β-glucuronidase as a marker for clonal analysis of tomato lateral roots

In higher plants, the root-shoot axis established during embryogenesis is extended and modified by the development of primary and lateral apical meristems. While the structure of several shoot apical meristems has been deduced by combining histological studies with clonal analysis, the application of this approach to root apical meristems has been limited by a lack of visible genetic markers. We have tested the feasibility of using a synthetic gene consisting of the maize transposable elementActivator (Ac) inserted between a 35S CaMV promoter and the coding region of a -glucuronidase (GUS) reporter gene as a means of marking cell lineages in roots. The GUS gene was activated in individual cells byAc excision, and the resulting sectors of GUS-expressing cells were detected with the histochemical stain X-Gluc. Sectors in lateral roots originated from bothAc excision in meristematic cells and from parent root sectors that bisect the founder cell population for the lateral root initial. Analysis of root tip sectors confirmed that the root cap, and root proper have separate initials. Large sectors in the body of the lateral root encompassed both cortex and vascular tissues. The number of primary initial cells predicted from the size and arrangement of the sectors observed ranged from two to four and appeared to vary between roots. We conclude that transposon-based clonal analysis using GUS expression as a genetic marker is an effective approach for deducing the functional organization of root apical meristems.     

The Emergence and Early Growth of the Lateral Root in Vicia faba L.

The duration of the mitotic cycle, as well as the proportionof cells with long and short cycle times and quiescent cells,have been investigated in the apical meristems of young lateralroots of Vicia faba. No changes took place in the duration ofC or in the phases of the mitotic cycle as the lateral rootemerged from the primary root, though the proportion of proliferatingcells increased and the quiescent fraction of cells decreased.It is suggested that the low frequency with which newly emergedlateral roots label with ³H-TdR is a result of the formationof a large endogenous pool of TdR in the meristems during theperiod they are temporarily quiescent. The changes which tookplace in the parameters of cell proliferation during the earlygrowth of the lateral root have been correlated with those inroot apical meristems following the onset of seed germination.     

The Response of Root Meristems to Colchicine and Indol-3yl-acetic acid in Vicia faba L.

The effects of colchicine and IAA treatments on mitotic activityin various root proliferating tissues have been determined.Lateral root primordia were not affected by IAA, though 24 hfollowing treatment mitotic activity was severely inhibitedin the apical meristems of 1-cm-long attached lateral rootsand primary roots. Primordia were also less sensitive to colchicinetreatment than root apical meristems. Thus telophase figureswere present in the former meristems 3 h following treatment,but not in the latter. Primordia and apical meristems respondedto the same extent, however, to the colchicine-induced increasein number of cells in metaphase, anaphase, and telophase, 3h after treatment began. The apparent difference between largeprimordia and root apical meristems in this respect was dueto the failure of colchicine to penetrate the cells of the formerproliferating tissues as rapidly as the latter. IAA was foundto prevent the increased MI found 24 h following colchicinetreatment only in those meristems where IAA inhibited mitoticactivity at this time. IAA treatments, either alone or withcolchicine, were also found to maintain mitotic activity in1-cm-long lateral roots which were excised from the primaryroots 24 h previously. In such laterals which were not treatedwith IAA, MI was zero at 24 h. It is concluded from the datareported in this paper that, during the development of rootapical meristems, changes take place in the response of cellsto factors affecting mitotic activity.     

Localization of naphthol AS-B1 phosphatase activity in lateral and main root meristems of pea and corn

Summary High levels of naphthol AS-B 1 phosphatase activity have been localized in the main and lateral root meristems of pea and corn. The highest activity in the primordia of each species occurs in the root cap and epidermis although all cells of the meristem show activity. Activity in the main root meristem is compared with that in the lateral root and it is concluded that this enzyme does not facilitate penetration of the lateral root through the parent cortex but rather that it is involved in the differentiation of the lateral root itself.     

The response of apical meristems of primary roots of Vicia faba L. to colchicine treatments

The effects of 0.5% and 0.025% solutions of colchicine on the passage of cells through the mitotic cycle in apical meristems of primary roots of Vicia faba have been examined. Both treatments affected cell progression through the mitotic cycle in the same way: S and G1 were shorter, and G2 and mitosis longer, than the corresponding control values. The duration of the various phases of the mitotic cycle were similar to those reported previously for apical meristems of lateral roots though cycle time itself was longer. Recovery of root proliferating tissues from colchicine-induced inhibition of growth is correlated with the presence of quiescent cells. Meristems which have no quiescent cells do not recover from eolchicine treatment, while meristems which contain many quiescent cells recover faster than those which contain few. The growth fraction and the proportion of proliferating cells with a short cycle time are linearly related to the duration of the S period in root meristems.     

