Andrew J. Bowling

Immunocytochemical characterization of tension wood: Gelatinous fibers contain more than just cellulose.

Gelatinous fibers (G-fibers) are the active component of tension wood. G-fibers are unlike traditional fiber cells in that they possess a thick, nonlignified gelatinous layer (G-layer) internal to the normal secondary cell wall layers. For the past several decades, the G-layer has generally been presumed to be composed nearly entirely of crystalline cellulose, although several reports have appeared that disagreed with this hypothesis. In this report, immunocytochemical techniques were used to investigate the polysaccharide composition of G-fibers in sweetgum (Liquidambar styraciflua; Hamamelidaceae) and hackberry (Celtis occidentalis; Ulmaceae) tension wood. Surprisingly, a number of antibodies that recognize arabinogalactan proteins and RG I-type pectin molecules bound to the G-layer. Because AGPs and pectic mucilages are found in other plant tissues where swelling reactions occur, we propose that these polymers may be the source of the contractile forces that act on the cellulose microfibrils to provide the tension force necessary to bend the tree trunk.

Gelatinous fibers are widespread in coiling tendrils and twining vines

Although the coiling of tendrils and the twining of vines has been investigated since Darwins time, a full understanding of the mechanism(s) of this coiling and twining ability has not yet been obtained. In a previous study (Planta 225: 485-498), gelatinous (G) fibers in tendrils of redvine occurred concomitantly with the ability to coil, strongly indicating their role in the coiling process. In this study, tendrils and twining vines of a number of species were examined using microscopic and immunocytochemical techniques to determine if a similar presence and distribution of these fibers exists in other plant species. Tendrils that coiled in many different directions had a cylinder of cortical G fibers, similar to redvine. However, tendrils that coiled only in a single direction had gelatinous fibers only along the inner surface of the coil. In tendrils with adhesive tips, the gelatinous fibers occurred in the central/core region of the tendril. Coiling occurred later in development in these tendrils, after the adhesive pad had attached. In twining stems, G fibers were not observed during the rapid circumnutation stage, but were found at later stages when the vines position was fixed, generally one or two nodes below the node still circumnutating. The number and extent of fiber development correlated roughly with the amount of torsion required for the vine to ascend a support. In contrast, species that use adventitious roots for climbing or were trailing/scrambling-type vines did not have G fibers. These data strongly support the concept that coiling and twining in vines is caused by the presence of G fibers.

Plasma Membrane-Associated SCAR Complex Subunits Promote Cortical F-Actin Accumulation and Normal Growth Characteristics in Arabidopsis Roots

The ARP2/3 complex, a highly conserved nucleator of F-actin polymerization, and its activator, the SCAR complex, have been shown to play important roles in leaf epidermal cell morphogenesis in Arabidopsis. However, the intracellular site(s) and function(s) of SCAR and ARP2/3 complex-dependent actin polymerization in plant cells remain unclear. We demonstrate that putative SCAR complex subunits BRK1 and SCAR1 are localized to the plasma membrane at sites of cell growth and wall deposition in expanding cells of leaves and roots. BRK1 localization is SCAR-dependent, providing further evidence of an association between these proteins in vivo. Consistent with plasma membrane localization of SCAR complex subunits, cortical F-actin accumulation in root tip cells is reduced in brk1 mutants. Moreover, mutations disrupting the SCAR or ARP2/3 complex reduce the growth rate of roots and their ability to penetrate semi-solid medium, suggesting reduced rigidity. Cell walls of mutant roots exhibit abnormal structure and composition at intercellular junctions where BRK1 and SCAR1 are enriched in the adjacent plasma membrane. Taken together, our results suggest that SCAR and ARP2/3 complex-dependent actin polymerization promotes processes at the plasma membrane that are important for normal growth and wall assembly.

The mechanism for explosive seed dispersal in Cardamine hirsuta (Brassicaceae)

PREMISE Although many highly successful weed species use a ballistic seed dispersal mechanism, little is known about the mechanics of this process. Bittercress (Cardamine hirsuta) siliques are morphologically similar to Arabidopsis siliques, but they can project their seeds up to 5 m, while Arabidopsis seeds are dispersed by gravity. Comparison of these species should enable us to determine which structures might be responsible for ballistic seed dispersal. METHODS Sections of Arabidopsis and bittercress siliques were immunolabeled with antibodies raised against a variety of polysaccharide epitopes. RESULTS In bittercress, the second endocarp layer (enB) of the valve had strongly asymmetrical cell wall thickenings, whereas the analogous cells in Arabidopsis were reinforced symmetrically and to a lesser extent. Additionally, an accumulation of mucilaginous pectins was found between the first and second endocarp (enA and enB) layers in the bittercress valve that was not present in Arabidopsis. However, in both species, highly de-esterified homogalacturonan was lost in the dehiscence zone (at the carpel/replum interface) as the siliques matured, thus allowing for separation of the valve at maturity. CONCLUSIONS Ballistic seed dispersal in bittercress may involve the contraction of the outer pericarp tissue against the highly asymmetrically thickened enB cells, which are hypothesized to bend in one direction preferentially. The stress generated by the differential drying of the inner and outer layers of the valve is released suddenly as the adhesion between the cells of the dehiscence zone is lost, leading to a rapid coiling of the valve and dispersal of the seeds.

