Marina Ageeva

The snap point : a transition point in Linum usitatissimum bast fiber development

The developing stem of fibre flax (Linum usitatissimum L.) contains a specific region called the ‘snap point’, where the fiber-enriched bast tissues considerably change their mechanical properties. The snap point was found to be present during a restricted period of plant development */the fast growth phase, and to disappear when stem growth was completed. To relate this snap point to bast fiber formation stages, the number of bast fiber cells and the thickness of their cell walls were followed on the stained cross-sections of the flax stem throughout plant development, using the progressing snap point as the reference. The snap point was shown to be the spot, above which the elongation of bast fiber cells is fully completed. This fast growth stage is the period when the maximum length of all bast fibers in the mature plant (a major characteristic of flax fiber quality) is fixed and would not be changed later. Autoradiography was used to visualize the mode of flax bast fiber elongation above the snap point. The even distribution of label was indicative for surface (diffusive) growth type. Elongation of individual fiber cells was estimated to take only 2 � /4 days with a rate of 1 � /2 cm per day, while cell wall thickening occurs mainly below snap point and lasts around 2 months. The special cell wall structural order, characteristic for mature bast fibers, first appeared at the snap point in the outer layer of the secondary cell wall. Schemes are included, illustrating the course of cell wall thickening and the localization of various stages of fiber formation on the stem of growing flax plant. The established exact localization and duration of flax bast fiber formation stages, and the existence of snap point as the manually identified morphological reference for the transition, permit to separate the bast fibers at different stages of development and make flax an attractive model system to study the functional genomics of fiber formation in technical crops. # 2003 Elsevier B.V. All rights reserved.

Specific type of secondary cell wall formed by plant fibers

The review sums data indicating that, in many plant fibers, the secondary cell wall contains so-called gelatinous layers of peculiar structure along with those of common (xylan) structure. Sometimes these gelatinous layers comprise the main bulk of the cell wall. Key characteristics of gelatinous cell wall are presented and compared with those of classic xylan-type cell wall. The process of gelatinous cell wall formation is considered in detail for flax phloem fibers; several characteristic features of this process were revealed: intense rearrangement of already deposited cell-wall layers, unusual dynamics of Golgi vesicles, the occurrence of the stage-specific polysaccharide with specific properties, high activity of β-galactosidase, and the presence of substantial amount of free galactose. Similarity and differences in the gelatinous cell wall formation in the fibers of various plant species are discussed.

Intrusive growth of flax phloem fibers is of intercalary type.

Flax (Linum usitatissimum L.) phloem fibers elongate considerably during their development and intrude between existing cells. We questioned whether fiber elongation is caused by cell tip growth or intercalary growth. Cells with tip growth are characterized by having two specific zones of cytoplasm in the cell tip, one with vesicles and no large organelles at the very tip and one with various organelles amongst others longitudinally arranged cortical microtubules in the subapex. Such zones were not observed in elongating flax fibers. Instead, organelles moved into the very tip region, and cortical microtubules showed transversal and helical configurations as known for cells growing in intercalary way. In addition, pulse-chase experiments with Calcofluor White resulted in a spotted fluorescence in the cell wall all over the length of the fiber. Therefore, it is concluded that fiber elongation is not achieved by tip growth but by intercalary growth. The intrusively growing fiber is a coenocytic cell that has no plasmodesmata, making the fibers a symplastically isolated domain within the stem.

Intrusive growth of sclerenchyma fibers

Intrusive growth is a type of cell elongation when the rate of its longitudinal growth is higher than that of surrounding cells; therefore, these cells intrude between the neighboring cells penetrating the middle lamella. The review considers the classical example of intrusive growth, e.g., elongation of sclerenchyma fibers when the cells achieve the length of several centimeters. We sum the published results of investigations of plant fiber intrusive growth and present some features of intrusive growth characterized by the authors for flax (Linum usitatissimum L.) and hemp (Cannabis sativa L.) fibers. The following characteristics of intrusive growth are considered: its rate and duration, relationship with the growth rate of surrounding cells, the type of cell elongation, peculiarities of the fiber primary cell wall structure, fibers as multinucleate cells, and also the control of intrusive growth. Genes, which expression is sharply reduced at suppression of intrusive growth, are also considered. Arguments for separation of cell elongation and secondary cell wall formation in phloem fibers and also data indicating diffuse type of cell enlargement during intrusive growth are presented.

Arrangement of mixed-linkage glucan and glucuronoarabinoxylan in the cell walls of growing maize roots.

