Litchi is a highly perishable fruit. Ripe litchi fruit loses quality quickly as they hang on tree, giving a very short hanging life and thus harvest period. This study attempted to explore the roles of cytokinin in regulating fruit ripening and senescence of litchi and examine the possibility of applying cytokinin in “on-tree storage” of the fruit. Exogenous cytokinin, forchlorfenuron (CPPU), was applied at 20 mg L⁻¹ 7 weeks after full bloom on litchi (Litchi chinensis cv. Feizixiao) fruit clusters. Color parameters, chlorophylls, anthocyanins, fruit and fruit part weights, total soluble solutes (TSSs), soluble sugars, organic acids, non-anthocyanin flavonoids, ethanol, and also CPPU residue in fruit were traced. CPPU residue was higher but decreased faster in the pericarp than in the aril, where it maintained < 10 μg kg⁻¹. CPPU had no significant effect on fruit weight but tended to increase pericarp weight. The treatment suppressed chlorophyll loss and anthocyanin accumulation in the pericarp, increased non-anthocyanin flavonoids in the aril, but had no significant effects on non-anthocyanin flavonoids in the pericarp and total sugar and organic acids in the aril. As the commercially ripe fruit hanged on tree, TSSs, total sugar, and sucrose decreased with ethanol and acetic acid accumulation in the aril. CPPU significantly suppressed the loss of sucrose and total sugar and the accumulation of ethanol and acetic acid in the aril and inhibited malondialdehyde accumulation in the pericarp of the overripe fruit. Soluble invertase, alcohol dehydrogenase, and pyruvate decarboxylase (PDC) activity and gene expression in the aril were downregulated by CPPU. The results suggest that cytokinin partially suppresses the ripening process in litchi and is effective to slow quality loss in the overripe fruit on tree.

The plasticity of root development represents a key trait that enables plants to adapt to diverse environmental cues. The pattern of cell wall deposition, alongside other parameters, affects the extent, and direction of root growth. In this study, we report that FASCICLIN-LIKE ARABINOGALACTAN PROTEIN 18 (FLA18) plays a role during root elongation in Arabidopsis thaliana. Using root-specific co-expression analysis, we identified FLA18 to be co-expressed with a sub-set of genes required for root elongation. FLA18 encodes for a putative extra-cellular arabinogalactan protein from the FLA-gene family. Two independent T-DNA insertion lines, named fla18-1 and fla18-2, display short and swollen lateral roots (LRs) when grown on sensitizing condition of high-sucrose containing medium. Unlike fla4/salt overly sensitive 5 (sos5), previously shown to display short and swollen primary root (PR) and LRs under these conditions, the PR of the fla18 mutants is slightly longer compared to the wild-type. Overexpression of the FLA18 CDS complemented the fla18 root phenotype. Genetic interaction between either of the fla18 alleles and sos5 reveals a more severe perturbation of anisotropic growth in both PR and LRs, as compared to the single mutants and the wild-type under restrictive conditions of high sucrose or high-salt containing medium. Additionally, under salt-stress conditions, fla18sos5 had a small, chlorotic shoot phenotype, that was not observed in any of the single mutants or the wild type. As previously shown for sos5, the fla18-1 and fla18-1sos5 root-elongation phenotype is suppressed by abscisic acid (ABA) and display hypersensitivity to the ABA synthesis inhibitor, Fluridon. Last, similar to other cell wall mutants, fla18 root elongation is hypersensitive to the cellulose synthase inhibitor, Isoxaben. Altogether, the presented data assign a new role for FLA18 in the regulation of root elongation. Future studies of the unique vs. redundant roles of FLA proteins during root elongation is anticipated to shed a new light on the regulation of root architecture during plant adaptation to different growth conditions.

