Targeted inactivation and identification of targets of the Gli2a transcription factor in the zebrafish.
Hedgehog (Hh) signaling is mediated by the Gli transcription factors and, in the zebrafish, plays an important role in patterning both the neural tube and myotome. Using a null allele of the gli2a gene induced by targeted mutagenesis, we show that Gli2a is completely dispensable in the fish but acts redundantly with Gli1 to regulate expression of known Hh targets, such as ptch2, prdm1a and eng2a, in the myotome and neural tube. To identify novel targets of Hh signaling, we performed chromatin immunoprecipitation sequencing (ChIP-seq) of whole embryo extracts. Samples were significantly enriched for 192 genomic regions, some of which are associated with four known Hh target genes, ptch1, ptch2, gli1 and olig2. Sequence analysis of these regions reveals a high level of conservation of Gli-binding sites from fish to mammals in some, but not all, cases. Expression analysis of other transcription units that are closely associated with peaks identified several putative targets not previously implicated as Hh targets, including myl10, hnmt, lrp4, efemp2, fras1, quo, and lamc1. Each of these genes shows loss of, or reduced expression in, embryos homozygous for an antimorphic allele of gli2a, you-too (yot), consistent with their being direct targets of Gli2a.
Institute of Molecular and Cell Biology , 61 Biopolis Drive , Singapore 138673 ; Present address: High Throughput Molecular Drug Discovery Center, Tianjin International Joint Academy of Biotechnology and Medicine, Tianjin, China.
Xingang
X
Wang
Zhonghua
Z
Zhao
Julius
J
Muller
Audrey
A
Iyu
Alexis Jiaying
AJ
Khng
Ernesto
E
Guccione
Yijun
Y
Ruan
Philip W
PW
Ingham
ChIP-seq; Gli transcription factor; Hedgehog signaling; Target genes; Zebrafish; gli2a null allele