BACKGROUND AND AIMS Plant cell enlargement is unambiguously coupled to changes in cell wall architecture, and as such various studies have examined the modification of the proportions and structures of glucuronoarabinoxylan and mixed-linkage glucan in the course of cell elongation in grasses. However, there is still no clear understanding of the mutual arrangement of these matrix polymers with cellulose microfibrils and of the modification of this architecture during cell growth. This study aimed to determine the correspondence between the fine structure of grass cell walls and the course of the elongation process in roots of maize (Zea mays). METHODS Enzymatic hydrolysis followed by biochemical analysis of derivatives was coupled with immunohistochemical detection of cell wall epitopes at different stages of cell development in a series of maize root zones. KEY RESULTS Two xylan-directed antibodies (LM11 and ABX) have distinct patterns of primary cell wall labelling in cross-sections of growing maize roots. The LM11 epitopes were masked by mixed-linkage glucan and were revealed only after lichenase treatment. They could be removed from the section by xylanase treatment. Accessibility of ABX epitopes was not affected by the lichenase treatment. Xylanase treatment released only part of the cell wall glucuronoarabinoxylan and produced two types of products: high-substituted (released in polymeric form) and low-substituted (released as low-molecular-mass fragments). The amount of the latter was highly correlated with the amount of mixed-linkage glucan. CONCLUSIONS Three domains of glucuronoarabinoxylan were determined: one separating cellulose microfibrils, one interacting with them and a middle domain between the two, which links them. The middle domain is masked by the mixed-linkage glucan. A model is proposed in which the mixed-linkage glucan serves as a gel-like filler of the space between the separating domain of the glucuronoarabinoxylan and the cellulose microfibrils. Space for glucan is provided along the middle domain, the proportion of which increases during cell elongation.

Intrusive growth of primary and secondary phloem fibres in hemp stem determines fibre-bundle formation and structure

Plant fibres – cells with important mechanical functions and a widely used raw material – are usually identified in microscopic sections only after reaching a significant length or after developing a thickened cell wall. We characterized the early developmental stages of hemp stem phloem fibres, both primary and secondary, when they still had only a primary cell wall. We gave a major emphasis to the role of intrusive elongation, the specific type of plant cell growth, by which fibres commonly attain large cell length. Intrusive growth is the key determinant of final bundle structure, both for primary and secondary phloem fibres.

Tissue-specific processes during cell wall formation in flax fiber

Flax (Linum usitatissimum L.) phloem fiber elongation is separate from secondary cell wall formation. The indicator for the developmental transition is the manually determinable “snap point”. Sharp increase in the mechanical strength at certain level of flax stem. It helped to characterize fiber-specific and stage-specific processes: soluble galactan turnover (revealed in pulse-chase experiments), specialized Golgi vesicle accumulation, and cell wall postsynthetic modification.

Plant fiber intrusive growth characterized by NMR method

Intrusively growing plant cells insert themselves between surrounding cells, thus increasing the number of membranes on the tissue cross-section. This parameter can be assessed by spin echo NMR method with a magnetic field pulse gradient. Diffusion echo decay was measured for stem regions of long-fiber flax (Linum usitatissimum L.) differing in the stages of primary fiber development, which elongate thousand-fold during intrusive growth. Additionally, the number of fibers on stem cross-sections was counted under microscope. An increase in the slow component of the echo diffusion decay was correlated with an increase in the number of fibers on the stem cross-section in the zone of intrusive growth, while other stem-structure characteristics remained unchanged. Thus, NMR method can be used for characterization of intrusive fiber growth in situ.

Formation of “Nonculturable” dormant forms of the phytopathogenic enterobacterium Erwinia carotovora

Reversible transition of the phytopathogenic gram-negative bacterium Erwinia carotovora, subsp. atroseptica, strain SCRI1043, to a dormant state was demonstrated; it was associated with a complete loss of cell ability to form colonies on the standard medium, i.e., with acquiring “non-culturability”. Entering of Erwinia cells to a nonculturable state occurred after long-term incubation (100–150 days) of the stationary-phase cell suspensions in either a fresh complete medium or in the carbon-free mineral medium or treatment with a chemical analogue of microbial anabiosis autoinducers (4 × 10−4 M of C12-alkylhydroxybenzene, AHB). However, confocal laser microscopy of the cells stained with the Live/Dead BacLight kit revealed that the majority of E. carotovora cells (90%) from long-incubated suspensions retained membrane integrity. In these suspensions, round cells of smaller size prevailed, with the envelope, containing an electron-dense outer layer and an underlying layer of lower density; the cytoplasm was coarse-granulated. Detection of “nonculturable” E. carotovora cells by quantitative real-time PCR analysis (Q-PCR) with specific primers by using standard procedures of sample preparation was shown to be inefficient. A special procedure including cell washing from the incubation medium in the absence of growth stimulation was developed, which promoted recovery of the colony-forming ability of the cells (up to 10% of the initial CFU number) and improved cell detection by Q-PCR from the number of genomic copies. The results provided further insight into the ways of long-term survival of phytopathogenic bacteria under environmental changes and carbon starvation.

Dissociation of a population of Pectobacterium atrosepticum SCRI1043 in tobacco plants: formation of bacterial emboli and dormant cells

The population dynamics of Pectobacterium atrosepticum SCRI1043 (Pba) within tobacco plants was monitored from the time of inoculation until after long-term preservation of microorganisms in the remnants of dead plants. We found and characterised peculiar structures that totally occlude xylem vessels, which we have named bacterial emboli. Viable but non-culturable (VBN) Pba cells were identified in the remnants of dead plants, and the conditions for resuscitation of these VBN cells were established. Our investigation shows that dissociation of the integrated bacterial population during plant colonisation forms distinct subpopulations and cell morphotypes, which are likely to perform specific functions that ensure successful completion of the life cycle within the plant.