The transposon-mediated Gal4 gene trapping identified genes expressing in unique patterns in the adult zebrafish

Mizusawa, K 1 , Asakawa, K 1 , Urasaki, A 1 , Kotani, T 1 , Nagayoshi, S 1 , Kishimoto, Y 1 , Kondo, S 2 and Kawakami, K 1

1 National Institute of Genetics, Japan, 2 Nagoya University, Japan

 Gene-trap and enhancer trap methods have recently been developed in zebrafish. The previous studies have shown that these methods are powerful to study the function of developmental genes. However, these methods have not been applied to the study of genes functioning in the adult stages. To identify and characterize genes that are expressed in spatially restricted patterns in the adult and play important roles to maintain the adult system, we have developed the Gal4-gene trap system in zebrafish and performed a screen for unique Gal4 expression patterns in the adult.
 First, 247 fish injected with the Gal4 gene trap transposon construct-donor plasmid and transposase mRNA were mated with the UAS-GFP reporter fish, and the offspring were screened for unique GFP expression during embryonic development. 154 unique GFP expression patterns were identified, and the F1 fish were raised. Then, 1252 F1 fish with these 154 embryonic patterns were analyzed for unique GFP expression at the adult stages. We observed unique GFP expression in various organs; i.e., eye, nose, jaw, fin, skin, gill, muscle, skeleton and part of the brain, and, in total, 121 unique GFP expression patterns were identified. To determine whether the prescreen at the embryo stages enriched fish with unique GFP expression patterns at the adult stage, we performed the adult screen without the embryonic prescreen. A total of 1984 F1 fish from 99 injected fish were analyzed at the adult stage, and 79 unique GFP expression patterns were identified. Thus, the adult screen with the embryonic prescreen (79%:121/154) was comparable with that without the prescreen (80%:79/99), suggesting that the adult and the embryonic patterns may be irrelevant.
 We are now focusing on fish lines exhibiting unique Gal4 expression in different parts of the adult brain (telencephalon, diencephalon, and cerebellum) and in the pigment cells on the skin (the inter-stripe and the stripe boundary regions). Further studies of these lines should disclose genes functioning these organs. Also, these Gal4 lines will be useful to modify the activities of those cells in the adult zebrafish.