Photoconvertible fluorescent proteins: a versatile tool in zebrafish skeletal imaging

Area of Science:

• Zebrafish (Danio rerio) research
• Skeletal biology
• Developmental biology

Background:

• Transgenic zebrafish lines are crucial for visualizing and manipulating specific cell populations.
• Standard fluorophores like GFP and mCherry are widely used, but photoconvertible fluorescent proteins (pcFPs) offer unique advantages.
• Photoconversion allows a change in emission spectrum upon UV light exposure, enabling advanced imaging techniques.

Purpose of the Study:

• To illustrate the benefits and versatility of pcFPs for single and dual fluorochrome imaging in zebrafish skeletal research.
• To validate the osx:Kaede transgenic line for imaging immature osteoblasts.
• To explore applications of osx:Kaede in lineage tracing and combined imaging strategies.

Main Methods:

• Generation and characterization of the osx:Kaede transgenic zebrafish line.
• Comparison of osx:Kaede expression patterns with osx:nuGFP line.
• In vivo skeletal staining and dual-color imaging with other transgenic lines.
• Photoconversion of Kaede from green to red fluorescence using UV light.

Main Results:

• The osx:Kaede line accurately reflects the expression pattern of immature osteoblasts in larval and adult zebrafish.
• Mineral staining confirmed osteoblast-independent notochord mineralization.
• Osteoblast lineage tracing revealed scleroblast migration and dedifferentiation during fin regeneration.
• Successful dual-color imaging was achieved by combining osx:Kaede with an osc:GFP transgenic line.

Conclusions:

• The osx:Kaede transgenic line is a valuable tool for studying immature osteoblasts in zebrafish.
• Photoconvertible fluorescent proteins significantly enhance imaging capabilities in skeletal research.
• This study demonstrates the feasibility of combining pcFPs with other transgenic lines for complex biological investigations.