CRX ChIP-seq reveals the cis-regulatory architecture of mouse photoreceptors

Abstract

Approximately 98% of mammalian DNA is noncoding, yet we understand relatively little about the function of this enigmatic portion of the genome. The cis-regulatory elements that control gene expression reside in noncoding regions and can be identified by mapping the binding sites of tissue-specific transcription factors. Cone-rod homeobox (CRX) is a key transcription factor in photoreceptor differentiation and survival, but its in vivo targets are largely unknown. Here, we used chromatin immunoprecipitation with massively parallel sequencing (ChIP-seq) on CRX to identify thousands of cis-regulatory regions around photoreceptor genes in adult mouse retina. CRX directly regulates downstream photoreceptor transcription factors and their target genes via a network of spatially distributed regulatory elements around each locus. CRX-bound regions act in a synergistic fashion to activate transcription and contain multiple CRX binding sites which interact in a spacing-and orientation-dependent manner to fine-tune transcript levels. CRX ChIP-seq was also performed on Nrl -/-retinas, which represent an enriched source of cone photoreceptors. Comparison with the wild-type ChIP-seq data set identified numerous rod-and cone-specific CRX-bound regions as well as many shared elements. Thus, CRX combinatorially orchestrates the transcriptional networks of both rods and cones by coordinating the expression of photoreceptor genes including most retinal disease genes. In addition, this study pinpoints thousands of noncoding regions of relevance to both Mendelian and complex retinal disease.

Active opsin loci adopt intrachromosomal loops that depend on the photoreceptor transcription factor network

“…Using ChIP-qPCR, we show that three transcription factors, CRX, NRL, and NR2E3, differentially bind to rod vs. cone opsin genes in a subtype-specific manner. CRX preferentially binds to each opsin in the respective photoreceptor subtype expressing it, in agreement with CRX ChIP-seq results (33) and consistent with CRX's role as a trans-activator. In contrast to the discrete binding sites detected by ChIP-seq, ChIPqPCR revealed that CRX is bound along the entire length of each active opsin locus.…”

The long noncoding RNA Vax2os1 controls the cell cycle progression of photoreceptor progenitors in the mouse retina

“…2, 3), provided preliminary evidence that it may represent a putative enhancer element, similar to what has been previously reported for other conserved sequences (Pennacchio et al 2006). Moreover, a previous finding of a binding site for CRX validated by ChIP-seq, within the Vax2os1 conserved region, seems to further support this hypothesis (Corbo et al 2010). CRX (Cone-Rod Homeobox) is a transcription factor known to play an important role in cone and rod maturation by regulating the expression of a number of photoreceptorspecific genes (Chen et al 1997;Freund et al 1997;Furukawa et al 1997).…”

Early-onset autosomal recessive cerebellar ataxia associated with retinal dystrophy: new human hotfoot phenotype caused by homozygous GRID2 deletion

“…30 Indeed, two cis-regulatory elements bound by CRX are located in the GRID2 region, and one of them is removed by the homozygous deletion in our proband (Supplementary Figure S4 online). Although we cannot exclude RD and concordant ERG changes in the GRID2 deletion mutants based on the clinical data reported by Utine et al, 4 Hills et al, 5 and Maier et al, 6 the absence of a retinal phenotype in these patients might be explained by intact cis-regulatory elements bound by CRX on at least one of the two alleles.…”