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Published March 14, 2012 | Published + Supplemental Material
Journal Article Open

PhOTO Zebrafish: A Transgenic Resource for In Vivo Lineage Tracing during Development and Regeneration

Abstract

Background: Elucidating the complex cell dynamics (divisions, movement, morphological changes, etc.) underlying embryonic development and adult tissue regeneration requires an efficient means to track cells with high fidelity in space and time. To satisfy this criterion, we developed a transgenic zebrafish line, called PhOTO, that allows photoconvertible optical tracking of nuclear and membrane dynamics in vivo. Methodology: PhOTO zebrafish ubiquitously express targeted blue fluorescent protein (FP) Cerulean and photoconvertible FP Dendra2 fusions, allowing for instantaneous, precise targeting and tracking of any number of cells using Dendra2 photoconversion while simultaneously monitoring global cell behavior and morphology. Expression persists through adulthood, making the PhOTO zebrafish an excellent tool for studying tissue regeneration: after tail fin amputation and photoconversion of a ~100µm stripe along the cut area, marked differences seen in how cells contribute to the new tissue give detailed insight into the dynamic process of regeneration. Photoconverted cells that contributed to the regenerate were separated into three distinct populations corresponding to the extent of cell division 7 days after amputation, and a subset of cells that divided the least were organized into an evenly spaced, linear orientation along the length of the newly regenerating fin. Conclusions/Significance: PhOTO zebrafish have wide applicability for lineage tracing at the systems-level in the early embryo as well as in the adult, making them ideal candidate tools for future research in development, traumatic injury and regeneration, cancer progression, and stem cell behavior.

Additional Information

© 2012 Dempsey et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Received December 18, 2011; Accepted February 1, 2012; Published March 14, 2012. Editor: Shree Ram Singh, National Cancer Institute, United States of America. Funding: WPD was supported by a Caltech Rosen Scholar Graduate fellowship. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We would like to thank Le Trinh and Sean Megason for sharing reagents and Le Trinh, Cambrian Liu, and Tatiana Hochgreb for assistance with the western blotting. We would like to thank Andres Collazo for access to the Confocal Microscopy Core at the House Research Institute, and Le Trinh and Christopher Dempsey for comments on the manuscript. Ethics Statement: The animal experiments conducted were covered under the animal license protocol number 1227 'Zebrafish Develomental Biology' approved by Caltech's Institutional Animal Care and Use Committee (IACUC). Author Contributions: Conceived and designed the experiments: WPD SEF PP. Performed the experiments: WPD PP. Analyzed the data: WPD PP. Wrote the paper: WPD SEF PP.

Attached Files

Published - Dempsey2012p18151PLoS_ONE.pdf

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Supplemental Material - VideoS2.mov

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Created:
August 19, 2023
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