Abstract
Motor proteins are responsible for transport of vesicles and organelles within the cell cytoplasm. They interact with the actin cytoskeleton and with microtubules to ensure communication and supply throughout the cell. Much work has been done in vitro and in silico to unravel the key players, including the dynein motor complex, the kinesin and myosin superfamilies, and their interacting regulatory complexes, but there is a clear need for in vivo data as recent evidence suggests previous models might not recapitulate physiological conditions. The zebrafish embryo provides an excellent system to study these processes in intact animals due to the ease of genetic manipulation and the optical transparency allowing live imaging. We present here the advantages of the zebrafish embryo as a system to study live in vivo processive transport in neurons and provide technical recommendations for successful analysis.
Highlights
Processive intracellular transport is essential for the distribution of organelles and cellular cargoes within the cell
The zebrafish embryo has emerged as an excellent model to pursue the characterization of processive transport in vivo as it can meet the need for more inclusive models, where the contribution of neuronal activity, glia and the cell cytoskeleton are taken into account
We outlined here some advantages and technical hints to use the zebrafish model for this type of analysis
Summary
Processive intracellular transport is essential for the distribution of organelles and cellular cargoes within the cell. Most approaches discussed here rely on the overexpression of fusion proteins, allowing in vivo detection of the bound fluorescent protein This can be achieved injecting DNA constructs to obtain single-cell labeling of cargoes, as shown here (Figure 1A), or creating stable transgenic lines, where restriction of expression can be achieved by a combination of Gal4- and UAS-expressing lines. While this technique produces a bright signal well suited to time-lapse imaging, overexpression of protein can lead to deleterious effects by interfering with endogenous expression and triggering stress response mechanisms (Cheng and Lee, 2010). Further advances in detection algorithms, based on in vivo data estimating how cargoes should behave, will surely be of benefit to researchers facing the tedious task of manual tracking
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