Effects of auxins and cytokinins on epigenetic instability of callus-propagated Kalanchoe blossfeldiana Poelln.

A main problem in the vegetative propagation of ornamental plants in vitro is the epigenetic instability of cells removed from their organized environment. With calluses of leaf explants of Kalanchoe blossfeldiana Poelln., cv. Yucatan, the role of plant growth regulators (PGRs) in the occurrence of fasciation was studied.In various combinations of auxins and cytokinins, the auxin 2,4-D (2,4-dichlorophenoxyacetic acid) gave only deformed, inseparable shoot primordia. The most rapid callus induction with regeneration of well-developed sprouts was obtained with the natural IAA (indoleacetic acid) and Z (zeatin).As a first symptom of fasciation, aberration in decussate phyllotaxis can be observed. At increasing concentrations of IAA + Z, this symptom gradually decreased but fasciation proper increased. The optimum concentration was at 1 M for both PGRs. Reduction of exposure to the PGRs from six to three weeks reduced the epigenetic instability.     

Occurrence of spontaneous shoot regeneration on leaf stipules in relation to hyperflowering response in micropropagated strawberry plantlets

Summary Stipular bud (SB) formation occurred spontaneously on “Gorella” strawberry leaf stipules during proliferation phase on Boxus medium. These buds gave rise to normal shoots on the inner median zone between the stipule tips. Some stipular buds also have been observed on other parts of adaxial surface of the stipule. Stipular bud formation took place directly on the stipule without an intermediate callus. The first stage consisted of subspherical protrusions of small cells which progressively differentiated into shoots that were able to proliferate and to root. Scanning electron microscopy was used to examine the developmental stages of this adventitious organogenesis and to describe morphologic abnormalities, e.g., multiapex formation and fasciation. In vitro cloning of plantlets was made from axillary and SBs produced by the same tufts. The greater flowering abundance of stipular plants resulted in a drastic reduction of the commercial production and the fruit caliber when compared to axillary plants. However, the general phenotype of the mother cultivar (Gorella) was not affected in either case.     

Strategy for shoot meristem proliferation in plants

Shoot apical meristem (SAM) of plants harbors stem cells capable of generating the aerial tissues including reproductive organs. Therefore, it is very important for plants to control SAM proliferation and its density as a survival strategy. The SAM is regulated by the dynamics of a specific gene network, such as the WUS-CLV interaction of A. thaliana. By using a mathematical model, we previously proposed six possible SAM patterns in terms of the manner and frequency of stem cell proliferation. Two of these SAM patterns are predicted to generate either dichotomous or axillary shoot branch. Dichotomous shoot branches caused by this mechanism are characteristic of the earliest vascular plants, such as Cooksonia and Rhynia, but are observed in only a small minority of plant species of the present day. On the other hand, axillary branches are observed in the majority of plant species and are induced by a different dynamics of the feedback regulation between auxin and the asymmetric distribution of PIN auxin efflux carriers. During evolution, some plants may have adopted this auxin-PIN system to more strictly control SAM proliferation.     

A comparative histological study between normal and fasciated shoots of <Emphasis Type="Italic">Prunus avium</Emphasis> generated <Emphasis Type="Italic">in vitro</Emphasis>

A combination of Murashige and Skoogs medium and N⁶–benzyladenine (BA) at various concentrations (0, 0.1, 0.25, 0.5, 0.75, 1.0, 1.25 and 1.5 mg l^–1) was supplied to shoot tips from root cuttings of a 50-year-old wild-cherry tree (Prunus avium). The concentration of BA in the growing medium was a determining factor with respect to the number of proliferated shoots per explant in vitro.Normal and fasciated shoots were generated when BA was present at 0.5, 0.75, 1.0 and 1.25 mg l^–1 in the medium and the mean numbers of normal shoots per explant were 3.63, 5.37, 8.93 and 7.30 respectively, and those of the fasciated shoots per explant were 0.03, 0.1, 0.47 and 0.4 respectively. Anatomical analysis by confocal microscopy of sections of paraffin-embedded specimens revealed that the cell structure and organization of the cortex and vascular cylinder in the fasciated shoots was similar to that in normal shoots. However, the cross-sectional area of stem of the fasciations was apparently greater than that of the normal shoots. In particular, the volume of vascular tissues, of pith and of some individual parenchyma cells in the cortex and pith was apparently greater in fasciated shoots than in normal shoots. Increases in cytokinesis and morphogenetic activity, such as the development of callus-like regions and the formation of adventitious shoots, were observed in the cortex and pith throughout the fasciations. The fasciated shoots had numerous buds and initiating new shoots at their apices while normal shoots had a single dominant axial bud.     

