Regenerative Xylem in Inflorescence Stems of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

By inserting entomological needles into the lower parts of young inflorescence stems of three-month-old Arabidopsis thaliana (L.) Heynh var. Colombia plants, we studied the process of regenerative xylem production. Regenerative xylem was formed only in one- to two-day-old inflorescence stems but not in older ones. The regenerative vessels originated from re-differentiation of cortical parenchyma. To characterize the process of regenerative xylem formation, we conducted a histological study from the time of wounding to day 30 after wounding. In the first day after wounding the tissues showed no structural responses except for the wounding itself. After six days, regenerative vessel members were already differentiating in a basipetal pattern, forming a vascular bypass around the wound. Regenerative vessel member formation reached a maximal level on the twelfth day after wounding. Sixteen days after wounding the pith parenchyma started to become loose as if indicating tissue senescence. Altogether, vascular regeneration following wounding in inflorescence stems of Arabidopsis thaliana is similar to that in other dicotyledon plants. These findings provide the basis for the use of Arabidopsis thaliana as a model system to study the genetics, physiology and cell biology of wound healing and regenerative vascular tissue formation.     

The Inflorescence Stem Fibers of <Emphasis Type="Italic">Arabidopsis thaliana Revoluta</Emphasis> (<Emphasis Type="Italic">ifl1</Emphasis>) Mutant

Arabidopsis thaliana is gradually gaining significance as a model for wood and fiber formation.revolute/ifl1 is an important mutant in this respect. To better characterize the fiber system of therevolute/ifl1 mutant, we grew plants of two alleles (rev-9 in Israel andrev-1 in the USA) and examined the fiber system of the inflorescence stems using both brightfield and polarized light. Microscopic examination of sections of plants belonging to the two different alleles clearly revealed that, contrary to previous views, in 18 (13 in Israel and 5 in Ohio) out of 30 stems (20 in Israel and 10 in Ohio) the mutant produced the primary wavy fiber system of the inflorescence stems. Our findings are further supported by the fact that fibers are seen in the figures published in other studies of the mutant even when it was stated that there were no fibers. The impression of a total lack of the wavy band of fibers is in many cases just a result of poorly lignified secondary walls. This specific gene that reduces lignification in fibers is of great significance for biotechnological developments for the paper industry and thus for the global economy and ecology. We propose thatrevoluta, the first name given to this mutant (Talbert and others 1995), is more appropriate thanifl1. Online publication: 7 April 2005     

<Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> as a Model System for Graft Union Development in Homografts and Heterografts

Homografting of Arabidopsis thaliana scions on stocks of A. thaliana and heterografting on other species were used to study the compatibility and the ontogeny of graft union formation. Highly compatible homografting with scions of young leafy inflorescence stems was obtained on stocks of inflorescence stems growing from large 3-month-old A. thaliana plants. Histologic analysis revealed four developmental stages of graft union formation in Arabidopsis homografting: (1) development of a necrotic layer, (2) callus proliferation in the grafted scion, (3) differentiation of new vascular tissues within the scion, and (4) a full vascular graft union formation between the scion and the stock. Vascular connections were formed within the callus bridge between rootstocks and scions 15 days after grafting. Heterografts of Arabidopsis on two members of Brassicaceae, cabbage (Brassica) and radish (Raphanus), showed partial incompatible interaction with a lower level of vascular differentiation. Arabidopsis grafting on tomato (Solanaceae) rootstock showed complete incompatibility and limited noncontinuous differentiation of new vascular tissues that did not cross the scion/stock boundary. Although lacking scion/stock vascular connections, Arabidopsis scions grafted onto tomato rootstock flowered and produced seeds. This may indicate some nonvascular functional connections between the two plants, probably of parenchyma cells, further emphasizing the usefulness of Arabidopsis as a model plant for studying various levels of the complicated scion/stock relationships expressed in grafting biology. Experiments with dye transport in the xylem showed that although in general there was an agreement between the histologic study and dye transport, in Arabidopsis homografts water transport frequency was lower than functional and histologic compatability. We conclude that homografting and heterografting of Arabidopsis inflorescence stems is a convenient and reproducible method for studying the fundamental cellular genetic and molecular aspects of grafting biology.     

Ethylene evolution changes in the stems of<Emphasis Type="Italic"> Metasequoia glyptostroboides</Emphasis> and<Emphasis Type="Italic"> Aesculus turbinata</Emphasis> seedlings in relation to gravity-induced reaction wood formation

Two-year-old Metasequoia glyptostroboides and 3-month-old Aesculus turbinata seedlings were tilted at a 45° angle to induce compression wood formation on the lower side of the former species and tension wood on the upper side of the latter. Two weeks later, the seedlings were tilted in an opposite direction at 45° so that the upper and lower sides changed to each other. This reverse tilting was kept for 7 weeks for M. glyptostroboides and 6 weeks for A. turbinata. The seedlings were sampled and analyzed at intervals throughout each experimental period so that an ethylene evolution kinetic was monitored. Ethylene evolution from the cambial region of the upper and lower sides of tilted stems was measured separately by gas chromatography with a flame ionization detector. Xylem production expressed as wood area during each experimental period was microscopically determined. In both tilting and reverse tilting periods, the rates of ethylene evolution from the lower side of M. glyptostroboides and the upper side of A. turbinata, where xylem production was accelerated and compression or tension wood formation was induced, had increased to high levels, whereas those from the opposite sides had either remained low (in tilting period) or rapidly recovered to low levels (in reverse tilting period). The cambial activity quantified by wood formation, including reaction wood, in both species showed the same tendency as ethylene evolution. The stem side with vigorous ethylene evolution, xylem development and reaction wood formation reversed with the reversal of tilting orientation. The roles of accelerated ethylene evolution in reaction wood formation in the tilted seedlings of gymnosperm and angiosperm trees are compared and discussed.This work was presented at the 5th Pacific Region Wood Anatomy Conference, Yogyakarta, Indonesia, 9–14 September 2002     

Stems of the <Emphasis Type="Italic">Arabidopsis pin1-1</Emphasis> mutant are not deficient in free indole-3-acetic acid

The pin1-1 mutant of Arabidopsis thaliana has been pivotal for studies on auxin transport and on the role of auxin in plant development. It was reported previously that when whole shoots were analysed, levels of the major auxin, indole-3-acetic acid (IAA) were dramatically reduced in the mutant, compared with the WT (Okada et al. 1991). The cloning of PIN1, however, provided evidence that this gene encodes a facilitator of auxin efflux, raising the question of how the pin1-1 mutation might reduce overall IAA levels as well as IAA transport. We therefore re-examined IAA levels in individual parts of pin1-1 and WT plants, focusing on inflorescence stems. Our data show that there is in fact no systemic IAA deficiency in the mutant. The previously reported difference between mutant and WT may have been due to the inclusion of reproductive structures in the WT harvest: we show here that the inflorescence itself contains high levels of IAA. We reconcile the normal IAA levels of pin1-1 inflorescence stems with their (previously-reported) reduced ability to transport IAA by presenting evidence that the auxin in mutant stems is not imported from their apical portion. Our data also indicate that levels of another auxin, indole-3-butyric acid (IBA), are very low in stems of the genotypes used in this study.     

Gibberellin mediates the development of gelatinous fibres in the tension wood of inclined Acacia mangium seedlings

Background and Aims

Gibberellin stimulates negative gravitropism and the formation of tension wood in tilted Acacia mangium seedlings, while inhibitors of gibberellin synthesis strongly inhibit the return to vertical growth and suppress the formation of tension wood. To characterize the role of gibberellin in tension wood formation and gravitropism, this study investigated the role of gibberellin in the development of gelatinous fibres and in the changes in anatomical characteristics of woody elements in Acacia mangium seedlings exposed to a gravitational stimulus.

Methods

Gibberellin, paclobutrazol and uniconazole-P were applied to the soil in which seedlings were growing, using distilled water as the control. Three days after the start of treatment, seedlings were inclined at 45 ° to the vertical and samples were harvested 2 months later. The effects of the treatments on wood fibres, vessel elements and ray parenchyma cells were analysed in tension wood in the upper part of inclined stems and in the opposite wood on the lower side of inclined stems.

Key Results

Application of paclobutrazol or uniconazole-P inhibited the increase in the thickness of gelatinous layers and prevented the elongation of gelatinous fibres in the tension wood of inclined stems. By contrast, gibberellin stimulated the elongation of these fibres. Application of gibberellin and inhibitors of gibberellin biosynthesis had only minor effects on the anatomical characteristics of vessel and ray parenchyma cells.

