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Foxf genes integrate tbx5 and hedgehog pathways in the second heart field for cardiac septation.

Hoffmann AD, Yang XH, Burnicka-Turek O, Bosman JD, Ren X, Steimle JD, Vokes SA, McMahon AP, Kalinichenko VV, Moskowitz IP - PLoS Genet. (2014)

Bottom Line: GLI1 and TBX5 synergistically activated transcription from this cis-regulatory element in vitro.This enhancer drove reproducible expression in vivo in the posterior SHF, the only region where Gli1 and Tbx5 expression overlaps.Our findings implicate Foxf genes in atrioventricular septation, describe the molecular underpinnings of the genetic interaction between Hedgehog signaling and Tbx5, and establish a molecular model for the selection of the SHF gene regulatory network for cardiac septation.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pediatrics, Pathology, and Human Genetics, The University of Chicago, Chicago, Illinois, United States of America.

ABSTRACT
The Second Heart Field (SHF) has been implicated in several forms of congenital heart disease (CHD), including atrioventricular septal defects (AVSDs). Identifying the SHF gene regulatory networks required for atrioventricular septation is therefore an essential goal for understanding the molecular basis of AVSDs. We defined a SHF Hedgehog-dependent gene regulatory network using whole genome transcriptional profiling and GLI-chromatin interaction studies. The Forkhead box transcription factors Foxf1a and Foxf2 were identified as SHF Hedgehog targets. Compound haploinsufficiency for Foxf1a and Foxf2 caused atrioventricular septal defects, demonstrating the biological relevance of this regulatory network. We identified a Foxf1a cis-regulatory element that bound the Hedgehog transcriptional regulators GLI1 and GLI3 and the T-box transcription factor TBX5 in vivo. GLI1 and TBX5 synergistically activated transcription from this cis-regulatory element in vitro. This enhancer drove reproducible expression in vivo in the posterior SHF, the only region where Gli1 and Tbx5 expression overlaps. Our findings implicate Foxf genes in atrioventricular septation, describe the molecular underpinnings of the genetic interaction between Hedgehog signaling and Tbx5, and establish a molecular model for the selection of the SHF gene regulatory network for cardiac septation.

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Atrioventricular septal defects in Foxf1a+/−; Foxf2+/− compound heterozygote embryos at E14.5.Foxf1a+/−; Foxf2+/− embryos displayed atrial septal defects including absence of the dorsal mesenchymal protrusion (D, D′, black arrows). Compound heterozygotes also displayed expanded mesenchymal cap of primary atrial septum (red arrow) (D, D′). Wild-type (A, A′), Foxf1a+/− (B, B′), and Foxf2+/− embryos (C, C′) showed no atrial septal defects. P-values (Fisher's exact test): Foxf1a+/− (9 embryos) vs wild-type (4 embryos) = 1; Foxf2+/− (2 embryos) vs wild-type = 0.33; Foxf1a+/−; Foxf2+/− (3 embryos) vs wild-type = 0.03.
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pgen-1004604-g004: Atrioventricular septal defects in Foxf1a+/−; Foxf2+/− compound heterozygote embryos at E14.5.Foxf1a+/−; Foxf2+/− embryos displayed atrial septal defects including absence of the dorsal mesenchymal protrusion (D, D′, black arrows). Compound heterozygotes also displayed expanded mesenchymal cap of primary atrial septum (red arrow) (D, D′). Wild-type (A, A′), Foxf1a+/− (B, B′), and Foxf2+/− embryos (C, C′) showed no atrial septal defects. P-values (Fisher's exact test): Foxf1a+/− (9 embryos) vs wild-type (4 embryos) = 1; Foxf2+/− (2 embryos) vs wild-type = 0.33; Foxf1a+/−; Foxf2+/− (3 embryos) vs wild-type = 0.03.

Mentions: We hypothesized that Foxf1a and Foxf2 were required in a dosage sensitive manner for atrioventricular septation. We analyzed the cardiac anatomy of embryos from an intercross between Foxf1a+/− and Foxf2+/− at E14.5, when cardiac septation is normally complete. Foxf1a+/−; Foxf2+/− double-heterozygote embryos all exhibited atrioventricular septal defects (Figure 4D, D′ asterisk; p = 0.03). Primum-type atrial septal defects, characterized by absence of the dorsal mesenchymal protrusion, were observed in each case (Figure 4D, D′). Additionally, Foxf1a+/−; Foxf2+/− double-heterozygotes displayed larger than normal mesenchymal caps covering the primary atrial septum (Figure 4D′ arrow), an observation in keeping with the known redundant requirement for Foxf1a and Foxf2 in limiting mesenchymal growth in other contexts [32]. Atrial septal defects were never observed in Foxf1a+/− (Figure 4B, B′) or Foxf2+/− (Figure 4C, C′) single-heterozygotes or wildtype control littermate embryos (Figure 4A, A′). We concluded that Foxf1a and Foxf2 are redundantly required for atrioventricular septation.


Foxf genes integrate tbx5 and hedgehog pathways in the second heart field for cardiac septation.

