where shoot‐ and root‐forming ‘potentials’ are present, a view also supported by the observation that meristems have particular shapes indicative of their determined status as potential shoots or roots. Then, one might also regard meristems as sites from which organ architecture is elaborated. That is, meristems could be seen as sites where new meristems arise which, in due course, come to reside in new locations, such as the axil of a leaf. In other words, meristems can beget meristems. This is certainly what occurs in dichotomous branching of organs. Other types of branching, such the monopodial or sympodial types, are variants of this. These different branching patterns result in sylleptic or proleptic buds which, when they grow out, assume plagiotropic or orthotropic orientations, and so on. So, clearly, as one views the meristem at different levels of organization—cellular, organ, whole plant—more words are required to describe what a meristem is. The present book deals with all these various aspects of meristem behaviour. The purely cellular characteristics of meristems, that is, how the mechanics of cell division are managed, are expertly covered by W. Dewitte and J. Murray. As might be expected, the emphasis is on cyclin genes and how cells progress through the mitotic cycle, and what type of checkpoints limit cycle activity. Expression of cell division activity and cell differentiation nicely come together in root meristems, reviewed by P. Mylona and L. Dolan. While it may not have been on the agenda of the authors of these two last‐mentioned chapters to discuss the cytoskeleton in the context of the meristem, the reader does have to look hard here and elsewhere in the book for any discussion of how this important item of the cell brings about division and establishes the patterns of growth that give meristems their shape. The generation of shoot apices and their meristems are described in three chapters that survey the meristem in embryogenesis (M. Aida and M. Tasaka), the vegetative meristem (J. Fletcher), through to the shoot meristem’s most complex expression in floral apices (D. Zhao, Q. Yu, C. Chen and H. Ma). The remarkable dissection of pathways of gene activity in these processes is largely the result of the pivotal discovery of reverse transcriptase and restriction endonucleases. With the help of these enzymes, the analysis of meristem maintenance and the way in which tissues are derived from them has a reached a degree of sophistication which would have been unthinkable 20 years ago. Indeed, this reviewer, at least, got the impression from certain pages replete with facts derived from the literature that there are some areas of shoot meristem research where the sheer volume of data outpaces the ability to accommodate it into a coherent conceptual framework. The topic of meristems as a source of architecture is skilfully tackled by V. Grbic (axillary meristems) and B. Veit and T. Foster (leaves). These chapters, as well as the introductory overview of meristems provided by I. Sussex and N. Kerk, remind us that there is an extraordinary wealth of morphological variation in plants that has its basis in meristematic behaviour. It would be interesting indeed to see whether such a wide range of forms could all somehow be discovered in Arabidopsis thaliana. In that useful weed, there are mutations, e.g. pinhead‐1 and leafy, which display morphological features resembling those normally found in other plants, but that are normally hidden in wild type A. thaliana. Even the genus Arabidopsis itself is a source of a large amount of morphological variation, presumably indicative of genes that affect meristem behaviour having been activated or de‐activated during evolution of the genus. The chapter on phyllotaxis by D. Reinhardt and C. Kuhlemeier is an excellent account of how and where cell division and meristem division both come together in shaping the shoot apex. The authors have also taken the opportunity to provide in this chapter a fine synthesis of classic experimental and modern molecular studies, and thereby to propose a theory of phyllotaxis that brings auxin back to a central position. In so doing, it firmly pushes downstream a biophysical view of phyllotaxis that held sway for a long time (perhaps for lack of mutants at the AtPIN1 locus). Curiously, the authors contrast the stem cells of plants and animals (p. 179), saying that in the former they are involved in growth and morphogenesis, but in the latter they are restricted to the replacement of tissues. Surely, the last‐mentioned attribute is the correct one, and that, in plants, morphogenesis is largely the product of cells that are derivatives of the stem cells, as is so vividly shown in phyllotaxis itself! It is a pity that no colour illustrations have been included since a number of the black and white photographs are distinctly ‘flat’ and their reproduction and the point of their inclusion not immediately evident, especially when printed at the small size favoured here. Figure 2.1, for example, was originally published in colour, but its present reproduction in black and white robs it of the precision of the original. It was probably a wise decision not to include a separate chapter on cambium, a secondary meristem, in what is essentially a book on primary meristems. For one thing, one would quickly get bogged down in messy topics like hormonal controls and, for another, there is little (yet) in the way of genetic analysis of cambial activity, though this is likely to change now that an arabidopsis cambium and its secondary vascular tissues are becoming better known (Chaffey et al., 2002). What is pertinent to cambium is covered in Chapter 1, though it should not be taken as generally true (see p. 12) that elongated cambial fusiform cells will not develop in tissue culture: these have, in fact, been described by Washer et al. (1977) for callus nodules of Pinus radiata where they gave rise, as might be expected, to tracheids at the inside of the nodule and to phloem at the outside. In all, this is a most useful volume, but at a price that perhaps only libraries will be able to afford. Nevertheless, the present book is an essential purchase for all students and researchers wishing to discover more about how plants grow.