The variability in leaf form in nature is immense. Leaf patterning occurs by differential growth, taking place during a limited window of morphogenetic activity at the leaf marginal meristem. While many regulators have been implicated in the designation of the morphogenetic window and in leaf patterning, how these effectors interact to generate a particular form is still not well understood. We investigated the interaction among different effectors of tomato (Solanum lycopersicum) compound-leaf development, using genetic and molecular analyses. Mutations in the tomato auxin response factor SlARF5/SlMP, which normally promotes leaflet formation, suppressed the increased leaf complexity of mutants with extended morphogenetic window. Impaired activity of the NAC/CUC transcription factor GOBLET (GOB), which specifies leaflet boundaries, also reduced leaf complexity in these backgrounds. Analysis of genetic interactions showed that the patterning factors SlMP, GOB and the MYB transcription factor LYRATE (LYR) coordinately regulate leaf patterning by modulating in parallel different aspects of leaflet formation and shaping. This work places an array of developmental regulators in a morphogenetic context. It reveals how organ-level differentiation rate and local growth are coordinated to sculpture an organ. These concepts are applicable to the coordination of pattering and differentiation in other species and developmental processes.

Summary The variability in leaf form in nature is immense. Leaf patterning occurs by differential growth, taking place during a limited window of morphogenetic activity at the leaf marginal meristem. While many regulators have been implicated in the designation of the morphogenetic window and in leaf patterning, how these effectors interact to generate a particular form is still not well understood. We investigated the interaction among different effectors of tomato (Solanum lycopersicum) compound-leaf development, using genetic and molecular analyses. Mutations in the tomato auxin response factor SlARF5/SlMP, which normally promotes leaflet formation, suppressed the increased leaf complexity of mutants with extended morphogenetic window. Impaired activity of the NAC/CUC transcription factor GOBLET (GOB), which specifies leaflet boundaries, also reduced leaf complexity in these backgrounds. Analysis of genetic interactions showed that the patterning factors SlMP, GOB and the MYB transcription factor LYRATE (LYR) coordinately regulate leaf patterning by modulating in parallel different aspects of leaflet formation and shaping, This work places an array of developmental regulators in a morphogenetic context. It reveals how organ-level differentiation rate and local growth are coordinated to sculpture an organ. These concepts are applicable to the coordination of pattering and differentiation in other species and developmental processes.

Litchi (Litchi chinensis Sonn.) is a subtropical fruit known for its attractive red pericarp color, semi-translucent white aril and unique flavor and aroma. Rapid post-harvest pericarp browning strictly limits litchi fruit marketing. In the current research, we hypothesized that modification of litchi fruit pericarp anatomy by hormone application may reduce fruit susceptibility to post-harvest pericarp browning. In this context, we hypothesized that cytokinin treatment, known to induce cell division, may yield fruit with thicker pericarp and reduced susceptibility for fruit surface micro-crack formation, water loss and post-harvest pericarp browning. Exogenous cytokinin treatment was applied at different stages along the course of litchi fruit development and the effect on fruit pericarp anatomy, fruit maturation and postharvest pericarp browning was investigated. Interestingly, cytokinin treatment, applied 4 weeks after full female bloom (WFB), during the phase of pericarp cell division, led to mature fruit with thicker pericarp, reduced rate of post-harvest water loss and reduced susceptibility to post-harvest pericarp browning, as compared to non-treated control fruit. Histological sections ascribe the difference in pericarp anatomy to increased cell proliferation in the parenchymatic tissue and the highly-lignified brachysclereid cell layer. In contrast, exogenous cytokinin treatment applied 7 WFB, following the phase of pericarp cell division, significantly increased epidermal-cell proliferation but had no significant effect on overall fruit pericarp thickness and only minor affect on post-harvest water loss or pericarp browning. Interestingly, the late cytokinin treatment also significantly postponed fruit maturation-associated anthocyanin accumulation and chlorophyll degradation, as previously reported, but had no effect on other parameters of fruit maturation, like total soluble sugars and total titratable acids typically modified during aril maturation. In conclusion, exogenous cytokinin treatment at different stages in fruit development differentially modifies litchi fruit pericarp anatomy by induction of cell-type specific cell proliferation. Early cytokinin treatment during the phase of pericarp cell division may prolong litchi fruit storage by reducing fruit susceptibility to post-harvest water loss and pericarp browning.