Conclusions

The results suggest that gibberellin is important for the development of gelatinous fibres in the tension wood of A. mangium seedlings and therefore in gravitropism.     

Effects of hypergravity conditions on elongation growth and lignin formation in the inflorescence stem of Arabidopsis thaliana

The effects of hypergravity on elongation growth and lignin deposition in secondary cell walls of the Arabidopsis thaliana (L.) Heynh. inflorescence stem were examined in plants grown for 3 days after exposure to hypergravity in the direction from shoot to root at 300 g for 24 h. The content of acetylbromide-extractable lignins in a secondary cell wall fraction prepared by enzyme digestion of inflorescence stem segments removing primary cell wall components was significantly increased by the hypergravity stimulus. Xylem vessels, particularly in a region closer to the base of the inflorescence stem, increased in number. Gadolinium chloride at 0.1 mM, a blocker of mechanoreceptors, partially suppressed the effect of hypergravity on lignin deposition in the secondary cell wall fraction. These results suggest that mechanoreceptors are responsible for hypergravity-induced lignin deposition in secondary cell walls in A. thaliana inflorescence stems.     

An Overview of the Biology of Reaction Wood Formation

Reaction wood possesses altered properties and performs the function of regulating a tree＇s form, but it is a serious defect in wood utility. Trees usually develop reaction wood in response to a gravistimulus. Reaction wood in gymnosperms is referred to as compression wood and develops on the lower side of leaning stems or branches. In arboreal, dicotyledonous angiosperms, however, it is called tension wood and is formed on the upper side of the leaning. Exploring the biology of reaction wood formation is of great value for the understanding of the wood differentiation mechanisms, cambial activity, gravitropism, and the systematics and evolution of plants. After giving an outline of the variety of wood and properties of reaction wood, this review lays emphasis on various stimuli for reaction wood induction and the extensive studies carried out so far on the roles of plant hormones in reaction wood formation. Inconsistent results have been reported for the effects of plant hormones. Both auxin and ethylene regulate the formation of compression wood in gymnosperms. However, the role of ethylene may be indirect as exogenous ethylene cannot induce compression wood formation. Tension wood formation is mainly regulated by auxin and gibberellin. Interactions among hormones and other substances may play important parts in the regulation of reaction wood formation.     

An unusual form of reaction wood in Koromiko [<Emphasis Type="Italic">Hebe salicifolia</Emphasis> G. Forst. (Pennell)], a southern hemisphere angiosperm

Koromiko [Hebe salicifolia G. Forst. (Pennell)] is a woody angiosperm native to New Zealand and Chile. Hebe spp. belong to the otherwise herbaceous family Plantaginaceae in the order Lamiales. Reaction wood exerting expansional forces was found on the lower side of leaning H. salicifolia stems. Such reaction wood is atypical for angiosperms, which commonly form contracting reaction wood on the upper side of leaning stems. Reaction wood typical for angiosperms is formed by species in other families in the order Lamiales. This suggests that the form of reaction wood is specific to the family level. Functionally the reaction wood of H. salicifolia is similar to that found in gymnosperms, which both act by pushing. However, their chemical, anatomical and physical characteristics are different. Typical features of reaction wood present in gymnosperms such as high density, thick-walled rounded cells and the presence of (1 → 4)-β-galactan in the secondary cell wall layer are absent in H. salicifolia reaction wood. Reaction wood of H. salicifolia varies from normal wood in having a higher microfibril angle, which is likely to determine the direction of generated maturation stresses.     

Induction of tension wood by 2,4-dinitrophenol and auxins

Summary The effect of DNP and auxins on the development of the secondary xylem in erect stems ofAcer rubrum was studied. DNP affected the development of the secondary xylem only locally in the treated internode. Tension wood is formed in the stem below the DNP treatment site whereas above the application site the development of tracheary elements is altered. InAcer rubrum seedlings that were treated with auxin, especially at low concentrations, a thick ring of tension wood is developed in the erect stem below the treatment site. Previous suggestions that the formation of tension wood in arborescent angiosperms is a developmental response to auxin deficiency are discussed in terms of the induction of tension wood inAcer rubrum by DNP and auxins.The following abbreviations will be used TIBA (2,3,5-tri-iodobenzoic acid) - IAA (indole-3-acetic acid) - GA (gibberellic acid) - NAA (naphthaleneacetic acid) - 2,4-D (2,4-dichlorophenoxyacetic acid) - DNP (2,4-dinitrophenol) This material was included in a doctoral thesis submitted by P. R.Morey to the graduate school of Yale University, New Haven.     