Several features make the root apex of maize (Zea mays L.) a good experimental system for studying cell cycle controls in relation to development in higher plants. Actively dividing meristematic cells, slowly-cycling quiescent centre cells, differentiating cap cells and senescent detaching cap cells are distinctively compartmented. In addition, the quiescent centre can be activated into rapid proliferation in response to stresses. Although there is a wealth of cytological and physiological information about the behaviour of cells in the root apex, very little is known about the molecular factors which control the patterns of cell division and differentiation. Differential screening of cDNA libraries obtained from discrete cell populations from the apex may provide a means to identify genes which are expressed in cell cycle- and differentiation-dependent manners.

Cell division in plants is one component of the process of organogenesis. In the case of roots, division can be viewed from two perspectives, one relating to its structural role in blocking out the cellularized pattern of the organ, the other emphasising its functional importance in supplying cells for growth. In neither case is division directly relevant to tissue differentiation since this probably results from positional cues superimposed on the cellularized whole; new cells created by division are, however, the units in which differentiation is accomplished. The structural aspect of division in relation to organogenesis emphasises the orientation of the new cell walls in various regions of the meristem. It also recognises two basic classes of division, the formative and the proliferative, properties of which are illustrated with examples from tomato and maize roots, respectively. Formative divisions occur in a programmed sequence which has been worked out for the cortex and the cap/dermatogen cell complexes. Programmes also govern the proliferative divisions and details are given of two of these for stele and cortex. Since the division sequences are recursive, they are amenable to analysis by means of L-systems. These afford an opportunity to formalize the portion of the epigenetic code that applies to cell patterning. At the deeper, cytological level, the code may resolve into recursive patterns of microtubule behaviour.

The levels that compose biological hierarchies each have their own energetic, spatial and temporal structure. Indeed, it is the discontinuity in energy relationships between levels, as well as the similarity of sub-systems that support them, that permits levels to be defined. In this paper, the temporal structure of living hierarchies, in particular that pertaining to Human society, is examined. Consideration is given to the period defining the lifespan of entities at each level and to a periodic event considered fundamental to the maintenance of that level. The ratio between the duration of these two periods is found to be approximately 2.5 x 10(4). A similar relationship is found when lower, non-living levels of molecules and atoms are considered. This suggests that there is a constant factor of amplification between analogous periodic events at successive levels of the Human hierarchy.

Roots of hydroponically grown maize (Zea mays cv LG11) have a greatly reduced growth rate at 5 degrees C (0.02 millimeters per hour) compared with those at 20 degrees C (1.2 millimeters per hour). Various physical parameters of roots growing at each temperature were compared. Turgor pressure of cells in the elongation zone increased from 0.59 +/- 0.05 megapascal at 20 degrees C to 0.82 +/- 0.04 megapascal after 70 hours at 5 degrees C; thus, growth rate was not limited by decreased pressure. On cooling, tissue plasticity (measured by Instron/tensiometer) decreased slowly over 70 hours. On rewarming to 20 degrees C from 5 degrees C, growth rate, turgor pressure, and tissue plasticity all returned concertedly to their original values over a period of days. However, immediately following cooling growth rate dropped rapidly from 1.8 to 0.12 millimeters per hour in 30 minutes but turgor pressure and tissue Instron plasticity remained unchanged. A plot of turgor pressure against growth rate indicated that, following cooling from 30 to 15 degrees C, the in vivo wall extensibility of the tissue was reduced by 75%. Yield threshold was unchanged. Cells whose expansion was arrested in the long-term cold treatment do not resume growth. Root growth recovers by the expansion of cells newly produced by the meristem. Cessation of extension growth is an effect on the individual expanding cell. Growth recovery is not a reverse of this effect but requires the generation of fresh cells.