The production of hydrophilic mucilage along the course of seed coat epidermal cell differentiation is a common adaptation in angiosperms. Previous studies have identified COBRA-LIKE 2 (COBL2), a member of the COBRA-LIKE gene family, as a novel component required for crystalline cellulose deposition in seed coat epidermal cells. In recent years, Arabidopsis seed coat epidermal cells (SCEs), also called mucilage secretory cells, have emerged as a powerful model system for the study of plant cell wall components biosynthesis, secretion, assembly and de muro modification. Despite accumulating data, the molecular mechanism of COBL function remains largely unknown. In the current research, we utilized genetic interactions to study the role of COBL2 as part of the protein network required for seed mucilage production. Using correlative phenotyping of structural and biochemical characteristics, unique features of the cobl2 extruded mucilage are revealed, including: 'unraveled' ray morphology, loss of primary cell wall 'pyramidal' organization, reduced Ruthenium red staining intensity of the adherent mucilage layer, and increased levels of the monosaccharides arabinose and galactose. Examination of the cobl2cesa5 double mutant provides insight into the interface between COBL function and cellulose deposition. Additionally, genetic interactions between cobl2 and fei1fei2 as well as between each of these mutants to mucilage-modified 2 (mum2) suggest that COBL2 functions independently of the FEI-SOS pathway. Altogether, the presented data place COBL2 within the complex protein network required for cell wall deposition in the context of seed mucilage and introduce new methodology expending the seed mucilage phenotyping toolbox.

Background and Aims Stomatal morphology and function have remained largely conserved throughout ∼400 million years of plant evolution. However, plant cell wall composition has evolved and changed. Here stomatal cell wall composition was investigated in different vascular plant groups in attempt to understand their possible effect on stomatal function. Methods A renewed look at stomatal cell walls was attempted utilizing digitalized polar microscopy, confocal microscopy, histology and a numerical finite-elements simulation. The six species of vascular plants chosen for this study cover a broad structural, ecophysiological and evolutionary spectrum: ferns (Asplenium nidus and Platycerium bifurcatum) and angiosperms (Arabidopsis thaliana and Commelina erecta) with kidney-shaped stomata, and grasses (angiosperms, family Poaceae) with dumbbell-shaped stomata (Sorghum bicolor and Triticum aestivum). Key Results Three distinct patterns of cellulose crystallinity in stomatal cell walls were observed: Type I (kidney-shaped stomata, ferns), Type II (kidney-shaped stomata, angiosperms) and Type III (dumbbell-shaped stomata, grasses). The different stomatal cell wall attributes investigated (cellulose crystallinity, pectins, lignin, phenolics) exhibited taxon-specific patterns, with reciprocal substitution of structural elements in the end-walls of kidneyshaped stomata. According to a numerical bio-mechanical model, the end walls of kidney-shaped stomata develop the highest stresses during opening. Conclusions The data presented demonstrate for the first time the existence of distinct spatial patterns of varying cellulose crystallinity in guard cell walls. It is also highly intriguing that in angiosperms crystalline cellulose appears to have replaced lignin that occurs in the stomatal end-walls of ferns serving a similar wall strengthening function. Such taxon-specific spatial patterns of cell wall components could imply different biomechanical functions, which in turn could be a consequence of differences in environmental selection along the course of plant evolution. © The Author 2017.

