Distribution of tension wood like gelatinous fibres in the roots of Acacia nilotica (Lam.) Willd.

KEY MESSAGE: The present study unravels the anatomical characteristics and distribution patterns of cell wall polymers in the G-fibres found in the roots of A. nilotica using different microscopy techniques (light, electron and immunofluorescence microscopy). The present study was aimed to investigate the anatomy of reaction xylem in the positively gravitropic roots of Acacia nilotica growing in compact and waterlogged soils. The roots collected from the two different sites showed occurrence of gelatinous fibres throughout xylem radii from a distance of 4 cm from the soil surface. The thickness of gelatinous layer (G-layer) increased in the root collected from the deeper soil. Further, the ultrastructural studies revealed a complete replacement of S2 and S3 layers in G-fibres nearer to root tip region as compared to the root portion close to upper part of the soil surface. In addition, these fibres demonstrated intense lignification in compound middle lamellae region of G-fibre walls. Moreover, the vessel density and their width increased considerably near the root tip region. The immunofluorescence analysis suggested that the ß-1,4-galactans were prevalent in G-layer, whereas the xylan was restricted to only regions of lignified secondary wall. The similarities in distribution pattern and anatomical features of G-fibres in waterlogged and non-waterlogged roots suggest the occurrence of G-fibres as inherent characteristics in the roots of Acacia nilotica.

Ultrastructural changes during nectar secretion from extrafloral nectaries of Pithecellobium dulce Benth.

The structural changes in the secretory cells are important to understand the ontogeny and nectar secretion process from the nectaries. In this study, we investigated the ultrastructural changes during different developmental/secretion stages of extrafloral nectaries (EFNs) of Pithecellobium dulce. The dense cytoplasm with active biosynthesis mechanisms such as ribosomes, mitochondria, large nucleus, and plastids with accumulated starch grains characterized the pre-secretion stage of young nectariferous cells. During the secretory phase, the cytoplasm showed distinct changes associated with endomembrane transport such as the predominant occurrence of Golgi, secretory vesicles, and ER resulting in the subsequent appearance of secretions in the intercellular and subcuticular spaces. Cell wall loosening following the dissolution of middle lamellae leading to the formation of subcuticular spaces was evident during advanced stages of nectar secretion. The characteristic cytoplasmic and apoplastic changes associated with cell death were noticed during the post-secretory stages. The structural evidence from the present study suggests the occurrence of two modes of secretion (merocrine and holocrine) during the early and late stages of secretion in the EFNs of P. dulce.

Chemical Composition in Juvenile and Mature Wood of Branch and Main Trunk of Leucaena leucocephala (Lam.) de Wit.

Secondary growth is the most dynamic developmental aspect during the terrestrialization of plants. The development of secondary xylem tissue composed of thick-walled cells with characteristic changes in its structure and chemistry facilitates the growth and development of woody plants. In the present study, the chemical composition of the secondary xylem of juvenile and mature wood from the branch and main trunk of Leucaena leucocephala, has been investigated and the differences established. The biochemical analysis of different cell wall components in the mature wood of the main trunk revealed high holocellulose and α-cellulose and less lignin content in the juvenile wood while its syringyl/guaiacyl (S/G) ratio was less than for the mature wood. As compared to the branch xylem, concentration of cell wall polysaccharides and lignin content was higher in both juvenile and mature wood collected from the main trunk. Thioacidolysis and GC-MS analysis of wood lignin from juvenile and mature wood showed that an increased concentration in lignin content in mature wood is associated with a corresponding increase in S/G ratio. The structural information of the acetylated lignin was investigated by 1H NMR spectroscopy. Our results indicate that the mature wood from the main trunk is superior in pulp yielding and lignin degradability as compared to the juvenile wood of the branch and trunk.

Down-regulation of Leucaena leucocephala cinnamoyl CoA reductase (LlCCR) gene induces significant changes in phenotype, soluble phenolic pools and lignin in transgenic tobacco.

cDNA and genomic clones of cinnamoyl CoA reductase measuring 1011 and 2992 bp were isolated from a leguminous pulpwood tree Leucaena leucocephala, named as LlCCR. The cDNA exhibited 80-85% homology both at the nucleotide and amino acid levels with other known sequences. The genomic sequence contained five exons and four introns. Sense and antisense constructs of LlCCR were introduced in tobacco plants to up and down-regulate this key enzyme of lignification. The primary transformants showed a good correlation between CCR transcript levels and its activity. Most of the CCR down-regulated lines displayed stunted growth and development, wrinkled leaves and delayed senescence. These lines accumulated unusual phenolics like ferulic and sinapic acids in cell wall. Histochemical staining suggested reduction in aldehyde units and increased syringyl over guaiacyl (S/G) ratio of lignin. Anatomical studies showed thin walled, elongated xylem fibres, collapsed vessels with drastic reduction of secondary xylem. The transmission electron microscopic studies revealed modification of ultrastructure and topochemical distribution of wall polysaccharides and lignin in the xylem fibres. CCR down-regulated lines showed increased thickness of secondary wall layers and poor lignification of S2 and S3 wall layers. The severely down-regulated line AS17 exhibited 24.7% reduction of Klason lignin with an increase of 15% holocellulose content. Contrarily, the CCR up-regulated lines exhibited robust growth, development and significant increase in lignin content. The altered lignin profiles observed in transgenic tobacco lines support a role for CCR down-regulation in improving wood properties of L. leucocephala exclusively used in the pulp and paper industry of India.

Immunolocalization of ß-(1-4)-D-galactan, xyloglucans and xylans in the reaction xylem fibres of Leucaena leucocephala (Lam.) de Wit.

Cell wall architecture of tension wood fibres represents a suitable biological system to study the mechanism of growth and maintenance of posture of trees growing under various physical and physiological growth constraints. In the present study, we investigated the spatial distributions of ß-(1-4)-D-galactan, xyloglucan and xylans (both less and highly substituted) in the opposite and tension wood fibres of bent Leucaena leucocephala by immunolabelling with monoclonal antibodies LM5, CCRCM1, LM10 and LM11 specific to these carbohydrate epitopes. The presence of non-lignified, tertiary wall layer is the typical tension wood characteristic associated with the reaction xylem fibres in Leucaena. LM5 labelling of opposite fibres showed weak labelling in the cell walls indicating less concentration of ß-(1-4)-D-galactans while tension wood showed strong labelling in the tertiary wall layer suggesting the gelatinous layer (G-layer) has a strong cross linking with ß-(1-4)-D-galactans. Xyloglucan distribution was more in the compound middle lamellae and the primary wall-S1 layer boundary of tension wood fibres as compared to that of opposite wood. A weak labelling was also evident near the boundary between the G-layer and the secondary wall of tension wood fibres. The secondary wall of opposite and tension wood fibres showed a strong distribution of both ls ACG Xs (LM10) and hs ACG Xs (LM11) while a weak labelling was noticed in the compound middle lamella. Tension wood fibres also showed strong xylan labelling mainly confined to the lignified secondary walls while the G-layer showed weak xylan labelling. In conclusion, our results suggest that ß-(1-4)-D-galactans and xyloglucans could be implicated in the tensile stress generation within the G-layer of tension wood fibres of Leucaena leucocephala.