The plant cytoskeleton has been implicated in a variety of morphogenetic events in higher plants. Most of this work, however, has concentrated on epidermal cells or primary tissues. We have investigated the cortical microtubular (CMT) and microfilament (MF) components of the cytoskeleton in a secondary tissue – active vascular cambium of Aesculus hippocastanum L. (horse-chestnut) – and followed the changes in these components during the early stages of differentiation of fusiform cambial derivatives to axial elements of the secondary vascular system. A correlative approach was used employing indirect immunofluorescence microscopy of α-tubulin on 6 μm sections, and transmission electron microscopy of 60 nm sections. The study has demonstrated a rearrangement of the CMT cytoskeleton, from random to helical, as fusiform vascular cambial cells begin to differentiate as secondary phloem vascular tissue. A similar CMT rearrangement is seen as fusiform cambial cells begin to differentiate as secondary xylem fibres. This rearrangement is interpreted as evidence of determination of cambial derivatives towards vascular development. Axially-oriented MF bundles are present in fusiform cambial cells and their axial orientation is retained in the vascular derivatives at early stages of their development even though the CMTs have become rearranged.

A small proportion of nuclei in root meristems ofAllium flavum, Bryonia dioica andLupinus angustifolius exhibit a dispersion of their chromocentres; the structure of such nuclei corresponds to the “Zerstaubungsstadium” (Z-phase) described byHeitz.

The dimensions of metaphase chromosomes and nuclear DNA contents were measured in eight species ofLuzula. The 2 C DNA contents ranged from 8.51 pg inL. purpurea to 0.55 pg inL. pilosa. Total chromosome volume shows a linear relationship with DNA content; however, the total chromosome length of the complement of the different species is approximately constant. Nucleolar volume and the number of chromocentres in the different species also show a relationship with DNA content. Taken together, these data suggest that while chromosome fragmentation could have generated the present-day range of chromosome numbers in the genus, there have also been changes in the total quantity of DNA with the result that species with similar chromosome numbers have different DNA contents. The relationships of DNA content with chromosome volume inLuzula and other genera are compared and the differences discussed.

The nuclear DNA content of metaxylem cells in roots of Zea mays cv. Golden Bantam reaches 16C or 32C by successive rounds of DNA endoreduplication. Each phase of endoreduplication (endo-S) is separated by a non-DNA synthetic phase (endo-G). These phases seem to occur in zones at fixed distances from the root tip. The duration of the phases in two of the endoreduplication cycles (4C–8C, 8C–16C) has been estimated in two ways. The first makes use of the rate of movement of cells through the positions along the root where the different phases of the cycle are occurring, the second uses labelling with methyl-[3H]thymidine and autoradiography. Both methods indicate that the endo-S phases which cause the nuclear DNA content to rise from 4C to 8C and from 8C to 16C last 8–10 h, and that the intervening endo-G phase lasts 8–12 h. DNA endoreduplication keeps pace with the increase of nuclear volume; cell volume increases at a more rapid rate, however. Comparison of the endoreduplication cycle in the metaxylem with the mitotic cycle in the adjoining files of parenchyma cells shows that the mitotic cells complete their cycle more slowly.

Summary Dormant meristematic cells of the unemerged radicle contained within the caryopsis of Zea mays L. are arrested in either the G1 or G2 phase of the mitotic cycle. Following germination and the resumption of root growth, each of these cells divides repeatedly to form multicellular groups, or ‘packets’. The thickened cell walls that bound each packet correspond to the walls of the formerly dormant mother cell, while the thinner walls that partition the packet correspond to walls laid down when the successive rounds of division are completed. The relative thickness of these partition walls corresponds to their age, the most recently inserted wall being the thinnest. The packets thus give evidence of not only the number of divisions that have occurred since germination, but also the sequence in which these divisions took place. In addition, the elongation of the packets during root growth allows their displacement away from the root tip into the zone beyond the margin of the meristem to be measured. Using roots fixed at different times during early growth, the kinetics of packet development has been followed in cells occupying different positions within the meristem at the start of root growth. By counting the number of cells in the packets at frequent intervals during root growth, the period between each round of division has been found to be fairly constant, even as the cells are displaced towards the margin of the meristem. Variability in the interdivisional period within a packet is insufficient to cause extensive overlapping of the different rounds of division. Exceptions are found in cortical and stelar cells around the quiescent centre, where the more distal cells in a packet often divide at up to half the rate of the more proximal cells. This is evidence of a steep gradient of cell extension rate near the quiescent centre; such a gradient does not occur along packets elsewhere in the meristem. In the quiescent centre itself, cells of its cortical portion divide more rapidly than cells of its stelar portion. Cells in the cortex (but not in the stele) often divide unequally at their first transverse division; the distal (apical) daughter is usually the longer of the two daughter cells. Asymmetric, transverse division also occurs in some cells during the next rounds of division. The more rapid entry into mitosis of the longer daughter cell results in packets with particular sequences of division. Asymmetric, longitudinal (periclinal) divisions also occur in the cortex, the inner daughter cell being wider than the outer daughter cell. These periclinal divisions occur in the distal portion of the cortex near its inner and outer borders with the stele and epidermis, respectively. At the start of root growth, periclinal divisions commence sooner in the outer cortex than in the inner cortex but do not persist, and the number of cell files across the width of the cortex declines. A concurrent loss of files also occurs in the stele. The first two rounds of periclinal divisions in the innermost file of the cortex show a definite spatial pattern. These divisions intrude into the quiescent centre and may account for the apparently anomalous faster cycling cells that have been reported here. The cellular packets give insights into certain of the fundamental aspects of root morphogenesis, the choice that confronts a cell of whether to divide transversely or longitudinally being of special importance. Particular ranges of values for the ratio between cell length and breadth are associated with these two classes of division.