ABSTRACTMost floating aquatic plants have stomata on their upper leaf surfaces, and usually their stomata are permanently open. We previously identified 3 distinct crystallinity patterns in stomatal cell walls, with angiosperm kidney-shaped stomata having the highest crystallinity in the polar end walls as well as the adjacent polar regions of the guard cells. A numerical bio-mechanical model suggested that the high crystallinity areas are localized to regions where the highest stress is imposed. Here, stomatal cell wall crystallinity was examined in 4 floating plants from 2 different taxa: basal angiosperms from the ANITA grade and monocots. It appears that the non-functional stomata of floating plants display reduced crystallinity in the polar regions as compared with high crystallinity of the ventral (inner) walls. Thus their guard cells are both less flexible and less stress resistant. Our findings suggest that the pattern of cellulose crystallinity in stomata of floating plants from different families was altered as a consequence of similar evolutionary pressures.

Background and Aims: Stomatal morphology and function have remained largely conserved throughout ∼400 million years of plant evolution. However, plant cell wall composition has evolved and changed. Here stomatal cell wall composition was investigated in different vascular plant groups in attempt to understand their possible effect on stomatal function. Methods: A renewed look at stomatal cell walls was attempted utilizing digitalized polar microscopy, confocal microscopy, histology and a numerical finite-elements simulation. The six species of vascular plants chosen for this study cover a broad structural, ecophysiological and evolutionary spectrum: ferns ( Asplenium nidus and Platycerium bifurcatum ) and angiosperms ( Arabidopsis thaliana and Commelina erecta ) with kidney-shaped stomata, and grasses (angiosperms, family Poaceae) with dumbbell-shaped stomata ( Sorghum bicolor and Triticum aestivum ). Key Results: Three distinct patterns of cellulose crystallinity in stomatal cell walls were observed: Type I (kidney-shaped stomata, ferns), Type II (kidney-shaped stomata, angiosperms) and Type III (dumbbell-shaped stomata, grasses). The different stomatal cell wall attributes investigated (cellulose crystallinity, pectins, lignin, phenolics) exhibited taxon-specific patterns, with reciprocal substitution of structural elements in the end-walls of kidney-shaped stomata. According to a numerical bio-mechanical model, the end walls of kidney-shaped stomata develop the highest stresses during opening. Conclusions: The data presented demonstrate for the first time the existence of distinct spatial patterns of varying cellulose crystallinity in guard cell walls. It is also highly intriguing that in angiosperms crystalline cellulose appears to have replaced lignin that occurs in the stomatal end-walls of ferns serving a similar wall strengthening function. Such taxon-specific spatial patterns of cell wall components could imply different biomechanical functions, which in turn could be a consequence of differences in environmental selection along the course of plant evolution.

Differentiation of the maternally derived seed coat epidermal cells into mucilage secretory cells is a common adaptation in angiosperms. Recent studies identified cellulose as an important component of seed mucilage in various species. Cellulose is deposited as a set of rays that radiate from the seed upon mucilage extrusion, serving to anchor the pectic component of seed mucilage to the seed surface. Using transcriptome data encompassing the course of seed development, we identified COBRA-LIKE2 (COBL2), a member of the glycosylphosphatidylinositol-anchored COBRA-LIKE gene family in Arabidopsis (Arabidopsis thaliana), as coexpressed with other genes involved in cellulose deposition in mucilage secretory cells. Disruption of the COBL2 gene results in substantial reduction in the rays of cellulose present in seed mucilage, along with an increased solubility of the pectic component of the mucilage. Light birefringence demonstrates a substantial decrease in crystalline cellulose deposition into the cellulosic rays of the cobl2 mutants. Moreover, crystalline cellulose deposition into the radial cell walls and the columella appears substantially compromised, as demonstrated by scanning electron microscopy and in situ quantification of light birefringence. Overall, the cobl2 mutants display about 40% reduction in whole-seed crystalline cellulose content compared with the wild type. These data establish that COBL2 plays a role in the deposition of crystalline cellulose into various secondary cell wall structures during seed coat epidermal cell differentiation.