The Control of Lateral Root Development in Cultured Pea Seedlings. II. Root Fasciation Induced by Auxin Inhibitors

Lateral root development in cultured seedlings of Pisum sativum (cv. Alaska) was modified by the application of auxin transport inhibitors or antagonists. When applied either to replace the root tip or beneath the cotyledonary node, two auxin transport inhibitors, 2,3,5-triiodobenzoic acid (TIBA) and 3,3a-dihydro-2-(p-methoxyphenyl)-8H-pyrazolo[5,1-α]isoindol-8-one (DPX-1840), increased cell division activity opposite the protoxylem poles. This resulted in the formation of masses of cells, which we are calling root primordial masses (RPMs), 2 to 3 days after treatment. RPMs differed from lateral root primordia in that they lacked apical organization. Some roots however developed both RPMs and lateral roots indicating that both structures were similar in terms of the timing and location of cell division in the pericycle and endodermis leading to their initiation. Removal of the auxin transport inhibitors allowed many of the RPMs to organize later into lateral root primordia and to emerge in clusters. When the auxin, indoleacetic acid (IAA) was added to the growth medium along with DPX-1840, 3 ranks of RPMs now in the form of fasciated lateral roots emerged from the primary root. The auxin antagonist, p-chlorophenoxy-isobutyric acid (PCIB), also induced RPM formation. In contrast to DPX-1840 treatment, the addition of IAA during PCIB treatment caused normal lateral root development.     

Fasciation in pea: Basic principles of morphogenesis

A study of fasciated pea Pisum sativum L. (Fabaceae) mutant Shtambovy in comparison with the wild type (Nemchinovsky cultivar) has shown that fasciation is a result of abnormal cohesion of axial or other structures which arise in a superfluous amount due to uncontrolled meristic processes. In some cases, the organs with the same number and position as in the wild type can be fascinated. Subsequent defasciation and some features of tissue differentiation suggest that the meristem of a fasciated shoot retains a certain degree of discreteness which reflects its complex structure. The number and position of leaves in a node is a function of the diameter of the leaf primordium inhibitory zone, size of the shoot apical meristem, and number of bundles in a shoot. In the absence of the apex proliferative activity combined with the reduction of phyllomes in the upper nodes, abnormal cohesion of the second order axes, racemes, can take place. As a result, inflorescences of special type develop.     

A novel Arabidopsis gene TONSOKU is required for proper cell arrangement in root and shoot apical meristems

Root apical meristem (RAM) and shoot apical meristem (SAM) are vital for the correct development of the plant. The direction, frequency, and timing of cell division must be tightly controlled in meristems. Here, we isolated new Arabidopsis mutants with shorter roots and fasciated stems. In the tonsoku (tsk) mutant, disorganized RAM and SAM formation resulted from the frequent loss of proper alignment of the cell division plane. Irregular cell division also occurred in the tsk embryo, and the size of cells in meristems and embryo in tsk mutant was larger than in the wild type. In the enlarged SAM of the tsk mutant, multiple centers of cells expressing WUSCHEL (WUS) were observed. In addition, expression of SCARECROW (SCR) in the quiescent center (QC) disappeared in the disorganized RAM of tsk mutant. These results suggest that disorganized cell arrangements in the tsk mutants result in disturbed positional information required for the determination of cell identity. The TSK gene was found to encode a protein with 1311 amino acids that possesses two types of protein-protein interaction motif, leucine-glycine-asparagine (LGN) repeats and leucine-rich repeats (LRRs). LGN repeats are present in animal proteins involved in asymmetric cell division, suggesting the possible involvement of TSK in cytokinesis. On the other hand, the localization of the TSK-GFP (green fluorescent protein) fusion protein in nuclei of tobacco BY-2 cells and phenotypic similarity of tsk mutants to other fasciated mutants suggest that the tsk mutation may cause disorganized cell arrangements through defects in genome maintenance.     

