A radioactive assay allowing the quantitative measurement of cuticular permeability of intact Arabidopsis thaliana leaves

Abstract Background The plant cuticle represents one of the major adaptations of vascular plants to terrestrial life. Cuticular permeability and chemical composition differ among species. Arabidopsis thaliana is a widely used model for biochemical and molecular genetic studies in plants. However, attempts to isolate the intact cuticle from fresh leaves of Arabidopsis have failed so far. The goal of this study was to optimise an enzymatic method for cuticle isolation of species with a thin cuticle and to test it on several A. thaliana wild types and mutants. Results We developed a method for isolation of thin cuticles that allows reducing the isolation time, the separation of abaxial and adaxial cuticles, and avoids formation of wrinkles. Optical microscopy was used for studying cuticle intactness and scanning electron microscopy for visualisation of external and internal cuticle structures after isolation. Wax extracts were analysed by GC–MS. Isolation of intact cuticle was successful for all tested plants. The wax compositions (very-long-chained fatty acids, alcohols and alkanes) of intact leaves and isolated cuticles of wild type Col-0 were compared. Conclusions We conclude that the optimised enzymatic method is suitable for the isolation of A. thaliana adaxial and abaxial cuticles. The isolated cuticles are suitable for microscopic observation. Analysis of wax composition revealed some discrepancies between isolated cuticles and intact leaves with a higher yield of wax in isolated cuticles.

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Arabidopsis thaliana Cuticle Composition Contributes to Differential Defense Response to Botrytis cinerea

Frontiers in Plant Science ◽

10.3389/fpls.2021.738949 ◽

2021 ◽

Vol 12 ◽

Author(s):

Wendy Aragón ◽

Damien Formey ◽

Norma Yaniri Aviles-Baltazar ◽

Martha Torres ◽

Mario Serrano

Keyword(s):

Arabidopsis Thaliana ◽

Botrytis Cinerea ◽

Molecular Mechanisms ◽

Defense Responses ◽

Biotic Stresses ◽

Solute Permeability ◽

Ros Accumulation ◽

Cuticular Permeability ◽

Induce Resistance ◽

Upregulated Genes

The chemical composition of a plant cuticle can change in response to various abiotic or biotic stresses and plays essential functions in disease resistance responses. Arabidopsis thaliana mutants altered in cutin content are resistant to Botrytis cinerea, presumably because of increased cuticular water and solute permeability, allowing for faster induction of defense responses. Within this context, our knowledge of wax mutants is limited against this pathogen. We tested the contribution of cuticular components to immunity to B. cinerea using mutants altered in either cutin or wax alone, or in both cutin and wax contents. We found that even all the tested mutants showed increased permeability and reactive oxygen species (ROS) accumulation in comparison with wild-type plants and that only cutin mutants showed resistance. To elucidate the early molecular mechanisms underlying cuticle-related immunity, we performed a transcriptomic analysis. A set of upregulated genes involved in cell wall integrity and accumulation of ROS were shared by the cutin mutants bdg, lacs2-3, and eca2, but not by the wax mutants cer1-4 and cer3-6. Interestingly, these genes have recently been shown to be required in B. cinerea resistance. In contrast, we found the induction of genes involved in abiotic stress shared by the two wax mutants. Our study reveals new insight that the faster recognition of a pathogen by changes in cuticular permeability is not enough to induce resistance to B. cinerea, as has previously been hypothesized. In addition, our data suggest that mutants with resistant phenotype can activate other defense pathways, different from those canonical immune ones.

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A quantitative image analysis method for the determination of cocontinuity in polymer blends

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100150307 ◽

1993 ◽

Vol 51 ◽

pp. 894-895

Author(s):

William A. Heeschen

Keyword(s):

Image Analysis ◽

Polymer Blends ◽

Quantitative Measurement ◽

Morphological Evolution ◽

Phase Ratio ◽

Irregular Shapes ◽

Quantitative Image ◽

Discrete Domains ◽

A Domains

Two new morphological measurements based on digital image analysis, CoContinuity and CoContinuity Balance, have been developed and implemented for quantitative measurement of morphology in polymer blends. The morphology of polymer blends varies with phase ratio, composition and processing. A typical morphological evolution for increasing phase ratio of polymer A to polymer B starts with discrete domains of A in a matrix of B (A/B < 1), moves through a cocontinuous distribution of A and B (A/B ≈ 1) and finishes with discrete domains of B in a matrix of A (A/B > 1). For low phase ratios, A is often seen as solid convex particles embedded in the continuous B phase. As the ratio increases, A domains begin to evolve into irregular shapes, though still recognizable as separate domains. Further increase in the phase ratio leads to A domains which extend into and surround the B phase while the B phase simultaneously extends into and surrounds the A phase.

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