The behaviour of cells within the root apex and its meristem is considered within the framework of the following propositions. 1. (1) The quiescent centre is the site of a founder cell population from which all other cells of the root can be derived. During steady-state growth of the root the founder cells progress through the mitotic cycle slowly and have an indeterminate reproductive life-span. 2. (2) Surrounding the quiescent centre are initial cells and their derivatives; both types of cells progress through the mitotic cycle quickly and the derivative cells have a determinate reproductive life-span. The number of founder cells remains approximately constant during root growth, so when such a cell divides one of its daughters, or another founder cell, must displace an initial and take on its function. 3. (3) It is supposed that in the root apex there are gradients of substances that regulate aspects of cellular and nuclear behaviour. Founder cells in the quiescent centre may be the source of a gradient of a substance (perhaps a cytokinin) which, as long as its concentration is at an appropriate level, triggers mitosis and cytokinesis. A second substance (perhaps an auxin) moves towards the quiescent centre from a source in the maturing cells of the root apex; it is supposed that its concentration is appropriate to maintain nuclear DNA synthesis in cells of the meristem and elsewhere in the apex. The relative concentrations of the two substances may regulate the rates of mitosis, cell growth and the switch from mitosis to endomitosis. 4. (4) Besides their role in maintaining cell division in the meristem, the founder cells, together with the cells immediately surrounding them, may constitute a pattern of cells that is self-perpetuating. 5. (5) Founder cells may constitute a pool of cells whose proliferative potential is not impaired by the passage of time. The propositions are predictive and possible experimental tests of, as well as evidence for, their veracity are presented.

Optical density profiles of Feulgen-stained nuclei ofBryonia dioica at different stages of the mitotic cycle were determined. Nuclei in the G2 phase have a greater fraction of dense chromatin than nuclei in G1 phase. However, nuclei at the end of the S phase have dispersed chromatin of minimal density. Thus, chromatin density oscillates during the mitotic cycle of this species, consequently the progressive increase in density previously recorded throughout the intermitotic period of two other species (onion and mouse) cannot be a general rule.

SUMMARYThe chromocentres in nuclei of germinating radicles of Bryonia dioica normally disperse before the onset of DNA synthesis. Inhibitors of DNA synthesis enhance the frequency of such disperse nuclei. Later, when the meristem of the rootlets is established, there is a good correspondence between the disperse Z phase morphology and DNA synthetic activity in such nuclei. When roots are exposed to inhibitors of DNA synthesis Z phase persists but the correspondence with DNA synthesis is broken. It is proposed that chromatin dispersion is not dependent upon DNA synthesis, and that chromatin condensation/decondensation and DNA synthesis are markers of two separate cyclical processes that occur in proliferating cells.

The growth of the nuclei and cells that constitute the hairs on the anther ofBryonia dioica has been investigated. The cell at the base of the hair reaches a volume of nearly 106 μm3 just before the flower opens, while the cells distal to it do not grow and have volumes of less than 104 μm3. The nucleus of the basal cell undergoes endopolyploidisation, doubling its DNA every 1.3 days in the early stage of this process, to reach a maximum DNA content of 256 C which is equal to 512 pg of DNA. Nuclei in the distal hair cells do not have more than a 4 C DNA content. The endopolyploid nucleus of the basal cell contains chromosome structures that are polytene. Homologous polytene chromosomes often appear to be paired.

