ChIP-Seq identification of weakly conserved heart enhancers

Blow, Matthew J.; McCulley, David J.; Li, Zirong; Zhang, Tao; Akiyama, Jennifer A.; Holt, Amy; Plajzer-Frick, Ingrid; Shoukry, Malak; Wright, Crystal; Chen, Feng; Afzal, Veena; Bristow, James; Ren, Bing; Black, Brian L.; Rubin, Edward M.; Visel, Axel; Pennacchio, Len A.

doi:10.1038/ng.650

Cited by 399 publications

(458 citation statements)

References 44 publications

Supporting

Mentioning

415

Contrasting

Unclassified

Order By: Relevance

“…Concerning the latter strategy, many investigations in recent years have used genome-wide chromatin immunoprecipitation to map epigenetic marks that are typical for enhancers and, in some cases, to test the elements in transgenic mouse analysis (reviewed in [24,25]). Such studies have detected a large proportion of enhancers that are conserved among several mammals but also a significant fraction of enhancers without apparent evolutionary constraint [22,26,27]. It remains to be tested, however, the relative importance of conserved versus weakly-or non-conserved enhancers in the regulation of the genes they control.…”

Section: (A) Enhancer Evolution In Vertebratesmentioning

confidence: 99%

Enhancer turnover and conserved regulatory function in vertebrate evolution

Domené

Bumaschny

Souza

et al. 2013

Phil. Trans. R. Soc. B

View full text Add to dashboard Cite

Mutations in regulatory regions including enhancers are an important source of variation and innovation during evolution. Enhancers can evolve by changes in the sequence, arrangement and repertoire of transcription factor binding sites, but whole enhancers can also be lost or gained in certain lineages in a process of turnover. The proopiomelanocortin gene ( Pomc ), which encodes a prohormone, is expressed in the pituitary and hypothalamus of all jawed vertebrates. We have previously described that hypothalamic Pomc expression in mammals is controlled by two enhancers—nPE1 and nPE2—that are derived from transposable elements and that presumably replaced the ancestral neuronal Pomc regulatory regions. Here, we show that nPE1 and nPE2, even though they are mammalian novelties with no homologous counterpart in other vertebrates, nevertheless can drive gene expression specifically to POMC neurons in the hypothalamus of larval and adult transgenic zebrafish. This indicates that when neuronal Pomc enhancers originated de novo during early mammalian evolution, the newly created cis- and trans- codes were similar to the ancestral ones. We also identify the neuronal regulatory region of zebrafish pomca and confirm that it is not homologous to the mammalian enhancers. Our work sheds light on the process of gene regulatory evolution by showing how a locus can undergo enhancer turnover and nevertheless maintain the ancestral transcriptional output.

show abstract

Section: (A) Enhancer Evolution In Vertebratesmentioning

confidence: 99%

Enhancer turnover and conserved regulatory function in vertebrate evolution

Domené

Bumaschny

Souza

et al. 2013

Phil. Trans. R. Soc. B

View full text Add to dashboard Cite

show abstract

“…2C). 4. Cut the tip off a P200 pipette tip, fit the tip to a 3ml syringe, and fill the syringe with 100% silicone rubber aquarium sealant.…”

mentioning

confidence: 99%

“…Use sterile forceps to place round Whatman Nuclepore filters, shiny side up, atop the medium in each well. 4. Under a dissecting microscope, use a sterile transfer pipette to transfer the retinas onto the filter, lens-side-down.…”

mentioning

confidence: 99%

Quantifying the Activity of cis-Regulatory Elements in the Mouse Retina by Explant Electroporation