Organization of the root apical meristem in angiosperms

Although flowers, leaves, and stems of the angiosperms have understandably received more attention than roots, the growing root tips, or root apical meristems (RAMs), are organs that could provide insight into angiosperm evolution. We studied RAM organization across a broad spectrum of angiosperms (45 orders and 132 families of basal angiosperms, monocots, and eudicots) to characterize angiosperm RAMs and cortex development related to RAMs. Types of RAM organization in root tips of flowering plants include open RAMs without boundaries between some tissues in the growing tip and closed RAMs with distinct boundaries between apical regions. Epidermis origin is associated with the cortex in some basal angiosperms and monocots and with the lateral rootcap in eudicots and other basal angiosperms. In most angiosperm RAMs, initials for the central region of the rootcap, or columella, are distinct from the lateral rootcap and its initials. Slightly more angiosperm families have exclusively closed RAMs than exclusively open RAMs, but many families have representatives with both open and closed RAMs. Root tips with open RAMs are generally found in angiosperm families considered sister to other families; certain open RAMs may be ancestral in angiosperms.     

Between Xylem and Phloem: The Genetic Control of Cambial Activity in Plants

Abstract: Post-embryonic development is controlled by two types of meristems: apical and lateral. There has been considerable progress recently in understanding the function of root and shoot apical meristems at the molecular level. Knowledge of analogous processes in the lateral, or secondary, meristems, i.e. the vascular cambium or cork cambium, is, however, rudimentary. This is despite the fact that much of the diversity in the plant kingdom is based on the differential functions of these meristems, emphasizing the importance of lateral meristems in the development of different plant forms. The vascular cambium is particularly important for woody plants, but it also plays an important role during the development of various herbaceous species, such as Arabidopsis thaliana. In this review, we focus on the two basic functions of cambial activity: cell proliferation and pattern formation.     

Effect of mutations in the PINHEAD gene of Arabidopsis on the formation of shoot apical meristems

The primary shoot apical meristem of angiosperm plants is formed during embryogenesis. Lateral shoot apical meristems arise postembryonically in the axils of leaves. Recessive mutations at the PINHEAD locus of Arabidopsis interfere with the ability of both the primary shoot apical meristem as well as lateral shoot apical meristems to form. However, adventitious shoot apical meristems can form in pinhead mutant seedlings from the axils of the cotyledons and also from cultred root explants. In this report, the phenotype of pinhead mutants is described, and a hypothesis for the role of the wild-type PINHEAD gene product in shoot meristem initiation is presented. © 1995 Wiley-Liss, Inc.     

Stem cell function during plant vascular development

While many regulatory mechanisms controlling the development and function of root and shoot apical meristems have been revealed, our knowledge of similar processes in lateral meristems, including the vascular cambium, is still limited. Our understanding of even the anatomy and development of lateral meristems (procambium or vascular cambium) is still relatively incomplete, let alone their genetic regulation. Research into this particular tissue type has been mostly hindered by a lack of suitable molecular markers, as well as the fact that thus far very few mutants affecting plant secondary development have been described. The development of suitable molecular markers is a high priority in order to help define the anatomy, especially the location and identity of cambial stem cells and the developmental phases and molecular regulatory mechanisms of the cambial zone. To date, most of the advances have been obtained by studying the role of the major plant hormones in vascular development. Thus far auxin, cytokinin, gibberellin and ethylene have been implicated in regulating the maintenance and activity of cambial stem cells; the most logical question in research would be how these hormones interact during the various phases of cambial development.     

The Arabidopsis D-type cyclin CYCD2;1 and the inhibitor ICK2/KRP2 modulate auxin-induced lateral root formation

The integration of cell division in root growth and development requires mediation of developmental and physiological signals through regulation of cyclin-dependent kinase activity. Cells within the pericycle form de novo lateral root meristems, and D-type cyclins (CYCD), as regulators of the G₁-to-S phase cell cycle transition, are anticipated to play a role. Here, we show that the D-type cyclin protein CYCD2;1 is nuclear in Arabidopsis thaliana root cells, with the highest concentration in apical and lateral meristems. Loss of CYCD2;1 has a marginal effect on unstimulated lateral root density, but CYCD2;1 is rate-limiting for the response to low levels of exogenous auxin. However, while CYCD2;1 expression requires sucrose, it does not respond to auxin. The protein Inhibitor-Interactor of CDK/Kip Related Protein2 (ICK2/KRP2), which interacts with CYCD2;1, inhibits lateral root formation, and ick2/krp2 mutants show increased lateral root density. ICK2/KRP2 can modulate the nuclear levels of CYCD2;1, and since auxin reduces ICK2/KRP2 protein levels, it affects both activity and cellular distribution of CYCD2;1. Hence, as ICK2/KRP2 levels decrease, the increase in lateral root density depends on CYCD2;1, irrespective of ICK2/CYCD2;1 nuclear localization. We propose that ICK2/KRP2 restrains root ramification by maintaining CYCD2;1 inactive and that this modulates pericycle responses to auxin fluctuations.     

NUCLEAR DNA CONTENT OF EMBRYONIC RADICLES AND CULTURED STATIONARY PHASE ROOT TIPS OF PEA (PISUM SATIVUM L.)

Cells in mature embryos and stationary phase (SP) root meristems of pea arrest in G1 and G2 of the cell cycle. The patterns of distribution of G2 nuclei in radicles and SP meristems, with and without G2 factor, were compared by using cytophotometric analysis of the relative amount of DNA/nucleus in sectioned material. Radicles and SP meristems were each divided into 5 zones and the ratio of G1 to G2 nuclei was determined for each zone. The G2 population in the radicle is restricted mainly to the embryonic cortex. A small part of the G2 population was located in the central cylinder and the root cap. In SP meristems without G2 factor, the pattern of distribution of G2 cells was similar to that in radicles. SP meristems with G2 factor contained G2 arrested nuclei in all regions of the root tip. In each region the percentage of G2 nuclei was higher than that in the same region of SP meristems without G2 factor. This indicates that the population of cells that responds to G2 factor is distributed throughout the root tip.     

Hormone signaling in plant development

Hormone signaling plays diverse and critical roles during plant development. In particular, hormone interactions regulate meristem function and therefore control formation of all organs in the plant. Recent advances have dissected commonalities and differences in the interaction of auxin and cytokinin in the regulation of shoot and root apical meristem function. In addition, brassinosteroid hormones have recently been discovered to regulate root apical meristem size. Further insights have also been made into our understanding of the mechanism of crosstalk among auxin, cytokinin, and strigolactone in axillary meristems.     

Regeneration of whole plants from apical meristems of Pinus radiata

A methodology to regenerate whole plants of Pinus radiata from apical meristems of 3- and 7-year-old trees was developed. Meristematic domes with two or three leaf primordia were excised from surface-sterilized branch tips of field-grown plants and cultured in LP medium with half strength macronutrients (1/2 LP) and full strength micronutrients. The early growth of meristems required approximately 12 weeks, including a recovery stage during the first 2 weeks. After 8 weeks, some meristems developed abnormal phenotypes and died during the subsequent stages of development. However, healthy meristems elongated and formed shoots when they were transferred to LP medium supplemented with MS vitamins, 30 mg l^–1 casein hydrolysate, and 0.4 g l^–1 agar plus 2.85 g l^–1 Gelrite. Meristems that developed vigorous shoots were used for rooting experiments when they were 2 cm in length. Whole plants were obtained after 5 days of root induction in water-agar medium containing 8.2 M IBA and 5.4 M NAA and 1 month culture in LP medium with 10 g l^–1 sucrose. Plants regenerated from meristems were further propagated by rooting of cuttings. Of the rooted cuttings, 10% were morphologically juvenile.     

The effect of Lead on early stages ofPhaseolus vulgaris L. growthin vitro conditions

Lead chloride (10^-5 M) inhibited the growth of the main root, the duration of development, the number and growth of lateral roots, primary and trifoliate leaves, and also the mitotic index in root apical meristems. Lead strongly inhibited root growth rate, mainly by reducing the number of dividing cells. Other mechanisms of this inhibition are discussed.     

Initial proliferation of cortical cells in the formation of root nodules in Pisum sativum L.

Summary Root nodule initiation in Pisum sativum begins with cell divisions in the inner cortex at some distance from the advancing infection thread. After penetrating almost the entire cortex, the branches of the thread infiltrate the meristematic area previously initiated in the inner cortical cells. These cells are soon invaded by bacteria released from the infection thread and subsequently differentiate into non-dividing, bacteriod-containing cells. As the initial meristematic centre in the inner cortex is thus lost to bacteroid formation, new meristematic activity is initiated in neighbouring cortical cells. As development proceeds, more cortical layers contribute to the nodule, with the peripheral layer and apical meristem of the nodule not invaded by bacteria.Lateral root primordia are initiated in a region separate from that in which nodules are formed, with the lateral primordia being closer to the root apex. This is interpreted to indicate that the physiological basis for nodule initiation is distinct from that for initiation of lateral roots. The role of a single tetraploid cell in nodule initiation is refuted, as is the existence of incipient meristematic foci in the root. It is suggested that the tetraploid cells in nodule meristems arise from pre-existing endoreduplicated cells, or by the induction of endoreduplication in diploid cortical cells by Rhizobium.     