Ultra-structural organisation of cell wall polymers in normal and tension wood of aspen revealed by polarisation FTIR microspectroscopy

Polarisation Fourier transform infra-red (FTIR) microspectroscopy was used to characterize the organisation and orientation of wood polymers in normal wood and tension wood from hybrid aspen (Populus tremula × Populus tremuloides). It is shown that both xylan and lignin in normal wood are highly oriented in the fibre wall. Their orientation is parallel with the cellulose microfibrils and hence in the direction of the fibre axis. In tension wood a similar orientation of lignin was found. However, in tension wood absorption peaks normally assigned to xylan exhibited a 90° change in the orientation dependence of the vibrations as compared with normal wood. The molecular origin of these vibrations are not known, but they are abundant enough to mask the orientation dependence of the xylan signal from the S₂ layer in tension wood and could possibly come from other pentose sugars present in, or associated with, the gelatinous layer of tension wood fibres.     

Ultrastructural Investigation of Tension Wood Fibre in Fraxinus mandshurica Rupr. var. japonica Maxim.

The ultrastructure of the fibre wall in Fraxinus mandshuricaRupr. var. japonica Maxim. was investigated by electron microscopy.The trees had been inclined artificially at an angle of 30°to the vertical at the beginning of the initiation of cambialgrowth in early spring. The secondary walls of tension woodfibres were of the outer (S₁) layer and gelatinous (G) layertype. The microfibrils in the gelatinous (G) layer were orientedas a steep Z-helix relative to the fibre axis with a deviationthat ranged from 0° to 25° but was mainly between 5°and 10°. The cross-sectional surface of tension wood fibresrevealed the relatively strong attachment of the G-layer tothe S₁ layer. The G-layer stained weakly with potassium permanganate.The S₁ layer of tension wood fibres stained less strongly thanthat of the normal and opposite wood fibres. These results indicatethat the tension wood in F. mandshurica var. japonica is nottypical and is somewhat anomalous. The secondary walls of normaland opposite wood fibres were composed of two layers, S₁ andS₂, and lacked an S₃ layer. Microfibrils in the S₃ layer ofjuvenile stems were extremely variable in orientation and weresparsely distributed without forming a layer. By contrast, avery thin S₃ layer was present in the wood fibres of maturestems. The variations in the formation of the S₃ layer in thefibre walls were probably due to the differences in the cambialage of the stems of F. mandshurica Rupr. var. japonica.Copyright1995, 1999 Academic Press Fraxinus mandshurica Rupr. var. japonica Maxim., Japanese ash, tension wood, fibre wall, G-layer, microfibrillar orientation, normal and opposite wood, juvenile stem, field-emission scanning electron microscopy, low accelerating voltage     

Bioethanol production from tension and opposite wood of <Emphasis Type="Italic">Eucalyptus globulus</Emphasis> using organosolv pretreatment and simultaneous saccharification and fermentation

During tree growth, hardwoods can initiate the formation of tension wood, which is a strongly stressed wood on the upper side of the stem and branches. In Eucalyptus globulus, tension wood presents wider and thicker cell walls with low lignin, similar glucan and high xylan content, as compared to opposite wood. In this work, tension and opposite wood of E. globulus trees were separated and evaluated for the production of bioethanol using ethanol/water delignification as pretreatment followed by simultaneous saccharification and fermentation (SSF). Low residual lignin and high glucan retention was obtained in organosolv pulps of tension wood as compared to pulps from opposite wood at the same H-factor of reaction. The faster delignification was associated with the low lignin content in tension wood, which was 15% lower than in opposite wood. Organosolv pulps obtained at low and high H-factor (3,900 and 12,500, respectively) were saccharified by cellulases resulting in glucan-to-glucose yields up to 69 and 77%, respectively. SSF of the pulps resulted in bioethanol yields up to 35 g/l that corresponded to 85–95% of the maximum theoretical yield on wood basis, considering 51% the yield of glucose to ethanol conversion in fermentation, which could be considered a very satisfactory result compared to previous studies on the conversion of organosolv pulps from hardwoods to bioethanol. Both tension and opposite wood of E. globulus were suitable raw materials for organosolv pretreatment and bioethanol production with high conversion yields.     

Gibberellin is required for the formation of tension wood and stem gravitropism in Acacia mangium seedlings

Background and Aims Angiosperm trees generally form tension wood on the upper sides of leaning stems. The formation of tension wood is an important response to gravitational stimulus. Gibberellin appears to be involved in the differentiation of secondary xylem, but it remains unclear whether gibberellin plays a key role in the formation of tension wood and plant gravitropism. Therefore, a study was designed to investigate the effects of gibberellin and of inhibitors of the synthesis of gibberellin, namely paclobutrazole and uniconazole-P, on the formation of tension wood and negative stem gravitropism in Acacia mangium seedlings. Methods Gibberellic acid (GA(3)), paclobutrazole and uniconazole-P were applied to seedlings via the soil in which they were growing. Distilled water was applied similarly as a control. Three days after such treatment, seedlings were tilted at an angle of 45° from the vertical, and samples of stems were collected for analysis 2 weeks, 2 months and 6 months after tilting. The effects of treatments on the stem recovery degree (Ro) were analysed as an index of the negative gravitropism of seedlings, together the width of the region of tension wood in the upper part of inclined stems. Key Results It was found that GA(3) stimulated the negative gravitropism of tilted seedling stems of A. mangium, while paclobutrazole and uniconazole-P inhibited recovery to vertical growth. Moreover, GA(3) stimulated the formation of tension wood in tilted A. mangium seedlings, while paclobutrazole and uniconazole-P strongly suppressed the formation of tension wood, as assessed 2 weeks after tilting. Conclusions The results suggest that gibberellin plays an important role at the initial stages of formation of tension wood and in stem gravitropism in A. mangium seedlings in response to a gravitational stimulus.     

Multilayered structure of tension wood cell walls in Salicaceae sensu lato and its taxonomic significance

Salicaceae have been enlarged to include a majority of the species formerly placed in the polyphyletic tropical Flacourtiaceae. Several studies have reported a peculiar and infrequently formed multilayered structure of tension wood in four of the tropical genera. Tension wood is a tissue produced by trees to restore their vertical orientation and most studies have focused on trees developing tension wood by means of cellulose‐rich, gelatinous fibres, as in Populus and Salix (Salicaceae s.s.). This study aims to determine if the multilayered structure of tension wood is an anatomical characteristic common in other Salicaceae and, if so, how its distribution correlates to phylogenetic relationships. Therefore, we studied the tension wood of 14 genera of Salicaceae and two genera of Achariaceae, one genus of Goupiaceae and one genus of Lacistemataceae, families closely related to Salicaceae or formerly placed in Flacourtiaceae. Opposite wood and tension wood were compared with light microscopy and three‐dimensional laser scanning confocal microscopy. The results indicate that a multilayered structure of tension wood is common in the family except in Salix, Populus and one of their closest relatives, Idesia polycarpa. We suggest that tension wood may be a useful anatomical character in understanding phylogenetic relationships in Salicaceae. Further investigation is still needed on the tension wood of several other putatively close relatives of Salix and Populus, in particular Bennettiodendron, Macrohasseltia and Itoa.     

Deposition and organisation of cell wall polymers during maturation of poplar tension wood by FTIR microspectroscopy

To advance our understanding of the formation of tension wood, we investigated the macromolecular arrangement in cell walls by Fourier transform infrared microspectroscopy (FTIR) during maturation of tension wood in poplar (Populus tremula x P. alba, clone INRA 717-1B4). The relation between changes in composition and the deposition of the G-layer in tension wood was analysed. Polarised FTIR measurements indicated that in tension wood, already before G-layer formation, a more ordered structure of carbohydrates at an angle more parallel to the fibre axis exists. This was clearly different from the behaviour of opposite wood. With the formation of the S₂ layer in opposite wood and the G-layer in tension wood, the orientation signals from the amorphous carbohydrates like hemicelluloses and pectins were different between opposite wood and tension wood. For tension wood, the orientation for these bands remains the same all along the cell wall maturation process, probably reflecting a continued deposition of xyloglucan or xylan, with an orientation different to that in the S₂ wall throughout the whole process. In tension wood, the lignin was more highly oriented in the S₂ layer than in opposite wood.