# Long-term labelling and tracing of endodermal cells using a perpetual cycling Gal4-UAS system

## TL;DR

A new system allows long-term tracking of endodermal cells in zebrafish, from embryos to adults, using continuous fluorescent labeling.

## Contribution

A perpetual cycling Gal4-UAS system with a novel Gal4 fusion protein enables long-term cell labeling and tracing.

## Key findings

- The perpetual cycling Gal4-UAS system ensures sustained reporter gene expression with minimal cytotoxicity.
- Zebrafish transgenic lines demonstrate effective long-term endoderm tracing from embryos to adults.
- The system visualizes endodermal differentiation from progenitor cells to mature tissues.

## Abstract

Cell labelling and lineage tracing are indispensable tools in developmental biology, offering powerful means with which to visualise and understand the complex dynamics of cell populations during embryogenesis. Traditional cell labelling relies heavily on signal stability, promoter strength and stage specificity, limiting its application in long-term tracing. In this report, we optimise and reconfigure a perpetual cycling Gal4-UAS system employing a previously unreported Gal4 fusion protein and the autoregulatory Gal4 expression loop. As validated through heat-shock induction, this configuration ensures sustained transcription of reporter genes in target cells and their descendant cells while minimising cytotoxicity, thereby achieving long-term labelling and tracing. Further exploiting this system, we generate zebrafish transgenic lines with continuous fluorescent labelling specific to the endoderm, and demonstrate its effectiveness in long-term tracing by showing the progression of endoderm development from embryo to adult, providing visualisation of endodermal cells and their derived tissues. This continuous labelling and tracing strategy can span the entire process of endodermal differentiation, from progenitor cells to mature functional cells, and is applicable to studying endoderm patterning and organogenesis.

Summary: A continuous fluorescent labelling strategy for long-term cell tracing, enabling effective visualisation of endodermal cells and their development from embryos to adults.