Roles for Class III HD-Zip and KANADI genes in Arabidopsis root development

Meristems within the plant body differ in their structure and the patterns and identities of organs they produce. Despite these differences, it is becoming apparent that shoot and root apical and vascular meristems share significant gene expression patterns. Class III HD-Zip genes are required for the formation of a functional shoot apical meristem. In addition, Class III HD-Zip and KANADI genes function in patterning lateral organs and vascular bundles produced from the shoot apical and vascular meristems, respectively. We utilize both gain- and loss-of-function mutants and gene expression patterns to analyze the function of Class III HD-Zip and KANADI genes in Arabidopsis roots. Here we show that both Class III HD-Zip and KANADI genes play roles in the ontogeny of lateral roots and suggest that Class III HD-Zip gene activity is required for meristematic activity in the pericycle analogous to its requirement in the shoot apical meristem.     

Growth Substances in the Roots of Vicia faba II. THE EFFECTS OF AGEING AND EXCISION OF THE MAIN TAP-ROOT MERISTEM

A quantitative chromatographic study has been made of the changesin the activity and distribution of the main ether-soluble acidauxins of Vicia faba seedling root systems during developmentand resulting from excision of the main tap-root meristem. AnIAA-like auxin (AP (ii)) is apparently synthesized predominantlyin apical and at a lower rate in lateral meristems. Productionseems to stop when meristematic growth stops. Its concentrationin mature extended cells is much lower and may fall to zeroin old cells, suggesting active degradation by an auxin-oxidase.Excision of the main tap-root tip gradually results in a greatlyaugmented production of AP (ii) in lateral meristems, conceivablythe result of correlative growth promotion. A second auxin and root-growth inhibitor (AP (iii)) is presentat higher activity levels than AP (ii) in tap-root meristemsand at the same level in lateral meristems. In mature cellsits activity is much lower than that of AP (ii). In contrastto AP (ii) it accumulates in both tap-root meristems and maturetissue as the root system ages. It could also be produced duringmeristematic growth but is not subsequently degraded. As withAP (ii), excision of the tap-root tip brings about a great increasein its concentration in lateral tips. A third auxin and root-growth accelerator (AP (i)) (accelerator?) is present in lower concentrations in both meristem and maturetissue. Its concentration tends to decrease with ageing and,in lateral meristems, is not affected by tap-root tip excision. It is suggested that AP (ii) produced by the meristem is normallyat suboptimal levels in the extending cells and may be the principalhormone controlling extension growth. AP (iii) accumulationmay account for growth deceleration on ageing. The role of AP(i) remains obscure. It is unlikely that correlative effectsof the taproot tip on lateral root growth are exercised directlyvia these auxins.     

Anatomy and development of the fern sporophyte

Recent research on the developmental anatomy and morphology of the fern sporophyte is reviewed. Detailed histological and experimental studies of the organization of the fern shoot apical meristem have reconfirmed the recently controversial role of the shoot apical cell as the single apical initial of the meristem. The shoot apical meristem is nevertheless an anatomically and functionally complex structure with a strongly zoned cytohistological organization. Fern shoot apex organization can be compared with that of seed plants. The control of leaf initiation and phyllotaxy remains poorly understood. Studies differ as to whether leaf initiation in ferns involves one leaf mother cell or a multicellular region of the shoot apex. The concept of non-appendicular fronds is refuted for living ferns. The later developmental changes in the determinate leaf apical and marginal meristems of the leaf primordium form an area that is still largely unexplored but could be investigated by methods similar to those used to study shoot and root apices. Branching in ferns is morphologiclaly and developmentally diverse. There is apparently more than one developmental mode of dichotomous branching, and several modes of lateral bud formation have been described, including the phyllogenous initiation of branches at the base of leaf primordia. Developmental changes in bud meristems related to apical dominance, inhibition, and bud activation is another major area for continued study. The traditional concept of the role of the root apical cell has been reestablished by studies similar to those made of the shoot apex. Detailed ultrastructural investigations of the root ofAzolla have given a sophisticated new picture of developmental processes in that organ. Fern roots show remarkably precise patterns of histogenesis in relation to apical segmentation. The formation of secondary vascular tissue inBotrychium suggests that the Ophioglossales may be related to the seed plants. The causal relationship of leaf (and branch and root) formation and the initiation of vascular tissue in the shoot needs more study. Although still poorly understood, protoxylem systems in ferns are variable and may have morphological and systematic significance. Recent investigations of hydraulic conductance in fern stems have found possible correlations of conductance levels with growth forms. The anatomical diversity of ferns makes comparative functional anatomy a promising field for future study.     

Histochemical localization of β-glycosidases in roots ofZea mays

Summary With 6-bromo-2-naphthyl--D-glucoside as substrate, -glucosidase activity has been histochemically localized in the lateral root meristems ofZea mays. Enzyme activity is highest in the epidermis and root cap of the young primordium. In contrast, enzyme activity is low in the parent root cortex, including the damaged areas around the lateral root. The results indicate that -glucosidase is involved in the development of the lateral root rather than its penetration through the cortex.