Hoffmann AD, Yang XH, Burnicka-Turek O, Bosman JD, Ren X, Steimle JD, Vokes SA, McMahon AP, Kalinichenko VV, Moskowitz IP - PLoS Genet. (2014)

Atrioventricular septal defects in Foxf1a+/−; Foxf2+/− compound heterozygote embryos at E14.5.Foxf1a+/−; Foxf2+/− embryos displayed atrial septal defects including absence of the dorsal mesenchymal protrusion (D, D′, black arrows). Compound heterozygotes also displayed expanded mesenchymal cap of primary atrial septum (red arrow) (D, D′). Wild-type (A, A′), Foxf1a+/− (B, B′), and Foxf2+/− embryos (C, C′) showed no atrial septal defects. P-values (Fisher's exact test): Foxf1a+/− (9 embryos) vs wild-type (4 embryos) = 1; Foxf2+/− (2 embryos) vs wild-type = 0.33; Foxf1a+/−; Foxf2+/− (3 embryos) vs wild-type = 0.03.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214600&req=5

pgen-1004604-g004: Atrioventricular septal defects in Foxf1a+/−; Foxf2+/− compound heterozygote embryos at E14.5.Foxf1a+/−; Foxf2+/− embryos displayed atrial septal defects including absence of the dorsal mesenchymal protrusion (D, D′, black arrows). Compound heterozygotes also displayed expanded mesenchymal cap of primary atrial septum (red arrow) (D, D′). Wild-type (A, A′), Foxf1a+/− (B, B′), and Foxf2+/− embryos (C, C′) showed no atrial septal defects. P-values (Fisher's exact test): Foxf1a+/− (9 embryos) vs wild-type (4 embryos) = 1; Foxf2+/− (2 embryos) vs wild-type = 0.33; Foxf1a+/−; Foxf2+/− (3 embryos) vs wild-type = 0.03.
Mentions: We hypothesized that Foxf1a and Foxf2 were required in a dosage sensitive manner for atrioventricular septation. We analyzed the cardiac anatomy of embryos from an intercross between Foxf1a+/− and Foxf2+/− at E14.5, when cardiac septation is normally complete. Foxf1a+/−; Foxf2+/− double-heterozygote embryos all exhibited atrioventricular septal defects (Figure 4D, D′ asterisk; p = 0.03). Primum-type atrial septal defects, characterized by absence of the dorsal mesenchymal protrusion, were observed in each case (Figure 4D, D′). Additionally, Foxf1a+/−; Foxf2+/− double-heterozygotes displayed larger than normal mesenchymal caps covering the primary atrial septum (Figure 4D′ arrow), an observation in keeping with the known redundant requirement for Foxf1a and Foxf2 in limiting mesenchymal growth in other contexts [32]. Atrial septal defects were never observed in Foxf1a+/− (Figure 4B, B′) or Foxf2+/− (Figure 4C, C′) single-heterozygotes or wildtype control littermate embryos (Figure 4A, A′). We concluded that Foxf1a and Foxf2 are redundantly required for atrioventricular septation.

Bottom Line: GLI1 and TBX5 synergistically activated transcription from this cis-regulatory element in vitro.This enhancer drove reproducible expression in vivo in the posterior SHF, the only region where Gli1 and Tbx5 expression overlaps.Our findings implicate Foxf genes in atrioventricular septation, describe the molecular underpinnings of the genetic interaction between Hedgehog signaling and Tbx5, and establish a molecular model for the selection of the SHF gene regulatory network for cardiac septation.

View Article: PubMed Central - PubMed

Affiliation: Departments of Pediatrics, Pathology, and Human Genetics, The University of Chicago, Chicago, Illinois, United States of America.

ABSTRACT
The Second Heart Field (SHF) has been implicated in several forms of congenital heart disease (CHD), including atrioventricular septal defects (AVSDs). Identifying the SHF gene regulatory networks required for atrioventricular septation is therefore an essential goal for understanding the molecular basis of AVSDs. We defined a SHF Hedgehog-dependent gene regulatory network using whole genome transcriptional profiling and GLI-chromatin interaction studies. The Forkhead box transcription factors Foxf1a and Foxf2 were identified as SHF Hedgehog targets. Compound haploinsufficiency for Foxf1a and Foxf2 caused atrioventricular septal defects, demonstrating the biological relevance of this regulatory network. We identified a Foxf1a cis-regulatory element that bound the Hedgehog transcriptional regulators GLI1 and GLI3 and the T-box transcription factor TBX5 in vivo. GLI1 and TBX5 synergistically activated transcription from this cis-regulatory element in vitro. This enhancer drove reproducible expression in vivo in the posterior SHF, the only region where Gli1 and Tbx5 expression overlaps. Our findings implicate Foxf genes in atrioventricular septation, describe the molecular underpinnings of the genetic interaction between Hedgehog signaling and Tbx5, and establish a molecular model for the selection of the SHF gene regulatory network for cardiac septation.

Show MeSH
Related in: MedlinePlus