Photoconvertible fluorescent proteins: A versatile tool in zebrafish skeletal imaging

Bek, J.W., De Clercq, A., De Saffel, H., Soenens, M., Huysseune, A., Witten, P.E., Coucke, P., Willaert, A.
Journal of Fish Biology   98(4): 1007-1017 (Journal)
Registered Authors
Coucke, Paul, Huysseune, Ann, Willaert, Andy, Witten, P. Eckhard
Kaede, Photoconvertible fluorescent protein, Zebrafish, osterix, skeletal imaging
MeSH Terms
  • Animals
  • Animals, Genetically Modified
  • Fluorescent Dyes/metabolism
  • Green Fluorescent Proteins/genetics
  • Green Fluorescent Proteins/metabolism
  • Luminescent Proteins/genetics*
  • Luminescent Proteins/metabolism*
  • Optical Imaging*
  • Zebrafish/metabolism*
  • Zebrafish Proteins/genetics
  • Zebrafish Proteins/metabolism
32242924 Full text @ J. Fish Biol.
One of the most frequently applied techniques in zebrafish (D. rerio) research is the visualisation or manipulation of specific cell populations using transgenic reporter lines. The generation of these transgenic zebrafish, displaying cell- or tissue- specific expression of frequently used fluorophores such as GFP or mCherry, is relatively easy with modern techniques. Fluorophores with different emission wavelengths and driven by different promoters can be monitored simultaneously in the same animal. Photoconvertible fluorescent proteins (pcFPs) are different from these standard fluorophores because their emission spectrum is changed when exposed to UV-light, a process called photoconversion. Here, we illustrate the benefits and versatility of using pcFPs for both single and dual fluorochrome imaging in zebrafish skeletal research in a previously generated osx:Kaede transgenic line. In this line, Kaede, which is expressed under control of the osterix, otherwise known as sp7, promoter thereby labelling immature osteoblasts, can switch from green to red fluorescence upon irradiation with UV-light. First, we demonstrate that osx:Kaede shows an expression pattern similar to a previously described osx:nuGFP transgenic line in both larval and adult stages, hereby validating the use of this line for the imaging of immature osteoblasts. More in-depth experiments highlight different applications for osx:Kaede, such as lineage tracing and its combined use with in vivo skeletal staining and other transgenic backgrounds. Mineral staining in combination with osx:Kaede confirms osteoblast independent mineralisation of the notochord. Osteoblast lineage tracing reveals migration and dedifferentiation of scleroblasts during fin regeneration. Finally, we show that combining two transgenics, osx:Kaede and osc:GFP with similar emission wavelengths is possible when using a pcFP such as Kaede. This article is protected by copyright. All rights reserved.
Genes / Markers
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes