Genome-wide discovery of Pax7 target genes during development

Abstract

Pax7 plays critical roles in development of brain, spinal cord, neural crest, and skeletal muscle. As a sequence-specific DNA-binding transcription factor, any direct functional role played by Pax7 during development is mediated through target gene selection. Thus, we have sought to identify genes targeted by Pax7 during embryonic development using an unbiased chromatin immunoprecipitation (ChIP) cloning assay to isolate cis-regulatory regions bound by Pax7 in vivo. Sequencing and genomic localization of a library of chromatin-DNA fragments bound by Pax7 has identified 34 candidate Pax7 target genes, with occupancy of a selection confirmed with independent chromatin enrichment tests (ChIP-PCR). To assess the capacity of Pax7 to regulate transcription from these loci, we have cloned alternate transcripts of Pax7 (differing significantly in their DNA binding domain) into expression vectors and transfected cultured cells with these constructs, then analyzed target gene expression levels using RT-PCR. We show that Pax7 directly occupies sites within genes encoding transcription factors Gbx1 and Eya4, the neurogenic cytokine receptor ciliary neurotrophic factor receptor, the neuronal potassium channel Kcnk2, and the signal transduction kinase Camk1d in vivo and regulates the transcriptional state of these genes in cultured cells. This analysis gives us greater insight into the direct functional role played by Pax7 during embryonic development.

Cell cycle regulation of embryonic stem cells and mouse embryonic fibroblasts lacking functional Pax7

“…Identification of Pax7 transcription factor targets in E16 mouse embryos revealed several genes involved in the regulation of proliferation 51 . Among them were genes positively impacting cell proliferation, such as CntfR , receptor of CNTF – a member of IL-6 family, which induces Jak/STAT and Ras/MAPK signaling, 52 Camk1d involved in calcium-calmodulin dependent signaling, 53 as well as Frk which also regulates cell cycle progression 54 .…”

The Cerebellin 4 Precursor Gene Is a Direct Target of SRY and SOX9 in Mice1

“…In this study, we identified Cbln4 as a direct target gene of SRY by employing in vivo ChIP analysis. In vivo ChIP is a powerful tool for studying protein-DNA interactions [50,51], offering the unique possibility of identifying novel target genes and their potential regulatory regions [52,53]. However, because of the occurrence of random protein-DNA cross-linking, nonspecific DNA fragments are likely to be precipitated together with the DNA genuinely bound by the protein of interest.…”

Genetic Biomarkers for ALS Disease in Transgenic SOD1G93A Mice

“…In skeletal muscle, many genes are involved in regenerative pathways, either by maintaining the integrity of the tissue and favoring skeletal myogenesis, as is the case for collagen, type XIX, alpha 1 ( Col19a1 ) [29], or by promoting muscle differentiation and regeneration, as is the case for the paired box gene ( Pax7 ), myogenic differentiation 1 ( Myod1 ), myogenic factor 5 ( Myf5 ), myocyte enhancer factor 2C ( Mef2c ) and myogenin ( Myog ) [30], [56]–[60].…”