Limits...
A discrete transition zone organizes the topological and regulatory autonomy of the adjacent tfap2c and bmp7 genes.

Tsujimura T, Klein FA, Langenfeld K, Glaser J, Huber W, Spitz F - PLoS Genet. (2015)

Bottom Line: The impact of engineered chromosomal rearrangements on the topology of the locus and the resultant gene expression changes indicate that this transition zone functionally organizes the structural partition of the locus, thereby defining enhancer-target gene allocation.This partition is, however, not absolute: we show that it allows competing interactions across it that may be non-productive for the competing gene, but modulate expression of the competed one.Altogether, these data highlight the prime role of the topological organization of the genome in long-distance regulation of gene expression.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Despite the well-documented role of remote enhancers in controlling developmental gene expression, the mechanisms that allocate enhancers to genes are poorly characterized. Here, we investigate the cis-regulatory organization of the locus containing the Tfap2c and Bmp7 genes in vivo, using a series of engineered chromosomal rearrangements. While these genes lie adjacent to one another, we demonstrate that they are independently regulated by distinct sets of enhancers, which in turn define non-overlapping regulatory domains. Chromosome conformation capture experiments reveal a corresponding partition of the locus in two distinct structural entities, demarcated by a discrete transition zone. The impact of engineered chromosomal rearrangements on the topology of the locus and the resultant gene expression changes indicate that this transition zone functionally organizes the structural partition of the locus, thereby defining enhancer-target gene allocation. This partition is, however, not absolute: we show that it allows competing interactions across it that may be non-productive for the competing gene, but modulate expression of the competed one. Altogether, these data highlight the prime role of the topological organization of the genome in long-distance regulation of gene expression.

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The Tfap2c-Bmp7 locus consists of two regulatory domains.(A) A schematic representation of the Tfap2c-Bmp7 locus, including the position of the SBlac insertions. Gene bodies are depicted as grey boxes with darker bars representing their exons. Tfap2c and Bmp7 are blue and green, respectively. The centromeric (CEN) and telomeric (TEL) ends of the chromosome are to the left and right of the diagram, respectively. The Sleeping Beauty transposon carrying a loxP site and LacZ reporter was first targeted into the immediate downstream region of Bmp7 along with an additional loxP sequence. Integration sites obtained upon remobilisation of the transposon are indicated by black arrowheads. (B) LacZ staining patterns of the transposon lines in the heart, limb, forebrain and the jaw as well as the staining of the BA0758 gene trap line in the forebrain and jaw are shown. Limbs: E12.5 embryos; other tissues: E11.5 embryos. Note that the intensity of the LacZ staining in the lateral and medial parts of the forebrain varies among BA0758, SB-A1 and SB-A2, as indicated by the blue arrows. Additional stages and views available in S1 Fig.
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pgen-1004897-g001: The Tfap2c-Bmp7 locus consists of two regulatory domains.(A) A schematic representation of the Tfap2c-Bmp7 locus, including the position of the SBlac insertions. Gene bodies are depicted as grey boxes with darker bars representing their exons. Tfap2c and Bmp7 are blue and green, respectively. The centromeric (CEN) and telomeric (TEL) ends of the chromosome are to the left and right of the diagram, respectively. The Sleeping Beauty transposon carrying a loxP site and LacZ reporter was first targeted into the immediate downstream region of Bmp7 along with an additional loxP sequence. Integration sites obtained upon remobilisation of the transposon are indicated by black arrowheads. (B) LacZ staining patterns of the transposon lines in the heart, limb, forebrain and the jaw as well as the staining of the BA0758 gene trap line in the forebrain and jaw are shown. Limbs: E12.5 embryos; other tissues: E11.5 embryos. Note that the intensity of the LacZ staining in the lateral and medial parts of the forebrain varies among BA0758, SB-A1 and SB-A2, as indicated by the blue arrows. Additional stages and views available in S1 Fig.

Mentions: To determine the regulatory organization of the Tfap2c-Bmp7 locus, we adapted the GROMIT (Genome Regulatory Organization Mapping with Integrated Transposons) strategy [14]. Firstly, at the 3′ end of the endogenous Bmp7 gene, we inserted a transgene consisting of a Sleeping Beauty transposon comprising 1) a regulatory sensor gene (a LacZ reporter under the control of a short naïve synthetic promoter region derived from the human β-globin gene [14], [50]) and 2) a loxP site. After establishment of a mouse line carrying the correct insertion, we removed the selection marker used to identify candidate targeted ES clones, a step which left behind an additional loxP site, next to the Sleeping Beauty transposon. We designated this allele as SB-B(3end) (Fig. 1). By serial remobilisation of the transposon in vivo[14], we obtained several insertions located in this region of mouse chromosome 2 (S1 Table). Of these, seven insertions were distributed along the Tfap2c-Bmp7 locus (Fig. 1A): three very close to SB-B(3end) (within 23 kb), one (SB-B(up)) 20 kb upstream of Bmp7, and another one in the first intron of Bmp7 (SB-B(in)). The remaining two (SB-A1 and SB-A2) lie within the large intergenic region separating Tfap2c and Bmp7. In parallel, we established a mouse line (BA0758) from an ES clone carrying a βgeo gene-trap insertion in Tfap2c[51].


A discrete transition zone organizes the topological and regulatory autonomy of the adjacent tfap2c and bmp7 genes.

