Mosaic analysis and tumor induction in zebrafish by microsatellite instability-mediated stochastic gene expression
- Authors
- Koole, W., and Tijsterman, M.
- ID
- ZDB-PUB-140410-8
- Date
- 2014
- Source
- Disease models & mechanisms 7(7): 929-36 (Journal)
- Registered Authors
- Keywords
- none
- MeSH Terms
-
- Green Fluorescent Proteins/metabolism
- Humans
- Animals, Genetically Modified
- Mosaicism/embryology*
- Genes, Reporter
- PubMed
- 24487406 Full text @ Dis. Model. Mech.
Mosaic analysis, in which two or more populations of cells with differing genotypes are studied in a single animal, is a powerful approach to study developmental mechanisms and gene function in vivo. Over recent years several genetic methods have been developed to achieve mosaicism in zebrafish, but despite their advances, limitations remain and different approaches and further refinements are warranted. Here, we describe an alternative approach to create somatic mosaicism in zebrafish that relies on the instability of microsatellite sequences during replication. We placed the coding sequences of various marker proteins behind a microsatellite and out-of-frame; in vivo frameshifting into the proper reading frame results in expression of the protein in random individual cells that are surrounded by wildtype cells. We optimized this approach for the binary Gal4-UAS expression system by generating a driver line and effector lines that stochastically express Gal4-VP16 or UAS:H2A-EGFP and self-maintaining UAS:H2A-EGFP-Kaloop, respectively. To demonstrate the utility of this system we stochastically expressed a constitutively active form of the human oncogene H-RAS and show the occurrence of hyperpigmentation and sporadic tumors within 5 days. Our data demonstrate that inducing somatic mosaicism via microsatellite instability can be a valuable approach for mosaic analysis and tumor induction in Danio rerio.