Hemp sesbania (Sesbania exaltata) control in rice (Oryza sativa) with the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene formulated in an invert emulsion

Abstract In greenhouse and field experiments, an invert emulsion (MSG 8.25) was tested with dried, formulated spores of the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene, a highly virulent pathogen of the leguminous weed Aeschynomene virginica (northern jointvetch), but considered ‘immune’ against another more serious leguminous weed, Sesbania exaltata (hemp sesbania). A 1:1 (v/v) fungus/invert emulsion mixture resulted in 100% infection and mortality of inoculated hemp sesbania seedlings over a 21-day period under greenhouse conditions. In replicated field tests of the fungus/invert formulation conducted in Stuttgart, AR, and Stoneville, MS, hemp sesbania was controlled 85 and 90%, respectively. These results suggest that this invert emulsion expands the host range of C. gloeosporioides f. sp. aeschynomene, with a concomitant improvement of the bioherbicidal potential of this pathogen.

Leaf abscission in Impatiens (Balsaminaceae) is due to loss of highly de-esterified homogalacturonans in the middle lamellae

PREMISE OF STUDY Abscission zones (AZ) are sites where leaves and other organs are shed. Investigating the AZ by classical biochemical techniques is difficult due to its small size and because the surrounding tissue is not involved in abscission. The goals of this study were to determine whether AZ cell walls are chemically unique from the other cells of the petiole, perhaps making them more susceptible to enzymatic degradation during abscission and to identify which cell wall polysaccharides are degraded during abscission. METHODS A battery of antibodies that recognize a large number of cell wall polysaccharide and glycoprotein epitopes was used to probe sections of the Impatiens leaf AZ at several time points in the abscission process. KEY RESULTS Prior to abscission, the walls of the AZ cells were found to be similar in composition to the walls of the cells both proximal and distal to the AZ. Of all the epitopes monitored, only the highly de-esterified homogalacturonans (HG) of the middle lamellae were found to be reduced post-abscission and only at the plane of separation. More highly esterified homogalacturonans, as well as other pectin and xyloglucan epitopes were not affected. Furthermore, cellulose, as detected by an endoglucanase-gold probe and cellulose-binding module staining, was unaffected, even on the walls of the cells facing the separation site. CONCLUSIONS In the leaf abscission zone of Impatiens, wall alterations during abscission are strictly limited to the plane of separation and involve only the loss of highly de-esterified pectins from the middle lamellae.

Immunohistochemical investigation of the necrotrophic phase of the fungus Colletotrichum gloeosporioides in the biocontrol of hemp sesbania (Sesbania exaltata; Papilionaceae)

UNLABELLED PREMISE OF THE STUDY Fungal plant pathogens exert much of their effect on plant cells through alterations in the host cell walls. However, obtaining biochemical proof for this change is difficult because of the relatively small number of cells that are affected by the pathogen relative to the bulk of host tissue. In this study, we examined the differences in host wall composition between infected and uninfected areas of seedlings of the weed hemp sesbania (Sesbania exaltata) that were treated with the biocontrol agent Colletotrichum gloeosporioides. • METHODS To determine the changes in cell wall composition, we used semi-thin sections and a battery of antibody probes that recognize components of the cell wall and immunogold-silver cytochemistry to visualize the probes. • KEY RESULTS A loss of specific plant cell wall polysaccharides in the region surrounding the primary fungal infection and the creation of a defensive layer by the plant to limit the fungal invasion were the two most obvious changes noted in this study. At the invasion site, there was significant loss of rhamnogalacturon-1 (RGI) and esterified and de-esterified homogalacturonan (HG)-reactive epitopes from the cell walls. In contrast, boundary tissue between the vascular tissue and the fungal lesion reacted more strongly with antibodies that recognize arabinogalactan proteins (AGPs) and xyloglucans than in unaffected areas. • CONCLUSIONS These data strongly indicate a role of pectinases in the invasion of the biocontrol agent and the importance of extensins, AGPs, and xyloglucans as defense by the host.

The Vestigial Root of Dodder (Cuscuta pentagona) Seedlings

Seedlings of dodder are unique among dicotyledonous plants in that they emerge as a leafless, cotyledonless shoot with only a small swollen rootlike structure at the base of the tissue. Although growth of the shoot end of the dodder seedling is dramatic, no change in “root” length occurs, and the root tip is withered and senescent within 7 d of germination. Unlike most roots, the dodder root has neither recognizable root cap nor apical meristem. A strand of vascular tissue extends all the way to the root apex and is already differentiated into vascular elements on germination. Cortical cells swell dramatically and contain large vacuoles with a small rim of cytoplasm. Nuclei in these cortical cells are extensively lobed and are much larger than nuclei in shoot tips, indicating endopolyploidy. Microtubules are detected, although they are much less abundant than in shoot tissue of dodder or roots of other dicots, especially in roots older than 1 d postgermination. Similarly, α‐tubulin protein, as detected by immunoblots, appear as faint bands in root extracts; both are easily detectable in extracts of shoot tissue. Cell walls of 1–2‐d‐old roots are normal in morphology and contain well‐defined cellulose microfibrils and well‐developed middle lamellae. In contrast, later stages of development reveal cell‐wall‐loosening complexes and the degradation of wall structure and loss of polysaccharides, especially those of pectin side chains, which also are lost in other senescent tissues. By 5–7 d postgermination, all of the cortical cells have degenerated, leaving the vascular strand as the last remnant of intact tissue in these roots. From these data, we conclude that the swollen appearance of the dodder root is due to the low level of microtubules, so that neither mitotic divisions nor cell elongation can occur, and the loosening/senescence of the cell wall allows for expansion, resulting in a swollen root phenotype. It is speculated that the degeneration of the root end of the dodder may allow a flow of carbon from this organ to sustain the continued growth of the shoot. Although the tuberous end is clearly differentiated from the shoot tissue, it probably should be considered a highly modified basal portion of stem tissue used as a food reserve and basal support rather than a root.