When primary roots of Zea mays are grown in culture solution at 5°C elongation practically ceases and cell division in the meristem comes to a halt. Roots grown at 5°C for 4 days and then returned to 20°C rapidly resume elongation after a lag of 2 days. The end of the lag period coincides with the onset of nuclear DNA synthesis and mitosis throughout the meristem. Cells of the quiescent centre (QC) are also stimulated to divide but do so at a rate faster than usual. Divisions are more persistent in the cortical portion of the QC than in the stelar portion, may of whose cells tend to return to a quiescent state after one division. Descendants of cells from the cortical portion displace the original root cap and regenerate a new cap. Apices with signs of cap regeneration are most frequently seen on the fifth and sixth day of recovery and the regeneration process is complete by the eigtth day. Some cells in the meristem fail to divide and die during the recovery period causing lesions which are most evident on the third day. The dead cells are crushed by the expansion of their neighbours which, when they divide, sometimes form new cell walls with unusual orientations. All damage induced by the cold treatment is eventually cleared from the meristem as a result of the growth and division of younger meristematic cells, including those of the QC.

Tropisms and other movements of a plant organ result from alterations in local rates of cell elongation and a consequent development of a growth differential between its opposite sides. Relative elemental rates of elongation (RELELs) are useful to characterize the pattern of growth along and round an organ. We assume that the value of the RELEL at a given point is dependent on distance from the tip and that the distribution of values along the organ surface can be characterized in terms of the spread and the position of the maximum value. A computer model is described which accomodates these parameters and simulates tropic curvatures due to differential growth. Additional regulatoru functions help to return the stimulated organ to its original orientation. Particular attention is given to the simulation of root gravitropism because here not only do each of the various growth and regulatory parameters have a known biological counterpart, but some can also be given an actual quantitative value. The growth characteristics relate to the biophysical properties of cells in the elongation zone of the root, while the regulatory functions relate to aspects of the graviperception and transmission systems. We believe that, given a suitably flexible model, computer simulation is a powerful means of characterizing, in a quantitative way, the contribution of each parameter to the elongation of plant organs in general and their tropisms in particular.

Summary A search was made for strand-breaks in the DNA of nuclei in different regions of the root cap of Zea mays using terminal deoxynucleotidyl transferase and 3 H-dCTP. Quantitative autoradiography of the resulting reaction product showed that DNA strand-breaks do not accumulate as the cells pass from a proliferative state in the cap meristem to a degenerative state on the outer surface of the cap. A microdensitometric investigation of root cap nuclei with a 4 C DNA content showed that the proportion of dense chromatin is higher in the outer cap cells than it is in either the central or meristematic cells.

A brief account is given of the discovery of abscisic acid (ABA) in roots and root caps of higher plants as well as the techniques by which ABA may be demonstrated in these tissues. The remainder of the review is concerned with examining the role of ABA in the regulation of root growth. In this regard, it is well established that when ABA is supplied to roots their elongation is usually inhibited, although at low external concentrations a stimulation of growth may also be found. Fewer observations have been directed at exploring the connection between root growth and the level of naturally occurring, endogenous ABA. Nevertheless, the evidence here also suggests that ABA is an inhibitory regulator of root growth. Moreover, ABA appears to be involved in the differential growth that arises in response to a gravitational stimulus. Recent reports that deny a role for ABA in root gravitropism are considered inconclusive. The response of roots to osmotic stress and the changes in ABA levels which ensue, are summarised; so are the interrelations between ABA and other hormones, particularly auxin (e.g. indoleacetic acid); both are considered in the context of the root growth and development. Quantitative changes in auxin and ABA levels may together provide the root with a flexible means of regulating its growth.

Summary Using primary roots of Zea mays we have found that removal of the root cap causes amyloplasts to develop in cells of the quiescent centre and elsewhere in the meristem. The course of amyloplast development from a proplastid has been followed in a semi-quantitative way with the electron microscope. As a new root cap regenerates, so the starch grains diminish in the cells of the meristem and reforming quiescent centre, but persist in the new root cap.

