Dataset Information

Developmental Nuclear Localization and Quantification of GFP-Tagged EB1c in Arabidopsis Root Using Light-Sheet Microscopy.

ABSTRACT: The development of the root apex is determined by progress of cells from the meristematic region to the successive post-mitotic developmental zones for transition, cell elongation and final cell differentiation. We addressed root development, tissue architecture and root developmental zonation by means of light-sheet microscopic imaging of Arabidopsis thaliana seedlings expressing END BINDING protein 1c (EB1c) fused to green fluorescent protein (GFP) under control of native EB1c promoter. Unlike the other two members of the EB1 family, plant-specific EB1c shows prominent nuclear localization in non-dividing cells in all developmental zones of the root apex. The nuclear localization of EB1c was previously mentioned solely in meristematic cells, but not further addressed. With the help of advanced light-sheet microscopy, we report quantitative evaluations of developmentally-regulated nuclear levels of the EB1c protein tagged with GFP relatively to the nuclear size in diverse root tissues (epidermis, cortex, and endodermis) and root developmental zones (meristem, transition, and elongation zones). Our results demonstrate a high potential of light-sheet microscopy for 4D live imaging of fluorescently-labeled nuclei in complex samples such as developing roots, showing capacity to quantify parameters at deeper cell layers (e.g., endodermis) with minimal aberrations. The data presented herein further signify the unique role of developmental cell reprogramming in the transition from cell proliferation to cell differentiation in developing root apex.

SUBMITTER: Novak D

PROVIDER: S-EPMC4700127 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Developmental Nuclear Localization and Quantification of GFP-Tagged EB1c in Arabidopsis Root Using Light-Sheet Microscopy.

Novák Dominik D Kuchařová Anna A Ovečka Miroslav M Komis George G Šamaj Jozef J

Frontiers in plant science 20160105

The development of the root apex is determined by progress of cells from the meristematic region to the successive post-mitotic developmental zones for transition, cell elongation and final cell differentiation. We addressed root development, tissue architecture and root developmental zonation by means of light-sheet microscopic imaging of Arabidopsis thaliana seedlings expressing END BINDING protein 1c (EB1c) fused to green fluorescent protein (GFP) under control of native EB1c promoter. Unlike ...[more]

PMID: 26779221

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Project description:Phospholipase D alpha 1 (PLD?1, At3g15730) and its product phosphatidic acid (PA) are involved in a variety of cellular and physiological processes, such as cytoskeletal remodeling, regulation of stomatal closure and opening, as well as biotic and abiotic stress signaling. Here we aimed to study developmental expression patterns and subcellular localization of PLD?1 in Arabidopsis using advanced microscopy methods such as light-sheet fluorescence microscopy (LSFM) and structured illumination microscopy (SIM). We complemented two knockout pld?1 mutants with a YFP-tagged PLD?1 expressed under the PLD?1 native promoter in order to study developmental expression pattern and subcellular localization of PLD?1 in Arabidopsis thaliana under natural conditions. Imaging of tissue-specific and developmentally-regulated localization of YFP-tagged PLD?1 by LSFM in roots of growing seedlings showed accumulation of PLD?1-YFP in the root cap and the rhizodermis. Expression of PLD?1-YFP in the rhizodermis was considerably higher in trichoblasts before and during root hair formation and growth. Thus, PLD?1-YFP accumulated in emerging root hairs and in the tips of growing root hairs. PLD?1-YFP showed cytoplasmic subcellular localization in root cap cells and in cells of the root transition zone. In aerial parts of plants PLD?1-YFP was also localized in the cytoplasm showing enhanced accumulation in the cortical cytoplasmic layer of epidermal non-dividing cells of hypocotyls, leaves, and leaf petioles. However, in dividing cells of root apical meristem and leaf petiole epidermis PLD?1-YFP was enriched in mitotic spindles and phragmoplasts, as revealed by co-visualization with microtubules. Finally, super-resolution SIM imaging revealed association of PLD?1-YFP with both microtubules and clathrin-coated vesicles (CCVs) and pits (CCPs). In conclusion, this study shows the developmentally-controlled expression and subcellular localization of PLD?1 in dividing and non-dividing Arabidopsis cells.

Dataset Information

Developmental Nuclear Localization and Quantification of GFP-Tagged EB1c in Arabidopsis Root Using Light-Sheet Microscopy.

Publications

Developmental Nuclear Localization and Quantification of GFP-Tagged EB1c in Arabidopsis Root Using Light-Sheet Microscopy.

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