Flower morphology and relationships of Schefflera subintegra (Araliaceae,Apiales): an evolutionary step towards extreme floral polymery

Gross morphology and the development of flowers in Schefflera subintegra (Araliaceae) are examined. The floral groundplan of this species is found to be very similar to that of Tupidanthus calyptratus representing a case of most extreme floral polymery within Araliaceae. Schefflera subintegra differs from T. calyptratus with respect to a lower floral merism (19–43 versus 60–172 stamens and 15–33 versus 60–138 carpels respectively) and by transformation from polysymmetry to disymmetry of flower in the course of its development. Close relationships between S. subintegra, T. calyptratus, and Schefflera hemiepiphytica have been confirmed by phylogenetic analysis based on nuclear ribosomal internal transcribed spacer sequences. These species form a subclade within the Asian Schefflera clade, with T. calyptratus as a sister taxon to two other species. Apart from more or less pronounced floral polymery, the species of this subclade share calyx and corolla without any traits of individual sepals and petals, and also a massive calyptra. As these data suggest, the extremely polymerous flowers of Tupidanthus apparently evolved in two steps: (1) the saltational multiplication of floral elements together with a loss of individuality of sepals in the calyx and petals in the corolla and (2) further polymerization of androecium and gynoecium. Mutation(s) in CLAVATA‐like gene(s) are suggested as a possible mechanism of the saltation event. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 553–597.     

Identification of two new phylogenetically distant phytoplasmas from Senna surattensis plants exhibiting stem fasciation and shoot proliferation symptoms

Sunshine trees (Senna surattensis) exhibiting unusual stem fasciation symptoms were observed in Yunnan, China. Morphological abnormalities of the affected plants included enlargement and flattening of stems and excessive proliferation of shoots. An electron microscopic investigation revealed presence of single membrane bound mycoplasma‐like bodies in sieve elements of symptomatic plants. With DNA templates extracted from diseased plants and phytoplasma universal primers P1/P7 and P1A/R16S‐SR, nested polymerase chain reactions produced amplicons of 1.5 kb. Subsequent restriction fragment polymorphism and nucleotide sequence analyses of the amplicons indicated that the diseased plants were infected by distinct phytoplasmas affiliated with two phylogenetically distant taxa classified in two 16Sr groups (16SrXII and 16SrV). This is the first report that sunshine tree is a natural host of two evolutionarily divergent phytoplasmas and the first report that a ‘Candidatus Phytoplasma australiense’‐related strain is present in China. The findings signal a significant expansion of both geographical distribution and host range of 16SrXII and 16SrV phytoplasmas.     

Flower structure and development in Tupidanthus calyptratus (Araliaceae): an extreme case of polymery among asterids

Flowers of Tupidanthus show an extreme case of floral polymery among asterids. Floral development and gynoecium structure have been examined. The floral meristem has a complex folded shape. The tiny calyx is initiated as a continuous ring primordium. The corolla is initiated as a lobed ring and develops into a calyptra. All stamen primordia appear simultaneously as a single whorl. The carpels, also in a single whorl, tend to alternate with the stamens. Some Schefflera species related to Tupidanthus are also studied. The flower of Tupidanthus is interpreted as a result of fasciation. Further investigation should determine whether mutation(s) in gene(s) of the CLAVATA family are responsible for the fasciation here. The significance of Tupidanthus for understanding spatial pattern formation in flowers of Araliaceae, and both functional and developmental constraints in angiosperm flowers with a single polymerous carpel whorl are discussed.