Biology Reference
In-Depth Information
Chapter 7
Visualizing and Quantifying the In Vivo Structure
and Dynamics of the
Arabidopsis
Cortical Cytoskeleton
Using CLSM and VAEM
Amparo Rosero , Viktor Žárský , and Fatima Cvr cková
Abstract
The cortical microtubules, and to some extent also the actin meshwork, play a central role in the shaping
of plant cells. Transgenic plants expressing fl uorescent protein markers specifi cally tagging the two main
cytoskeletal systems are available, allowing noninvasive in vivo studies. Advanced microscopy techniques,
in particular confocal laser scanning microscopy (CLSM) and variable angle epifl uorescence microscopy
(VAEM), can be nowadays used for imaging the cortical cytoskeleton of living cells with unprecedented
spatial and temporal resolution. With the aid of suitable computing techniques, quantitative information
can be extracted from microscopic images and video sequences, providing insight into both architecture
and dynamics of the cortical cytoskeleton.
Key words
Actin, Microtubules, Fluorescent proteins, CLSM, VAEM, Image analysis
1
Introduction
Cortical microtubules are long known to play a major part in the
morphogenesis of plant cells, in particular due to their intimate
relationship with the biosynthesis of the cellulosic cell wall micro-
fi brils (
see
ref. [
1
]). However, the actin cytoskeleton, which under-
goes constant dynamic remodeling [
2
], is crucial for processes such
as trichome morphogenesis [
3
], tip growth in root hairs [
4
], or
development of epidermal cell lobes [
5
] and apparently contrib-
utes to the localization of exocytosis, affecting also the positioning
of cellulose synthase complexes [
1
,
6
]. Detailed characterization of
the spatial structure and temporal behavior of the two main cyto-
skeletal systems in vivo may thus substantially contribute to our
understanding of plant cell shaping.
Such studies depend on several prerequisites. Suffi ciently spe-
cifi c and nondisruptive fl uorescent cytoskeletal markers must be
introduced into the tissues of interest, and suitable high-resolution
imaging technology must be available, even if the aim of the study
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