There is a certain measure of perplexity concerning the significance of endopolyploidy. It seems that this results from a narrow frame of reference from which investigators view the phenomenon; that is, a predilection for emphasizing the specialized functional aspect of endopolyploidy as it operates in species at the present time overrides any consideration of the role that this state may play in the life of a species in its encounter with the forces of natural selection either in the past or in the future. There does not seem to be any obvious relationship between the degree of endopolyploidy that a species can exhibit and either its basic DNA content or the structure of its nucleus. The significance of endopolyploidy may reside not so much in any specialized function that the condition can support, but rather in the properties that are consequent upon the endopolyploid condition itself and which are distinct from those that apply to diploid cells. Some of the properties of the endopolyploid state, and examples of their manifestation in plants and animals, are discussed. The conclusion is that these properties have a potential that opens possibilities for new paths of development and serves as a factor upon which natural selection can operate.

Primary roots of maize (Zea mays L.) were maintained at 5°C for various periods and then placed at 20°C to allow their regrowth. The growth of both roots and the first leaves in the recovery period decreased in proportion to the period spent in the cold. The anatomy of the portion of root exposed to 5°C and of the subsequently formed root was studied. The cold-treated portion contained aborted lateral root primordia and incompletely developed metaxylem elements. The regenerated portion of root often showed an abnormal vascular system, particularly where the original root had been exposed to the cold conditions for 8 days or more. The abnormalities included the loss of xylem elements and were accompanied by a thinning of the root. In other regenerated roots the vascular patterning was quite irregular and seemed to be triggered by the death, or reduction in diameter, or metaxylem elements in the more proximal, cold-treated portion of root.

Summary The rate of RNA synthesis was investigated in three regions of the root apex of Zea mays , including the quiescent centre, using 3 H-adenosine as a precursor and autoradiography of tissue sections. The rate of synthesis in the Q.C. appears to be higher than would be expected if the RNA content of its cells double during their intermitotic period. However, microdensitometry indicates that the mean RNA content of these cells is maintained at a constant level during root growth. Estimates of the rate of halving of the radioactivity in the cells of each region give no indication that the RNA detected by autoradiography is degraded once it is made. The half-times can be accounted for by the dilution of radioactivity through the action of cell division and cell growth in each region. It seems possible that estimation of the rate of RNA synthesis, at least as it normally proceeds within the Q.C., may be distorted by the feeding of a precursor.

Summary The entire cap can be removed from the apex of the primary root of Zea mays. When this is done the nuclei of cells of the quiescent centre are stimulated to synthesize DNA and subsequently enter mitosis; the labelling and mitotic indices reach maximum values 12 and 18 h, respectively, after decapping. Division of cells of the quiescent centre and their descendants leads to the regeneration of a new cap which is completed about 3-4 days after decapping. At this time a new quiescent centre is also established. The development of a new cap is a consequence of the changes in the polarity of growth and division of cells at the apex. Quantitative data are presented of the changes in rates and planes of division of the cells involved in cap regeneration. The data suggest that cells of the quiescent centre, stimulated to divide by de-capping, complete about four mitotic cycles before a new quiescent centre forms. Experiments in which decapped roots were grown in a solution of colchicine, to induce polyploid nuclei, also showed that cells of the quiescent centre have the potential to enter four or five successive mitotic cycles as, in the quiescent centre of roots so treated, nuclei with up to 64C DNA content were found after 4 days. Mature cells of the cap of intact roots contain nuclei with an 8-16C DNA content. In regenerating caps nuclei with an 8C DNA content were frequent on the fourth day after decapping. The pattern of [3H]glucose incorporation, as judged by quantitative autoradiography, by regenerating caps was compared with that by normal caps. On the fourth day of cap regeneration the patterns were similar. These two sets of results suggest that differentiation of the regenerating cap proceeds normally at the levels of both nucleus and cytoplasm. It is suggested that the initials of the cap collumella constrain the growth of the cells that constitute the quiescent centre and this in some way slows their rate of mitosis. Removing the cap relieves the constraint to cell growth and allows cell division, thus providing the trigger for regeneration. The changes in the pattern of cells in the regenerating apex, that accompany the reorganization of a new cap and quiescent centre, are explained in terms of the stresses that the cells impose upon each other.

The ultrastructure of the hair cells on the anthers ofBryonia dioica has been examined at two stages during their development. In the mature hair the cell at its base contains extensive rafts of endoplasmic reticulum while in the distal hair cells the ER forms a tubular network. In the basal cell the mitochondria degenerate and their contents pass into the cell wall; no such degeneration of mitochondria was observed in the distal hair cells. The plastids contain starch and an unusual membrane system of tubules and crescents. During hair development the wall becomes thickened with cuticular deposists. Many of the ultra-structural features of the cell contents could be accounted for by excessive growth of the membrane systems. There was no evidence that the hair cells manufacture or export any secretion.