Tsujimura T, Klein FA, Langenfeld K, Glaser J, Huber W, Spitz F - PLoS Genet. (2015)

The Tfap2c-Bmp7 locus consists of two regulatory domains.(A) A schematic representation of the Tfap2c-Bmp7 locus, including the position of the SBlac insertions. Gene bodies are depicted as grey boxes with darker bars representing their exons. Tfap2c and Bmp7 are blue and green, respectively. The centromeric (CEN) and telomeric (TEL) ends of the chromosome are to the left and right of the diagram, respectively. The Sleeping Beauty transposon carrying a loxP site and LacZ reporter was first targeted into the immediate downstream region of Bmp7 along with an additional loxP sequence. Integration sites obtained upon remobilisation of the transposon are indicated by black arrowheads. (B) LacZ staining patterns of the transposon lines in the heart, limb, forebrain and the jaw as well as the staining of the BA0758 gene trap line in the forebrain and jaw are shown. Limbs: E12.5 embryos; other tissues: E11.5 embryos. Note that the intensity of the LacZ staining in the lateral and medial parts of the forebrain varies among BA0758, SB-A1 and SB-A2, as indicated by the blue arrows. Additional stages and views available in S1 Fig.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4288730&req=5

pgen-1004897-g001: The Tfap2c-Bmp7 locus consists of two regulatory domains.(A) A schematic representation of the Tfap2c-Bmp7 locus, including the position of the SBlac insertions. Gene bodies are depicted as grey boxes with darker bars representing their exons. Tfap2c and Bmp7 are blue and green, respectively. The centromeric (CEN) and telomeric (TEL) ends of the chromosome are to the left and right of the diagram, respectively. The Sleeping Beauty transposon carrying a loxP site and LacZ reporter was first targeted into the immediate downstream region of Bmp7 along with an additional loxP sequence. Integration sites obtained upon remobilisation of the transposon are indicated by black arrowheads. (B) LacZ staining patterns of the transposon lines in the heart, limb, forebrain and the jaw as well as the staining of the BA0758 gene trap line in the forebrain and jaw are shown. Limbs: E12.5 embryos; other tissues: E11.5 embryos. Note that the intensity of the LacZ staining in the lateral and medial parts of the forebrain varies among BA0758, SB-A1 and SB-A2, as indicated by the blue arrows. Additional stages and views available in S1 Fig.
Mentions: To determine the regulatory organization of the Tfap2c-Bmp7 locus, we adapted the GROMIT (Genome Regulatory Organization Mapping with Integrated Transposons) strategy [14]. Firstly, at the 3′ end of the endogenous Bmp7 gene, we inserted a transgene consisting of a Sleeping Beauty transposon comprising 1) a regulatory sensor gene (a LacZ reporter under the control of a short naïve synthetic promoter region derived from the human β-globin gene [14], [50]) and 2) a loxP site. After establishment of a mouse line carrying the correct insertion, we removed the selection marker used to identify candidate targeted ES clones, a step which left behind an additional loxP site, next to the Sleeping Beauty transposon. We designated this allele as SB-B(3end) (Fig. 1). By serial remobilisation of the transposon in vivo[14], we obtained several insertions located in this region of mouse chromosome 2 (S1 Table). Of these, seven insertions were distributed along the Tfap2c-Bmp7 locus (Fig. 1A): three very close to SB-B(3end) (within 23 kb), one (SB-B(up)) 20 kb upstream of Bmp7, and another one in the first intron of Bmp7 (SB-B(in)). The remaining two (SB-A1 and SB-A2) lie within the large intergenic region separating Tfap2c and Bmp7. In parallel, we established a mouse line (BA0758) from an ES clone carrying a βgeo gene-trap insertion in Tfap2c[51].

Bottom Line: The impact of engineered chromosomal rearrangements on the topology of the locus and the resultant gene expression changes indicate that this transition zone functionally organizes the structural partition of the locus, thereby defining enhancer-target gene allocation.This partition is, however, not absolute: we show that it allows competing interactions across it that may be non-productive for the competing gene, but modulate expression of the competed one.Altogether, these data highlight the prime role of the topological organization of the genome in long-distance regulation of gene expression.

View Article: PubMed Central - PubMed

Affiliation: Developmental Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Despite the well-documented role of remote enhancers in controlling developmental gene expression, the mechanisms that allocate enhancers to genes are poorly characterized. Here, we investigate the cis-regulatory organization of the locus containing the Tfap2c and Bmp7 genes in vivo, using a series of engineered chromosomal rearrangements. While these genes lie adjacent to one another, we demonstrate that they are independently regulated by distinct sets of enhancers, which in turn define non-overlapping regulatory domains. Chromosome conformation capture experiments reveal a corresponding partition of the locus in two distinct structural entities, demarcated by a discrete transition zone. The impact of engineered chromosomal rearrangements on the topology of the locus and the resultant gene expression changes indicate that this transition zone functionally organizes the structural partition of the locus, thereby defining enhancer-target gene allocation. This partition is, however, not absolute: we show that it allows competing interactions across it that may be non-productive for the competing gene, but modulate expression of the competed one. Altogether, these data highlight the prime role of the topological organization of the genome in long-distance regulation of gene expression.

Show MeSH
Related in: MedlinePlus