Montana

Myers

Corbo

2012

Methods in Molecular Biology

View full text Add to dashboard Cite

Transcription factors within cellular gene networks control the spatiotemporal pattern and levels of expression of their target genes by binding to cis-regulatory elements (CREs), short (˜300-600 bp) stretches of genomic DNA which can lie upstream, downstream, or within the introns of the genes they control. CREs (i.e., enhancers/promoters) typically consist of multiple clustered binding sites for both transcriptional activators and repressors 1-3 . They serve as logical integrators of transcriptional input giving a unitary output in the form of spatiotemporally precise and quantitatively exact promoter activity. Most studies of mammalian cis-regulation to date have relied on mouse transgenesis as a means of assaying the enhancer function of CREs 4-5 . This technique is time-consuming, costly and, on account of insertion site effects, largely non-quantitative. On the other hand, quantitative assays for mammalian CRE function have been developed in tissue culture systems (e.g., dual luciferase assays), but the in vivo relevance of these results is often uncertain.Electroporation offers an excellent alternative to traditional mouse transgenesis in that it permits both spatiotemporal and quantitative assessment of cis-regulatory activity in living mammalian tissue. This technique has been particularly useful in the analysis of cis-regulation in the central nervous system, especially in the cerebral cortex and the retina 6-8 . While mouse retinal electroporation, both in vivo and ex vivo, has been developed and extensively described by Matsuda and Cepko 6-7,9 , we have recently developed a simple approach to quantify the activity of photoreceptor-specific CREs in electroporated mouse retinas 10 . Given that the amount of DNA that is introduced into the retina by electroporation can vary from experiment to experiment, it is necessary to include a co-electroporated 'loading control' in all experiments. In this respect, the technique is very similar to the dual luciferase assay used to quantify promoter activity in cultured cells.When assaying photoreceptor cis-regulatory activity, electroporation is usually performed in newborn mice (postnatal day 0, P0) which is the time of peak rod production [11][12] . Once retinal cell types become post-mitotic, electroporation is much less efficient. Given the high rate of rod birth in newborn mice and the fact that rods constitute more than 70% of the cells in the adult mouse retina, the majority of cells that are electroporated at P0 are rods. For this reason, rod photoreceptors are the easiest retinal cell type to study via electroporation. The technique we describe here is primarily useful for quantifying the activity of photoreceptor CREs. (Fig. 2A). The metal rails should be completely sealed to the bottom of the slide. 2. Use a Dremel tool to cut a handle off a plastic microcentrifuge tube rack. Cut the handle into 5 small rectangular pieces each with the following dimensions: length 0.8cm, height 0.6cm, width 0.3cm (Fig. 2B). These plastic pieces are reusable spa...

show abstract

“…However, neither the strong HS nor the rest of the intronic region fulfill the criteria to be considered a CNS (70% identity over 100 bp length). Therefore, and as described for other tissue-specific enhancers, 28 CD69 gene contains an intronic DNA regulatory region that is not sequence conserved under canonical basis.…”

Section: Resultsmentioning

confidence: 88%

Evidence for an intronic cis-regulatory element within CD69 gene

et al. 2012

View full text Add to dashboard Cite

CD69 is one of the earliest proteins expressed after leukocyte activation and its engagement is essential in the control of innate and adaptive immune responses. Inducible CD69 expression is strongly controlled at the transcriptional level. The molecular basis for developmental-and stage-specific regulation in T cells is beginning to be elucidated while it remains largely unknown in the rest of immune cells. DNase I hypersensitivity experiments in lymphocytes identified a novel hypersensitive region within mouse and human intron I, which was inducible upon stimulation. In silico analysis of CD69 gene revealed that this open chromatin region was present in different cell types and was associated with positioned nucleosomes. Analysis of histone post-translational modifications of intron I indicated that acetylation and lysine 4 dimethylation of histone H3 were dynamically regulated during thymocyte development and were constitutively high in resting and stimulated mature T lymphocytes. Thus, we provide evidence for the existence of a cis-acting element in intron I that is more accessible to DNase I digestion and that it is developmentally regulated at the chromatin level.

show abstract

ChIP-Seq identification of weakly conserved heart enhancers

Cited by 399 publications

References 44 publications

Enhancer turnover and conserved regulatory function in vertebrate evolution

Enhancer turnover and conserved regulatory function in vertebrate evolution

Quantifying the Activity of cis-Regulatory Elements in the Mouse Retina by Explant Electroporation

Evidence for an intronic cis-regulatory element within CD69 gene

Contact Info

Product

Resources

About